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The Impacts Of Ultra
Yiolet B Radiation On
Biological Systems:
A Study Relate
Stratospheric Ozone
Depletion
Submitted To:
The Stratospheric Impact Research
and Assessment Program (SIKA)
The U.S. Environmental Protection Agency
Washington, D.G. 2O6O4
Volume I
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DISCLAIMER
THIS REPORT HAS NOT BEEN REVIEWED FOR APPROVAL BY THE
AGENCY AND HENCE ITS CONTENTS DO NOT REPRESENT THE VIEWS AND
POLICIES OF THE U.S. ENVIRONMENTAL PROTECTION AGENCY, NOR DOES
MENTION OF TRADE NAMES OR COMMERCIAL PRODUCTS CONSTITUTE ENDORSEMENT
OR RECOMMENDATION FOR USE.
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CONTENTS
SIRA FILE #
VOLUME I
1. Study Of Increase In Skin Cancer As A Function
Of Time And Age And Changing Stratospheric Ozone:
Need For Careful Measure Of The Ultraviolet Dose ............. # 132.11
2. The Influence Of Age, Year Of Birth, And Date On
Mortality From Malignant Melanoma In The Populations
Of England & Wales, Canada And The White Population
Of The United States # 132.31
3. Non Melanoma Skin Cancer Surveys In The United States
- An Environmental Epidemiologic Project ...................... # 142.11
4.
5. Biological Effects Of Ultraviolet Radiation On Plant
Growth And Function ,. # 142.21
6. Effects Of UV-B Radiation On Selected Leaf Pathogenic
Fungi And On Disease Severity # 142.21g
7. The Effect Of Ultraviolet (UV-B) Radiation On Englemarm
Spruce And Lodgepole Pine Seedlings // 142.22
VOLUME II
8. UV-B Biological And Climate Effects Research # 142.23
9. Ultraviolet Effects Of Physiological Activities Of
Blud-Green Algage '... # 142.24
10. Impact Of Solar UV-B Radiation On Qfops And Crop
Canopies ....*..ซ # 142.25
11. High Altitude Studies Of Natural, Supplemental
And Deletion Of UV~B On Vegetables And Wheat ................. # 142.26
VOLUME III
12. UV-B Radiation Effects On Photosynthesis And
Plant Growth tf 142.27
13. Influence Of Broad Band UV-B On Physiology And
Behavior Of Beneficial And Harmful Insects // 142.28
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SIRA FILE
14. A Study Of The Effects Of Increased UV-B
Irradiation On Environmental Dissipation Of
Agricultural Chemicals ....... # 142 v. 29
15. Biological Effects Of Ultraviolet Radiation
On Plant Growth And Development In Florist And
Nursery Crops .# 142.210
16. Biological Effects Of Ultraviolet Radiation On Cattle:
Bovine Ocular Squamous Cell Carcinoma # 142.211
17. Radiation Sources And Related Environmental Control
For Biological And Climatic Effects UV Research (EAGER) # 142.212
18. Instrumentation For Measuring Irradiance In The
UV-B Region # 142.213
19o Annual Report To EPA, Bacer Program For
Fiscal Year 1978 ....... # 142.34
20. Penetration Of UV-B Into Natural Waters # 142.36'
21. Higher Plant Responses To Elevated Ultraviolet:
Irradiance // 142.4.1
22. Assessment Of The Impact Of Increased Solar
Ultraviolet Radiation Upon Marine Ecosystems // 142.42
23. UV-B Instrumentation Development ............................ # 142.51
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STUDY OF INCREASE IN SKIN CANCER
AS A FUNCTION OF TIME AND AGE AND CHANGING STRATOSPHERIC OZONE:
NEED FOR CAREFUL MEASURE OF THE ULTRAVIOLET DOSE
By June Morita and Elizabeth L.'Scott
Statistical Laboratory
University of California, Berkeley
The work which we are reporting is part of a large cooperative study
of the Panel to Review Statistics on Skin Cancer of the Committee on
National Statistics of the National Research Council, of an informal
Skin Cancer Workshop at the University of California, Berkeley, and of
members of the staff of the National Center for Health Statistics, and
of the National Cancer Institute. We have been' studying both melanoma
and nonmelanoina skin cancer as well as actinic skin damage. These are
responses to ultraviolet radiation, perhaps associated in some cases with
certain chemicals. They are rather different but each is important and
the studies support each other. We are estimating the increase in skin
cancer for a specified scenario of the change in ultraviolet radiation, as
a function of time, age, sex, location, and other predictor variables.
Note that a scenario about change in ultraviolet radiation implies that
an individual at a given locality will receive different doses as time
changes. Actually the present dose is not constant; there is diurnal
change, seasonal change, year to year change (influenced by changes in the
pollution of the earth's atmosphere and stratosphere as well as changes on
the sun), and also local changes within our atmosphere. The individuals at
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a given locality have different life styles, different sensitivity spectrum
of the skin, and so forth, all of which affect the dose received.
C#i1iซ'ซป
The effect of,new pollutants entering the stratosphere will be to change
the proportion of ozone and thus the amount of ultraviolet radiation that
passes through to the earth's surface. Because the vertical mixing in the
atmosphere is very slow and because cancer has a long latent period, any
change in ultraviolet radiation will have a long-time effect on skin cancer.
We will need to consider ultraviolet flux as a function of time as well as
of locality, life style, and other variables.
Observational data about skin cancer are sparse and inaccurate, due
partly to the gross under-reporting of nonmelanoma skin cancer. We need
to use many different sources of data from independent studies, employing
individual sets to obtain reinforced conclusions.
Our method of study is to estimate the increase in incidence, preva-
lence or mortality, as the case may be, for each age and sex category,
as we consider one locality after another with increasing flux. In addition
to our difficulties with the uncertainties in the skin cancer data, we also
have difficulties with uncertainties in the dose of ultraviolet radiation
received by an individual at a specified locality. In the first place,
we do not have measurements of the ultraviolet flux received at each
locality during any short period of time. In the second place, we do not
have measurements of the variability in dosage from one person to another
at the same locality or from one time to another for the same person. We
need to know the average dosage at each locality for each short time period,
and we also need to know how this dose varies according to the life style
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of the individual,'according to his skin sensitivity7 his year of birth,
and so forth and so forth. This information is not availablej we have
made use of estimates.
We estimate the average ultraviolet flux dosage, for each, locality
with the help of extensive computations made for us by S.V, Venkateswaran
and his colleagues, D.E. St. John and N. Sundaranaraman.
As shown in the schematic drawing in '.Figure 1", . ' -
. the ultraviolet radiation received by an individual ฑs the product of, the
solar irradiance in the ultraviolet times the transmittanee.through the
stratospheric ozone layer and the earth's atmosrphere (as computed for us
by Venkateswaran using ozone measures) times the transmittanee through
the layer of clouds and murk at the' earth's surface times the transmittanee
through the individual's clothing and other protection and finally into
his skin. We have labelled this product ''sensed -flux," Each of the factors
entering the product sensed flux is uncertain, particularly the estimated
. sensitivity of the skin as a function of wavelength. Following Setlow, v?e
have used the relative sensitivity, as a function of wavelength, in terms
of damage to DNA by radiation. It is important to have a check on the
calculated values, of. . . . .."'''
accuracy of the/flux of ultraviolet radiation at the'earth's surface and
also the amount entering the individual's skin. Ultraviolation radiation
meters, in particular, the Berger meter, provide a partial check. We are
now engaged in a comparison of the results we deduced from the Venkateswaran
computations, with those, obtained by. the Berger meter.
A further complication is that the ultraviolet radiation'dose varies
markedly from one individual to another in the same locality, depending
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OZONE
LAYER
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FIGURE "L. Schematic dirawing of path of ultraviolet: radiation through the
stratosphere and atmosphere of the earth.
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on life style, protective clothing, and so forth.. This means that some
individuals will be subject to higher risk than others, It also intro-
duces a statistical complication. The standard methods of estimating the
relationship between skin cancer raLes and ultraviolet flux will be biased,
underestimating the slope. To correct for the bias du6 to the variation
in ultraviolet dose from one individual to another at the same locality,
we need data on the variability. Direct measurements are hot yet available,
but estimates can be made from the surveys of Urbach for the two localities.
Philadelphia, Pennsylvania, and Galway, Ireland. Further information is
available from the exposure data of the Health and Nutrition Examination
Survey of the National Center for Health Statistics. However, these are
only estimates a.nd they are not scaled. We need actual measurements from
personal dosimeters on individuals in different localities, some in the
South and some in the North, in different age and sex categories, with
various skin sensitivities,, various occupations, and so forth.
We want to make a strong plea for personal dosimeter data, collected
on a regular basis on each of many individuals for.successive short periods
of time, preferably daily, so as to obtain estimates'of the dose under
v .
specified employment a.nd recreation regimes, and. as a function of micro-
climate,, age, sex, etc. We have already stated the'importance of verifying
the estimated avex'age flux at each, locality, and the importance of estimating
how this flux dosage varies in time and varies from one individual to
another at the same locality and time. We also should emphasize the dose-
rate problem. There is clinical evidence that persons who expose themselves
to a large dose of ultraviolet radiation, even for a relatively short
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period of. time, are much more likely to develop actinic skin damage and
even malignant melanoma than persons who receive the same total dose in
small sub-doses distributed over a long period of time, This, effect is
very striking, for example, in Scandinavians who spend a, short vacation
in the Mediterranean area, but it is also noticeable in office workers
who enjoy an outdoor weekend. Personal dosimeter observations are needed
to establish, the dose-rate relationships for different types of individuals,
and to compaje the estimated relationships with those predicated from
studies of laboratory mice.
Our study comparing the ultraviolet flux estimates based on the compu-
tations of Venka.te3wa.rain a.nd the. observations provided by the Berger meter
is still preliminary-, Therefore., the report'we are. presenting today is
an interim report.
First, we need to be .clear.!about what, we are comparing. On the one
hand, we have sensed flux, an estimate of what affects the cells of the skins-
computed as the product of many factors, as described above. This is to be
compared with the measurements from the Berger meter t as provided to us by
W. N. Haas and his colleague G. Cotton at the Air Resources Laboratories,
National Oceanic and Atmospheric Administration.
Speaking roughly, the Robertson-Berger meter integrates over the radi-
ation actually observed at a specified locality during each' time ; . .
interval. The integration is weighted so that the result approximates what
the cells of the skin would receive if the sensitivity spectrum of the skin
were a standard erythema (sunburn) spectrum. For each place where a Berger
meter is located, we have observations for each half-hour over the recorded
life of the meter there, usually from January 1, 1974 through October 30, 1976,
with .later observations to become available shortly. The sensed flux and the
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Berger meter flux are not strictly comparable. The sensed flux is a
product of extensive theoretical computations based on long-range ozone
observations, adjusted for altitude and for long-range cloudiness, .for skin
absorption and for the DNA-damage spectrum. It is available for each of
74 localities in the United States on a per month basis. The Berger
meter flux is strictly observational, subject to the vagaries of real clouds
and other absorbing material at its real altitude, but with the observations
weighted in an approximate fashion designed to correspond to the erythema
spectrum rather than-DNA-damage spectrum. Nevertheless, it is of interest
to see how well the two sets agree and to examine any systematic differences.
In particular, are there systematic differences in the higher flux (southern)
. *
localities that are different from those in the lower flux (nortbern)
localities?
The short time-period observations from the Berger meter were combined
into daily observations which were then combined to obtain averages over the '
month when the meter was in operation. Missing observations were interpo-
lated or were supplied from the auxilliary record?" at each locality. For
purposes of comparison with sensed flux, averages were taken over the years
the meter was in operation. The results are illustrated in Figure 2 for the
meter in Minneapolis. The triangles connected by solid-lines are the Berger
meter readings for a particular month. The crosses connected by dashed' lines
are monthly averages over the typically three years the meter was in operation
drawn again for each year to illustrate the variability from year to year
contrasted with the average over only three years. For example, the spring
of 1976 tended to have higher flux than the three-year average.
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MONTH
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7/76
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A Berger Meter Flux
X AvQ.(Mean)For Each Month
FIGURE 2. Comparison of Berger meter readings for each month (triangles
connected by solid lines) and monthly averages over three years
(crosses connected by dashed lines). > . . .
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These Berger meter monthly averages are drawn again on Figure 3
along with the monthly sensed flux values (as circles connected by solid
lines). Since the units are arbitrary for .both the Berger meter values and
the sensed flux (because the scale of the action spectrum is unknown in
both cases, and is arbitrarily set equal to unity at one wavelength), we
have adjusted the plot in Figure 3 so that distances from minimum to maximum
agree.
We have studied the relation between the monthly Berger meter measure-
ments and the corresponding monthly sensed flux in those localities where
< both are available to us: Albuquerque, El Paso, Des Moines, Oakland,
Minneapolis, PHiladelphia and Ft. Worth (except that we made an error in
computation on the Ft. Worth data). The localities group nicely into pairs
by climate: clear (Albuquerque and El Paso), mixed (Des Moines and Oakland),
and cloudy (Minneapolis and Philadelphia).
In Figure 4 is shown a direct comparison between the sensed flux and
the Berger meter flux for each month of the year 1975 for the two locations,
El Paso and Albuquerque. We have used different symbols for the six months
January to June and the remaining six months July to December. For the first
v
year in which the Berger meters were in operation the relationship between
Berger meter and latitude measures was systematically different for the first
half of the year and the second half. This effect persists when comparing
sense flux with Berger meter in each of the years 1974, 1975, 1976. For the
clear stations (which are the southern stations in our comparisons), the
relation for the first half of the year tends to be higher than that of the
second. This is also true for the cloudy stations (which are further north),
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. .. hy dashed lines) with the monthly sensed flux values (circles
connected hy solid lines).
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MONTHLY RESIDUALS
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MONTHLY RESIDUALS
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FIGURE 5. Direct comparison between the sensed flux and the Berger meter
flux for each month of the year 1975 for the two locations,,
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as shown in Figure 6. On the other hand, the early months tend to be lower
for the two mixed climate stations which we have studied, Oakland and
Des Moines. That is, if one were to trace the points in the scatter plot
for El Paso (Figure A), starting with January, then to February, and so
forth to December, one would find oneself tracing an ellipse. Thus it seems
that for the first half of the year the observed Berger meter flux values
are consistently higher than the corresponding calculated sensed flux values,
while for the second half of the year, the Berger meter values are consistently
lower than the corresponding sensed flux values. Seasonal deviations were
already noticed by Urbach in the 1974 data.
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MONTHLY RESIDUALS
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MONTH
FIGURE 6. Direct comparison between the sensed flux and the Berger meter
. flux for each month of the year 1975 for the two locations,,
s and Phi
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Both of these stations happen to be at middle latitude. We should point
out that the cloudiness we are using refers to the middle of the day in
July, the period of highest ultraviolet radiation intensity. In spite of
the suggestion of the systematic differences between the two sets of obser-
vations, which could be explained by inadequacies in the factor correcting
'for cloudiness in the computations of the sensed flux, the relationship
between the two measures is very reasonable and nearly linear. (We could
also explain this discrepancy by prolonged periods of unusual cloudiness in
the years under study.)
We have started to make a more detailed examination of the residuals.
The results are shown in Figures 7, 8 and 9 for the eight localities under
study, again paired according to climate. In the plots, the least-square
line relating the Berger meter flux measures to the sensed flux has been
eliminated and only the residuals from the line are plotted. Our original
plan had been to compare the residuals with cloudiness data for each month
shown. We have not extracted the cloudiness data so that part of the study
is unfinished. The plots of the residuals do give an impression of smooth
changes with time and it is of interest to determine whether these changes
reflect long periods of unusual weather.
. The last three Figures again show the residuals from a straight-line
relationship between the two measures of flux, but the residuals have -.been'.,'.
smoothed by Tukey's 3R or Running Medians-of three\ .repeated .until.convergence.
The.suggestion of systematic changes does persist but the evaluation of'the
contribution of unusual weather has still not been done.
We are continuing our studies comparing Berger meter flux with the sense
flux. As stated above, we are searching for systematic differences between
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MONTHLY FLUX VALUES 1975
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FIGURE 7. Monthly residuals from the line relating the Berger meter flux
measures to the sensed flux for El Paso and Albuquerque".
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FIGURE 9. Monthly residuals from the line relating the Berger meter flux
measures to the sensed flux for Minneapolis and Philadelphia.
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FIGURE 10. Running medians of monthly residuals for El Paso and Albuquerque.
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FIGURE 11. Running medians of monthly residuals for Oakland and Des Moines.
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the two systems which cannot be explained by prolonged periods of. unusual
weather. When observations are available from the Berger meters over a,
long period, the effects of irregularities of weather will be minimal.
At the present time, with less than three years of data at hand, we can
only estimate the effects of unusual weather and we need further meteorolog-
ical data to do even this. In order to check the adequacy of the cloudiness
corrections we have been using, we need many more than three years of com-
parison. We suspect that our cloudiness corrections are too simplistic.
The means of studying the adequacy of cloudiness corrections lies in the
direct comparison of Berger meter readings with the sensed flux data, com-
bined with meteorological observations on cloud cover, and extended over a
reasonably long period of time.
We have been troubled by the cloud corrections we, and everyone else
who makes any correction at all, have been using because we noted systematic
grouping of the residuals in age-adjusted mortality from malignant melanoma
considered as a function of flux. Localities with low cloudiness tend to
have higher than predicted mortality; localities with more cloudiness tend
to have lower than predicted mortality even when corrections for microclimate
are included. Even though residuals in our more recent studies do not show
any such pronounced effects, we realize that we should employ better means
for considering the attenuation by high and low clouds and other absorbing
materials. A careful study of the Berger meter readings over a period of
years, combined with a corresponding series of meteorological observations,
should provide the information we need.
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LEGENDS FOR FIGURES
FIGURE 1. Schematic drawing of path of ultraviolet radiation through the
stratosphere and atmosphere of the earth.
FIGURE 2. Comparison of Berger meter readings for each month (triangles
connected by solid lines) and monthly averages over three years
(crosses connected by dashed lines).
FIGURE 3. Comparison of Berger meter monthly averages (crosses connected
.ซ. hy dashed lines) with the monthly sensed flux values (circles
connected by solid lines).
FIGURE 4. Direct comparison between the sensed flux and the Berger
meter flux for each month of the year 1975 for the two locations,
El Paso and Albuquerque.
FIGURE 5. Direct comparison between the sensed flux and the Berger meter
v
flux for each month of the year 1975 for the two locations,
Oakland and Des Moines.
FIGURE 6. Direct comparison between the sensed flux and the Berger meter
. flux for each month of the year 1975 for the two locations,
Minneapolis and Philadelphia.
FIGURE 7. Monthly residuals from the line relating the Berger meter flux
measures to the sensed flux for El Paso and Albuquerque.
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FIGURE 8. Monthly residuals from the line relating the Berger meter flux
measures to tVie sensed flux for Oakland and Des Moines.
FIGURE 9. Monthly residuals from the line relating the Berger meter flux
measures to the sensed flux for Minneapolis and Philadelphia.
FIGURE 10. Running medians of monthly residuals for El Paso and Albuquerque.
FIGURE 11. Running medians of monthly residuals for Oakland and Des Moines.
FIGURE 12. Running medians of monthly residuals for Minneapolis and
Philadelphia.
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. 3t
THE INFLUENCE OF AGE, YEAR OF BIRTH, AM) DATE ON MORTALITY FROf!
MALIGNANT MELANOMA IN THE POPULATIONS OF ENGLAND & WALES, CANADA
AND THE WHITE POPULATION OF THE UNITED STATES
John A. H. Lee (1)
Gerald R. Petersen (2)
Richard G. Stevens (3)
Kajo Vesanen (1)
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Page 2
FOOTNOTES FOP. TITLE PAGE
Received for publication :
(1) From the Department of Epidemiology SC-36, University of
Washington Seattle, WA 98195. Address reprint requests to Dr. Lee.
K. Vesanen is witli the Department of Biostatistics at the University
of Washington
(2) San Jose State University, San Jose, CA 95192
(3) The Fox Chase Cancer Center, Institute for Cancer Research,
7701 Burholme Ave., Philadelphia, PA 19111
The study was supported by Environmental Protection Agency
Research Grant R805363010 from the Office of Health and Ecological
Effects.
A preliminary summary of these findings was presented at the
1977 meeting of the Environmental Protection Agency, Program
Planning and Review Workshop for the Biological and Climatic Effects
Research Program.
We are indebted to Mr. Larry C. Clark for data tapes.
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ABSTRACT
The age-adjusted death rates from malignant melanoma of the
skin have increased from 1951 to 1975 by about 3% per year in the
populations of England & Wales and Canada, and in the white
population of the U.S. This is due to large increases in risk of
successively later born cohorts. Any effects of earlier diagnosis
or improved treatment within the period 1951-75 have been
sufficiently steady to to fail to alter these trends. The slope of
the log rates with log age is about 3.5. Projections of rates for
at least the next decade can be made with some confidence, and
provide a basis for evaluating control measures.
MALIGNANT MELANOMA : COHORTS : AGE DISTRIBUTION : CANCER
CONTROL : INTERNATIONAL COMPARISONS
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Page
INTRODUCTION
The incidence of malignant melanoma has been rising rapidly in
many white populations in recent years (e.g. 1-6). The death rate
has also been rising sharply in the populations of England ฃ Wales
(7), and Canada (8). Mortality data for malignant melanoma for the
U.S. have been incompletely published for the years 1950-67 (9) and
for later years without specification by age. There are indications
from these data that mortality is also rising in the U.S.
population.
A major component of the causation of this rising mortality is
a systematic increase of risk of successively later born cohorts
(7,8,10). This produces a large difference between the age
distribution of the mortality in a population, and the age
distribution within each cohort (11). The prognosis of malignant
melanoma has improved (12), probably due to earlier diagnosis. Such
a change may also affect the observed age distribution of a disease
(13).
Mortality from malignarit- melanoma in white populations is
inversely related to latitude of residence (14,15), an exception
being the inhabitants of Sweden who have more melanomas than would
be expected from their latitude of residence (16). Long duration of
residence in Israel of persons of European origin leads to an
increased incidence of melanoma (17,18). The incidence of malignant
melanoma per unit skin area is greater on exposed than on unexposed
sites (1,19). The intensity of ultra-violet light at ground level
will rise due to the expected depletion of the stratospheric ozone
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Page 5
layer (20). Although the action spectrum of neoplastic change in
the human melanocyte is unknown, there are reasonable, grounds for
assuming that the ultra-violet component is important. At a time
when administrative measures, such as the control of refrigerating
agents, are under consideration to protect the ozone layer, it is
important to have projections of the future trends of mortality from
malignant melanoma. These can provide a base line for the
evaluation of control measures.
In this paper we report the U.S. mortality data for whites from
1951 through 1975, and compare the rates and their trends and age
distributions with those from Canada and England & Wales.
DATA AND METHODS
The numbers of deaths from malignant melanoma by age and sex
for England & Wales and for Canada, and the population data for all
three countries were obtained from the standard publications of the
respective offices of vital statistics. The U.S. melanoma mortality
data for 1950-67 were obtained from special tabulations from the
V
Department of Epidemiology, University of North Carolina, using data
originating with the National Center for Health Statistics. Data
for 1968-76 were obtained from special tabulations from the National
Center for Health Statistics, who take no responsibility for the
analyses that we have made.
Rates and rates of change were calculated as shown in the Table
descriptions. As a check that there were no serious interactions in
the data matrices, multivariate methods were also used to estimate
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Page 6
the parameters. The results are shown in the Tables.
RESULTS
The age adjusted death rates from malignant melanoma of skin
(ICD172) for England & Wales, Canada, and the United States white
population by 5-year periods 1951-55 through 1971-75 are shown in
table 1. The rates are rising most rapidly in Canada (Table 2), but
the changes are marked in all three countries. The least
proportionate increase is in U.S. females.
The prognosis of malignant melanoma has been improving (12), so
that each cohort will suffer a smaller increment of mortality per
unit time in the later time periods than in the earlier. However,
the mean slopes of rate against age (Table 3) show no consistent
changes with time. A constant rate of improvement with time will
not change the rates of change observed, and a steady improvement is
probably what is occurring in the diagnosis and treatment of
melanoma.
The rates of change with increasing age appear to be similar at
i
all ages (Table 4). They are lower in females than in males,
probably reflecting the better female prognosis, and are higher in
Canada than in the U.S. or England & Wales. Estimates using
multivariate techniques (Table 4) give similar results to the simple
means, and indicate that interactions between parameters are not
large.
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Page 7
The increase of mortality with increasing age is greater than
increase of incidence with age reported from Connecticut (11).
Case-fatality increases with age (12), which could account for the
difference. The differences between the cohorts in their mortality
rates are large and consistent (Table 5 and the Figure as an
example), particularly when it is considered that neighboring
cohorts are only separated by 5 years in date of birth. The decline
in the 1931 female cohort compared to the 1936 is consistent and
interesting.
Projections of mortality rates are an important guide to the
effectiveness of administrative or therapeutic measures. For
malignant melanoma rates they are likely to be reliable as the rates
of change with age do not show any sharp changes with time, and the
current behavior of the cohorts that will provide the bulk of the
deaths in the next decade is well established. Projected rates
derived from the present matrices of data, using the rates for
1971-75, and the rates of change for 1971,(13), are shown in
Table 1.
*,
DISCUSSION
The driving force behind the current increase in the mortality
from malignant melanoma is a systematic increase in risk with
successively later years of birth. It is difficult to imagine a
reasonable mechanism involving better diagnosis or reporting that
could produce these regular cohort differences, particular in the
light of the parallel increase in incidence rates . Why people born
five years later should go through life with a substantially
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Page 8
increased risk of dying from malignant melanoma compared with their
elder peers in the same population is unknown. Whether there are
systematic differences in life style that are distributed in this
way, or whether there is some deeper biologic mechanism at work,
will require, in the first instance, a series of studies in clinical
epidemiology.
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Page 9
Table I - Death Rates* and Projected** Death Rates from Malignant
Melanoma of Skin by Sex and Time Period: England & Wales, Canada,
and United States whites.
Time Period 1951- 1956- 1961- 1966- 1971- 1976- 1981-
55 60 65 70 75 80 85
MALE
England&Wales 6.8 7.4 8.6 10.6 11.8 : 13.4 15.2
Canada 7.1 10.1 11.7 13.5 15.8 : 18.9 '22.1
U.S.white 14.5 16.6 19.9 22.9 26.3 : 28.7 33.5
FEMALE
England&Wales 7.2 8.5 9.8 10.8 13.3 : 16.3 19.8
Canada 6.0 8.2 10.1 11.5 12.3 : 13.0 14.1
U.S.white 11.2 12.6 13.8 15.4 16.5 : 17.7 18.8
*In this, and the subsequent tables, rates are given per
million per year, age adjusted using the UICC standard European
population (21).
** Rates to the right of the colons are derived from each national
set of age and sex specific rates 1971-75, and the sets of cohort
slopes for 1971. These are the age-specific increments of the log
rates divided by the increments of the log ages. Rates for new cohorts
entering the projection at age 15-19 were given the 1971-75 values.
The method is discussed in (13).
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Page 10
Table 2 - Mean Annual Percentage Increase in Malignant Melanoma
Death Rate*, England & Wales, Canada, and U.S. whites by Sex.
Male %
England & Wales 2.9
Canada 3.5
U.S. whites 3.0
Female
England & Wales 2.9
Canada 3.3
U.S. whites 1.9
* The slope of the line fitted to the national period rates in
Table 1 as a % of the national means.
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Page 11
Table 3 - Mean* Cohort Slopes by Mid-year of Observation,
Malignant Melanoma Mortality Rates, England & Wales; Canada;
and United States whites.
MALE FEMALE
Mid-Year England England
& Wales Canada U.S.w & Wales Canada U.S.w
1956
1961
1966
1971
3.59
3.41
4.43
3.48
6.34
2.98
3.92
3.94
3.54
3.87
3.76
3.66
3.82
3.43
2.64
3.91
5.38
5.01
3.56
2.69
3.21
2.92
3.25
2.96
^Arithmetic means of the age-specific increments of the log rates
divided by the increments of log ages (13). The data for 1956
are the means of the age-specific slopes between the years
1951-55 and 1956-60.
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Page 12
Table A - Mean age-specific Cohort Slopes* for Death Rates from
Malignant Melanoma by Sex: England & Wales, Canada, and United
States whites, 1951-55 through 1971-75.
England
&
AGE
20
25
30
35
40
45
50
55
60
65
70
75
80
85
Mean
20-85
Estimates
Wales
7.01
3.79
2.05
3.44
1.88
3.09
4.90
2.24
3.94
4.30
4.48
2.31
7.24
3.32
3.86
fron least
squares
**
3.32
Estimates
from maximum
likeli-
hood*** 3.58
MALE
Canada
6.37
2.36
3.71
1.81
4.14
4.56
3.83
3.49
3.27
7.51
4.43
4.28
2.21
16.33
4.88
5.06
3.67
U.S.
Whites
4,
3.
3.
,83
,44
31
2.65
2.27
3.33
3.51
,80
,86
.00
,85
.93
59
6.08
3.89
3.27
3.4!
England
& Wales
4.66
3.37
2.49
3.74
2.89
2.91
2.91
2.13
2.45
4.60
4.89
3.61
3.76
4.54
FEMALE
Canada
4.21
4.52
2.71
3.69
4.23
0.42
3.05
4.74
3.06
6.28
4.70
3.33
9.85
7.49
3.50
3.31
3.19
4.45
4.68
3.34
U.S.
Whites
4.27
3.82
2.57
2.
2.
2.
2.
2.
2.
3.
,63
,12
,50
,10
,69
,95
,48
3.59
4.79
3.90
6.72
3.44
3.15
2.93
*Cohort slopes as described in Table 2, arithmetic means of these
for successive time periods 1951-55 to 1956-60; 1956-60 to 1961-65
etc.
**Slopes of lines fitted to the age parameters for each data set (22).
***Slopes of lines fitted to the age parameters for each data set (23).
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Page 13
Table 5 - Mean Percentage Increase of Death Rates from Malignant
Melanoma between successive Birth Cohorts over the same Age Range
Cohorts
Age
Range
Mean % change
between cohorts
England
& Wales
1936/1931 20-39
1931/1926 25-44
1926/1921 30-49
1921/1916 35-54
1916/1911 40-59
1911/1906 45-64
1906/1901 50-69
1901/1896 55-74
1896/1891 60-79
1891/1886 65-84
1886/1881 70+
6
18
27
6
30
31
15
26
12
13
- 12
MALE
Canada
26
23
9
28
27
22
29
4
25
23
16
U.S.
whites
1
10
19
16
24
17
21
19
12
21
11
England
& Wales
- 6
18
8
38
21
40
22
4
9
18
6
FEMALE
Canada U.S.
wh i t e s
1
13
20
15
27
13
16
7
52
1
17
- 5
10
18
11
12
10
13
13
8
5
1
15.6% 21.1% 15.6% 16.2% 16.4% 8.7%
Mean of age-specific inter-cohort ratios expressed as a percentage.
-------
Page 14
150
125
100
50-
I
a
25-
10-
5-
2-
'46
15
\ 25 i 35 I 45 i 55 16
20 30 40 50 60
Figure: Cohort Diagram of Mortality Rates from Malignant Melanoma
in U.S. white Males. 1951-75
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Page 15
REFERENCES
1. Elwood JH, Lee JAM: Recent Data on the Epidemiology of
Malignant Melanoma. Sera in Oncology 2:149-154,1975
2. Lee JAH: The current rapid increase in incidence and mortality
from Malignant Melanoma in developed Societies. Pigment Cell
v.2 ed. Riley V pps.414-420 Basel Karger 1976
3. Magnus K: Incidence of Malignant Melanoma of the Skin in the
Five Nordic Countries: Significance of Solar Radiation. Int
J Cancer 20:477-485,1977
4. Teppo L,Pakkanen M, Hakulineri T: Sunlight as a Risk Fetor of
Malignant Melanoma of the Skin. Cancer 41:2018-2027,1978
5. Malec E, Eklund C: The Changing Incidence of Malignant Melanoma
of the Skin in Sweden, 1959-68. Scand J Plast Reconstr Surg
12:19-27,1978
6- Soodalter-Toman DL, West DW, Derrick LR: Epidemiology of
Malignant Melanoma in Utah 1966-76. In press
7. Lee JAM, Carter AP:Secular trends of Mortality from Malignant
Melanoma. J Natl Cancer Inst 45:91-97,1970
8. Elwood JM, Lee JAH:Trends in Mortality from Primary Tumours
of Skin in Canada: J Canad Med Assoc 110:913-915,1974
9. Burbank F: Patterns in Cancer Mortality in the United States:
1950-67. Natl. Cancer Inst Mono 33 Washington USDHEW
10. Gordon T, Crittenden M, Haenszel K: End Results and Mortality
Trends in Cancer II Cancer Mortality Trends in the United
States. Natl Cancer Inst Mono 6 pps 133-350 Washington USDHEW
11. Lee JAH: Letter. Am J Epidera 107:259-260,1978
12. End Results Section: Cancer Patient Survival No.5 pps 223-227
Washington USDHEW
13. Stevens RG, Lee JAH: Tuberculosis Generation Effects and
Chemotherapy. Am J Epidera 107:120-126,1978
14. Lancaster HO: Some Geographical Aspects of the Mortality
from Malanorna in Europeans. Med J Aust 1:1082-1087,1956
15. Elwood JM, Lee JA1I, Walter SD, Mo T, Green AES: Relationship
of Melanoma and Other Skin Cancer Mortality to Latitude
and Ultraviolet Radiation in the United States and Canada.
Int. J. Epidera 3:325-332,1974
-------
Page 16
16. Lee JAK, Issenborg HJ: A Comparison between England and Males
and Sweden in the Incidence and Mortality of Malignant Skin
Tumours. Brit J Cancer 26:59-66,1972
17. Movshovitz M, .Modan B: Role of Sun Exposure in the Etiology
of Malignant Melanoma: Epidemiological Inference. J Hatl
Cancer Inst 51:777-779
18. Anaise D, Steinitz R, Ben Hur N: Solar Radiation: A Possible
Etiological Factor in Malignant Melanoma in Israel.
A Retrospective Study (1960-72). Cancer 42:299-304
19. Committee on the Impacts of Stratospheric Change: llalocarbons:
Environmental Effects of Chloroflouromethane Release, pps
101-106 Washington National Academy of Sciences
20. Committee on the. Impacts of Stratospheric Change: Response to
the Ozone Protection Sections of the Clean Air Act Amemdments
of 1977: An Interim Report. Washington National Academy of
Sciences
21. Doll R, Cook P: Summarizing Indices for Comp-.rison of Cancer
Incidence Data. Int J Cancer 2:269-279,1967
22. Barrett JC: Age, time and cohort factors in mortality from
cancer of the cervix. J Hyg Camb 71:253-259,1973
23. Breslou ME, Day NE: Indirect Standardization and
Multiplicative Models for Rates with Reference to the Age
Adjustment of Cancer Incidence and Relative Frequency Data.
J Chron Dis 28:289-303,1975
-------
NCI - Human Health Effects Research Project
for the BACER Program
Project Title: Non Melanoma sk,in cancer surveys in the United States
An Environmental Epidemiologic Project
Interim Report - As of December -IS, 1977
TYPE OF CONTRACT: EPA/NCI Cooperative Interagency Agreement
Amount of Funding Provided by EPA (for the BACER Program): $840,000
Survey Period - One year, beginning June 1, 1977
Funding Period - One year's funding applied through
September 30, 1978
Key Personnel:
NCI: Joseph Scotto, Senior Health Services Officer and
Thomas Fears, Ph.D., FieTd Studies and Statistics Program,
V Division of Cancer Cause and Prevention
EPA: Herbert Wiser, Ph.D. and Alphonse Forziati, Ph.D.
Research and Development
-------
Summary
c
In response to the BACER program's urgent need for more epidemiologi -1
data on the possible human health effects of ozone depletion, the Natioru;
Cancer Institute is presently conducting simultaneous surveys on the annual
*
incidence of r.onmelanoma skin cancers in various locations in the United
States. All new cases of basal cell carcinoma and squamous cell carcinoma,
diagnosed as of June 1, 1977 (through May 31, 1978) will be reported. In
addition, a sample interview of patients and the general population will be
sought in each'location. It will be necessary to evaluate the new morbidity
data in light of known and suspected etiological factors (both genetic and
environmental).
At this juncture the incidence phase is well under way and promises
fruitful results. A pilot test of the interviewing phase has already proved
beneficial in paving the way for a successful telephone interview survey.
Preliminary results from the pilot study indicate that there are important
questions to be taken into consideration, e.g., outdoor exposure patterns,
skin reaction to sunlight (burn, tan)5 skin conditions (freckles), ethnic
group (genetics), skin complexion, industrial exposure, etc.
-------
TABLE OF CONTENTS
v '
Page
Introduction 1
Progress Update - Incidence Pha,se 3
Interviewing Phase - Report on Pilot Study 7
General Procedures 7
Results 11
Direct Responses vs. Proxy Responses 12
Plan A vs. Plan B 12
Telephone Interview vs. Personal Interview... .' 18
Patient Responses vs. General Population Responses 18
References .;. .... 22
Tables and Figure
Table 1.. ................."... .... 4
Table 2 9
Table 3.. 10
Table 4................................................. 13
Table 5 ...16
%
Table 6 .......................'....... 19
Figure 1 8
-------
Introduction
o
The National Cancer Institute (NCI) in collaboration with the Environmental
Protection Agency (EPA) is conducting basic epidemiologic research to study the
potential human health effects of stratospheric ozone depletion. It was
4
essential that this project be implemented as soon as possible in order to
provide the Biological and Climatic Effects Research (BACER) program information
ป
which is urgently needed to decide on guidelines and restrictions for our
country's use of certain chemicals known and suspected of depleting the earth's
protective ozone "shield".
The NCI had previously conducted a short-term study of the incidence of
nonmelanoma skin cancer in four locations which were part of the Third National
Cancer Survey (TNCS 1969-72): Dallas-Ft. Worth, San Francisco-Oakland, Iowa
and Minneapolis-St. Paul. ' While the results of this study were being
analyzeds the Department of Transportation (DOT) had already embarked on its
famous Climatic Impact Assessment Program (CIAP). This program was initially
concerned with the effects of stratospheric pollution (e.g., ozone depletion)
from supersonic aircraft (SST) exhausts, such as nitrogen oxides. Responding
- to DOT's need, the NCI formed the Ultraviolet Radiation and Skin Cancer Working
Group. Members included representatives from Temple University; Queensland,
Australia, the National Oceanic Atmospheric Administration (NOAA), prominent
scientists in the field of medicine and phys'ics as well as the leaders of the
DOT's CIAP program. Under the auspices of this working group, measurements
of ultraviolet radiation (UV-B) were made in various locations of the United
States. By design, the four locations of the TNCS's skin cancer survey were
(?)
also included, NCI produced a. monograph^c/ of the measurements from each of
ten locations for one entire year (48S half-hour "readings per day), 1974, and
compared UV-B measurements with available morbidity data: four locations for
-------
nonmelanoma skin cancer, and nine locations for melanoma of the skin. As
ป
expected, NCI showed that the incidence of skin cancer was associated with
increased UV exposure, and that UV count was correlated with latitude,
altitude sky cover, season of the year, and time of day. ' ' '
4
Concurrently., other scientists 'had reported that chlorofluoromethane
(CFMS) gases such as those used as propellants in aerosol cans and as
refrigerants (freons) could deplete stratospheric ozone in potentially
devastating proportions. One estimate was that the earth may realize a 7 to
9 percent total ozone level depletion by the year 2000, if continued use of
CFM's persisted at current levels.
It was further recommended by another interfederal agency task force3
the Inadvertent Modification of the Stratosphere (IMOS) Committee., that
much basic research was needed to study and estimate potential harmful
biologic effects of ozone depletion both human and nonhuman in the very
near future. ' It was specifically noted that more epidemiologic data on
skin cancer were urgently needed to provide better estimates of the amount
of increased risk to human skin cancer which may result from various amounts
of increased UV-BS resulting from suspected ozone depletion. Utilizing the
existing data from the NCIS the current or tentative estimates of the
V
biological amplification factor is put at 2, for increases of UV-B of less
than 10 percent. This means that a one percent increase in UV-B is expected
to result in a two percent increase in skin cancer. Combined with a physical
amplification factor, also put at 2 (the physical amplification factor of 2
means that a one percent decrease of ozone results in a two percent increase
in UV-B), this means that a fourfold increase in skin cancer may result from
low levels of ozone depletion. With the present state of the art, it is also
known that the effects will be amplified to much greater degrees for ozone
-------
3
depletion levels greater than 10 percent. However, there still remains a
great deal of variability in many"of these estimates thus the need for
quick-yielding basic research projects.
Again, the NCI responded to the need by recommending the utilization
of its continuing surveillance, epidemiology and end results (SEER) program.
Seven SEER participants are now engaged in the latest epidemiologic effort.
In addition to basic morbidity information, the NCI will attempt to obtain
relevant information from the general populations as well as a sample of
patients in each location. The new information sought deals with factors
which may be related to genetic susceptibility (e.g. ethnic group, eye
color, hair color,, skin color, etc.) and environmental susceptibility (e.g.
outdoor exposure habits, occupational exposure, sunburning, tanning,
protective measures, etc.). The method of obtaining information employs
sample telephone interviewing techniques. The specific contractors engaged
in this project are listed in Table 1. A pilot test of the interviewing
phase is being conducted at the University of Minnesota. This location was
chosen because the same contractor had provided NCI with incidence information
from the TNCS and had independently followed-up the survival of skin cancer
patients diagnosed in the earlier study, thus providing a test data base for
patient interviewing. Computer support services are provided by the Geomet
Company, and sampling and interviewing support are provided by the Westat
Company.
Progress_Update - Incidence Phase
At a recent general meeting of the BACER program, held at the University
of Maryland, a presentation of all activities through mid-September was made (see
attachment 1, written transcript, September 19, 1977). As of December 9, 1977
-------
TABLE 1
List of Contractors
Location
Seattle
San Francisco
Detroit
Atlanta
New Mexico
New Orleans
Utah
Minneapolis-
St. Paul
Support Services
Bethesda, Md.
Bethesda, Md.
ซ
Contractor
ป
Fred Hutchinson
Cancer Research
Center
California Tumor
Registry
Michigan Cancer
Foundation
Emory University
New Mexico
Tumor Registry
Tulane University
Utah Cancer
Registry
University of
Minnesota
Contracts
Geomet
Westat '
No. of Documents
submitted through
December 9, 1977
606
456
770
13257
965
460
1,100
688
Purpose
Data Processing
Sampling,
Funding
Appropriated*
$74,842
$85,720
$87,190
$123,469
$55S785
$91,201
$34,060
$71S505
$123,870
$92 , 358
Interviewing
"Funding appropriations for the seven SEER field
offices cover only phase 1, incidence reporting
-------
participating areas have submitted 5664 incidence-related documents to NCI
for processing. This represents about 20 percent of the total amount we
expect for one year's survey. Slow starts in some areas like
i
San Francisco-Oakland, New Orleans and Atlanta were due mainly to
administrative problems.
Each location has reported that cooperation from the medical community
has been received, and that each expects to provide meaningful incidence
data during the course of the survey. Examples of the kinds of activities
going on in several field offices are provided in attachment, 2. There are
a few individual physicians' in some locations who may not participate in this
study; however3 the amount of cases involved appears to be minimal and
accountable by various means.
As site visits were made to each field office, it became apparent that
several new codes could be added to our abstracting'procedures to facilitate
reports by cell type. In particular, it v/as observed that some locations
were accessing a large number of in-situ carcinomas and "Bowen's disease" of
the skin. The item (#25) for cell type on our abstract forms was expanded
to include several additonal codes.
V
An interactive central computer system has been developed and installed.
As documents are received from the field offices, they are edited manually as
well as by computer. Change documents are submitted by field office personnel
to correct errors and update files. A summary of the computer system design
is given in attachment 3.
-------
In related work, the New Mexico field office has estimated that the
amount of malignant melanomas of the skin observed for "Anglos", i.e.,
Caucasians other than those Spanish ancestry, in the Albuquerque area,
was consistent with what would have been projected from the UV and melanoma
data provided by the NCI.
Further, the city of Albuquerque, Second Council., has endorsed the
New Mexico Melanoma project of the New Mexico Cancer Control program which
is engaged in a prevention program* making people aware of the sun related
association of increased risk to melanomas of the skin (see attachment 4).
Obviously, it is too soon to provide new estimates of nonmelanoma
skin cancer morbidity from the new SEER locations. Completion of
documentation for a full year's survey is expected by the end of the
summer of 1978. Meanwhile, the NCI will evaluate melanoma data which is
being routinely collected by the same SEER participants. Preliminary
indications are that the incidence of melanoma of the skin are continuing
to increase during the 1970's. It is not clear,, however, to what degree
increased amounts of UV-B or changing lifestyles may have affected the
apparent rising trend.
-------
Intervi ewi ng Phase - Reporb on Pj^ot Study
There are various epidemiologic factors known or suspected of influencing
the development of skin malignancies in humans. The objectives of the pilot
study conducted at the University of Minnesota were to develop a telephone
interviewing technique which would yield-high response rates., and to produce
a meaningful set of questions which would provide answers to pertinent factors
relating to the individual's genetic and environmental susceptibility.
The pilot study design is given in figure 1. Definition of terms are
given in Table 2. Numbers of households and individuals interviewed are
given in Table 3.
The decision to attempt a short, 10 minute telephone interview was made
because the alternate approach (person-to-persor. interview) was determined to
be too costlya both in time and money. In the pilot study a systematic sample
of telephone numbers from the Minneapolis-St. Paul area was used to provide
sample responses from the general population. In the final study designs we
expect to employ a random digit dialing technique developed by the Westat Co.
(see attachment 5). This would allow the inclusion of samples of households
with unlisted telephone numbers* In the Minneapolis-St. Paul areas it was
estimated that only a small proportion of numbers were unlisted and that over
93% of the households have telephones.
General Procedures
Simultaneously, samples of households from the general population were
selected and interviews were obtained via Plan A or Plan B. After a decision
was made as to which plan was to be used, in this case. Plan B was selected, a
-------
8
FIGURE 1
PILOT STUDY DESIGN
PLAN A
Telephone
150 Direct
39 Proxy
PLAN B
Telephone
198 Direct
61 Proxy
PLAN A
Personal
63
PLAN B
Personal
75
PLAN B
Patients
84
-------
TABLE 2
Pilot Study Plan Definition of Terms
1. Instrument: Questionnaire containing 24 questions or less,
including skin complexion chart.
2. Interview: Telephone communication to obtain responses to
questionnaire.
3. Direct Interview: Interview conducted directly with sampled
household member.
4. Proxy Interview: Interview conducted with a household member
who provides responses for another member of the household.
5. Personal Interview: Person to person interviews obtained after
telephone interview is obtained (a repeat of the interview is
given to confirm telephone responses).
6. General population: Households obtained from sampling of area
telephone directory.
7ซ Patients: Sample of patients who were diagnosed with basal cell
or squamous cell carcinoma of the skin in Minneapolis-St. Paul
(SMSA) during the TNCS (1971-72). Physicians permission must
have been obtained prior to initial contact.
8. Plan A:
1. Telephone sampled household from General/Operation.
2. Obtain free and informed consent to interview.
3. Conduct interview of selected respondent,
V
9. Plan B:
1. Telephone sampled household.
2. Obtain mailing address from proposed respondent.
3. Mail out questionnaire to household.
4. Obtain free and informed consent to interview.
5, Conduct.
-------
10
TABLE 3
Numbers of Households and Individuals in Pilot Study
General Population Patients
Plan A Plan B Plan B
No. Household
contacted \ 144 170 157
No. Household
Interviewed 117 145 84
No. Household . .
Refused '27 25 73
Response Rate (%) 81 85 54*
No. Individuals
included 189 _259_ 84
by direct interview 150 198 ;19
by proxy interview 39 61 65
*
This response rate was low mainly because old patients
(over 75 years) refuse to response for various reasons
(senilitys infirmity, etc.)ซ The patients, who were old
to begin with, were five years older than they were at
time of diagnosis (1971-72). Although the response rate
is expected to improve for current patients, we may in
subsequent studies, limit our comparison group to those
under 75 years, if responses from older individuals
continue to be unreliable.
-------
11
sample of the patients were interviewed according to the preferred plan. To
verify that responses received by telephone were valid and reliable, a sample
of respondents were also given person to person interviews. Sample sizes
are provided in Table 3. Responses from person to person interviews were
then compared with telephone responses. Between 20 and 25% of the individual
responses from the general population were obtained by proxy, interview.
Therefore, it was necessary to verify that oroxy data were also reliable.
Responses for each question (see attachment 6) were evaluated in total and
for each plan type to determine if the questions were appropriate,
understandable, and productive. . .
Results
After reviewing the responses from each question and conferring with
consultants to the BACER program, we found several problem areas which
required major changes.
General occupational history as well as industrial exposure questions
were found difficult to administer and were therefore made more specific.
Questions on childhood exposure patterns, use of sunscreens and sunlamps, use
of hats or long sleeves were not as productive as had been anticipated.
V
Vacation information questions produced variable response and required
improvement. Questions on outdoor exposure patterns on the job also needed
to be spelled out more specifically for various ages. The question of utmost
consideration was that on skin complexion. Although the skin complexion
chart appeared to work, there was room for improvement. We have found we
could obtain better skin color tones by employing a printing process
recommended by representatives of the National Geographic magazine. As a
result of the pilot, a new instrument has been designed (see attachment 7),
-------
12
which deletes olds nonproductive questions,, restructures the more meaningful
questions, and provides an improved skin complexion chart.
Direct Responses vs. Proxy Responses
The average proxy response was found to be comparable to the average
direct response for most questions (see Table 4). Proxy responses, however,
must be interpreted with caution, since most proxy interviews were obtained
from female members of the households. Of particular concern are questions
relating to sunburn effects and tanning ability.
Any misgivings about proxy information that the investigators may have
had were found to be outweighed by the increased sample sizes they provide,
at minimal costs (Table 3). ' We anticipate that proxy information will be
obtained in subsequent study. .
Plan A vs. Plan B .
Questions asked in Plan A were identical to those asked in Plan B except
for those relating to skin complexion. Mo basic differences in responses
were found between Plan A and Plan B except of course for that skin complexion
information (see Table 5). In Plan As the individual was asked whether he
considered his skin color to be "fair", "medium"9 or "dark". In Plan B9 a
skin complexion chart was provided on the bottom of the questionnaire (see
attachment 4) which the individual received'in the mail, and he was asked to
compare his skin complexion against the chart. The latter has the apparent
advantage of providing a standard of comparison and thereby eliminating any
personal or regional biases which may exist with respect to skin color. The
investigators were, however, concerned that two telephone contacts (i.e..
Plan B) would result in a lower response rate compared to Plan A. However,
the response rate was 81% for Plan A and 85% for Plan B essentially no
difference (Table 3).
-------
TABLE 4
Direct Responses vs. Proxy Responses
Telephone Interviewing Method
No. responses
PLAN A
Proxy
39
PLAN A
Direct
150
PLAN B
Proxy
61
PLAN B
Direct
198
Hours - Outdoors^
Weekdays x"
Weekends
m
x
m
Hours - Outdoor^
Vacation x
m
0x
Sunbathe
Frequently
Occasionally
Rarely
Never
Reaction to Sun
No Reaction--
Some Redness
Burn
Painful Burn
Not in Sun
Other Reaction
Age of Respondent
x"
m
Type of Tan
None
Light Tan
Average Tan
Deep Tan
Other
Not in Sun
Freckles
Yes
No
19.2
13.0
2.5
12.1
11.9
1.0
40.0
33
3.6
17.9
10.3
23.1
48.7
19.0
35.9
30.8
20.
10.
,5
.3
2.6
41.7
38.0
2.3
2.6
30.8
23.1
43.6
14,
1,
11.5
9.0
.5
43.3
41.0
1.8
20.0
27.3
28,0
24.7
40:9
39
10
10
.6
,7
,7
,7
,7
38.5
61.5
39.4
35.0
1.2
6.7
11.4
47.0
33.6
1.3
37.3
62.7
.21.6
15.0
2.1
12.0
11.3
1.0
42.9
41
2.8
9.8
26.2
16.4
47.5
36.1
49.2
8.2
1.6
4-9
40.5
35.5
2.0
4.9
14.8
35.1
44.3
16.2
12.0
.9
10.6
9.0
.4
40.2
40.0
1.5
19.3
25.4
29.4
25.9
34.2
40.3
16.8
5.6
3.1
"18.0
82.0
5.6
19.2
41.4
31.3
1.0
1.5
45.7
54.3
-------
Table 4 (cont'd.)
14
Eye Color
Blue 43.6
Brown 38.5
Green 15.4
Other 2.6
Hair Color
Black 5.3
Brown 60
Red t'-
Other 2f:.;
Years Worked on
Primary Occupation
Y . 16.1
m 14.5
ฐx 2.0
Skin Conditions
Acne 35.9
Psoriasis 5.1
Chemical Exposure
Yes 35.9
No 64.1
Ethnic Background
Irish 17.9
English - 10.3
German 33.3
Scandinavian 30.8
Skin Complexion
Chart
3
4
5
6
' 7
8
9
10
Skin Cancer
Complexion Chart
Matches
Exactly
Closely
Not Closely
42.7
40.7
10.7'
6.0
11.3
69.3
2.0
17.3'
12.8
9.5
1.0
56.0
3.3
26.0
74.0
25.3
20.7
SOoO
34.7
49.2
29.5
11.5
9.8
6.6
73.8
1.6
18.0
13.9
10.0
1.5
39.3
1.6
31.1
68.9
14.8
26.2
47.5
24.6
5.8
21.2
9.6
5.8
21.2
3.8
32.7
40.4
29.8
15.7
14.1
3.5
62.6
2.5
31.3
13.3
10.0
.8
59.1
2.5
18.7
81.3
25.8
21.7
49.5
39.9
1.6
3.2
7.0
7.0
3.2
11.9
30.8
35.1
3.9
86.3
9.8
5.5
81.3
13.2
-------
15
Table 4 (cont'd.)
Chart Comparison
with SKIN
Lighter
Darker
Skin Color
Fair
Medium
Dark
34.9
65.1
33.7
66.3
46.2
35.9
17.9
44.2
49.0
6.8
-------
TABLE 5
Plan A vs. Plan B
16
No. responses
Telephop.e Interviews vs.
PLAN A PLAN B
189 259
Personal Interviews
PLAN A
63
PLAN B
75
Hours - Outdoors
Weekdays x"
Weekends
m
a
x
m
cr
19,0
14.5
1.1
11.6
9.6
0.5
17.5
13.0
0.8
10.9
10.0
0.4
18.9
13.0
1.8
10.7
9.5
.7
17.7
10.0
1.7
11.0
9.0
0.7
Hours - Outdoors
Vacation x"
m
a
42.6
40
1.6
40.8
40
1.3
44.1
41.0
2.5
41.0
36.0
2.6
Sunbathe %
Frequently 19.6
Occasionally 23.8
Rarely 27.0
Never 29.6
Reaction to Sun %
No Reaction 39.9
Some Redness 37.8
Burn 12.8
Painful Burn 8.0
Not in Sun 1.1
Other Reaction 0.5
Age of Respondent $
x 39.9
m 35.5
ฐTT 1.0
17.1
25.6
26.4
31.0
34.6
42.4
14.8
4.7
CD Bป
3.5
40.9
37.0
0.9
19.0
22.2
28.6
30.2
30.2
47.6
12.7
9.5
36.6
34.5
1.5
13.3
21.3
30.7
34.7
32.0
45.3
17.3
4.0
1.3
39.8
34.0
1.8
Type of Tan
None
Light Tan
Average Tan
Deep Tan
Other
Not in Sun
Freckles
Yes
No
5.9
15.4
42.0
35.6
.1.1
37.6
62.4
5.4
18.1
40.2
34.4
0.8
1.2
39.1
60,9
6.3
19.0
38.1
35.5
46.0
54.0
5.5
17.8
41.1
32.9
2.7
41.3
58.7
-------
Table 5 (cont'd.) . 17
Eye Color % % % %
Blue 42.9 42.5 33.3 44.0
Brown 40.2 v . 29.7 46.0 25.3
Green 11.6 14/7 14.3 16.0
Other 5.3 13.1 6.3 14.7
Hair Color ' % % % %
Black 10.1 4.2 7.9 8.0
Brown 67.6 ' . 65.3 69.8 64.0
Red 2.7 2.3 6.3 1.3
Other 19.7 28.2 15.9 26.7
Years Worked on
Primary Occupation % ' % % %
x 13.6 13.4 11.7 13.5
m ' 9.0 10.0 9.5 10.0
ฐ~. ' 0.9 0.7 1.1 1.0
X
Skin Conditions % % % %
Acne 51.9 54.4 58.7 58.7
Psoriasis .. 3.7 2.3 -- 5.3
Chemical Exposure % % % %
Yes 28.0 21.6 39.7 25.3
No 72.0 78.4 60.3 74.7
Ethnic Background % % % %
Irish 23.8 23.2 27.0 29.3
English 18.5 22.8 28.6 18.7
German 46.6 - 49.0 57.1 60.0
Scandinavian 33.9 36.3 34.9 44.0
Skin Complexion % % %
Chart 1 . 1.2
3 1.3 .1.6
4 3.8 1.6 1.3
5 10.1 19.0 21.3
... 6 ' , 7.6 6.3 9.3
7 3.8
8 13.9 3.2 5.3
9 24.9 . 31.7 26.7
10 34.6 36.5 4.6
Skin Color Complexion
Chart Matches % % %
Exactly 5.2 3.2 2.8
Closely 82.4 84.1 83.3
Not Closely 12.4 11.1 13.9
Chart Comparison with ซ ป, /
SKIN % k
Lighter 34.0 19.0 18.8
Darker 66.0 " 81.0 81.2
Skin Color % % % %
Fair 44.6 39.7 48,3 41.2
Medium 46.2 52.6 45.0 52.9
Dark 9.1 7.7 6.7 5.9
-------
18
Telephone Interview vs. Personal Interview
o
Responses obtained from telephone interviews were found to be in agreement
with responses obtained in person, within reasonable limits for a great
majority of questions. The level of agreement ranged between 60% and 95%
for telephonic responses compared to personal responses. The proportion of
responses in exact agreement varied by question (e.g., sunbathing, 63%,
reaction to sun, 64%, type of tan, 70%s eye color, 86%, state of major
residence, 88%). Just as importantly, however, the average response for
telephone interviews was comparable to the average response for personal
interviews, in Plan A and Plan B (see Table 5).
Patient Responses vs. General Population Responses
In addition to providing a test of the questions and methods of
Interviewing, the pilot study also offers9 on a small scale, a comparison
of responses from patients and the general population. It must be
emphasized that these results are from" a small pilot study, arid not to be
interpreted as definitive.
All patient interviews were administered via Plan B. Because the
patient group was relatively old (only five patients were under 50), averaging
about 65 years, the comparison group used was that for Plan Bs among the
general population over 50 years of age. Comparisons were made separately
for each sex group. Table 6 shows the results for selected questions of
particular interest. It was surprising to find that the difference between
comparison groups for hours of outdoor exposure were small. A greater
surprise was the finding that the proportion of individuals of Scandinavian
ancestry in the patient population was found not to be dramatically large
compared to that found for the general population group. Individuals of
-------
TABLE 6 . Iy
Patients compared to General Population Surveys Individuals 50 years and over
Plan B Only
Telephone Interviewing Method
No. responses
PATIENTS
MALE .
46
PATIENTS
FEMALE
35
PLAN B over 49
MALES
33
PLAN B over 49
FEMALES
42
Hours - Outdoors
Weekdays x"
m
a
Weekends
x
x
m
a
x
21.1
16
2.3
13.2
10
1.6
13.8
10
2.0
9.9
7.9
1.1
26.6
20
3.2
12.2
10
1.1
14.2
10
1.3
8.9
7.9
.76
Hours - Outdoors^
Vacation x"
m
cr
41.1
41.0
3.0
35.6
29
4.3
36.6
36
3.5
35.8
32.5
3.8
Sunbathe
Frequently
Occasionally
Rarely
Never , __
Reaction to Sun
No Reaction
Some Redness
Burn
Painful Burn
Not in Sun
Other Reaction
Age of Respondent
x"
m
19.6
8.7
21.7
50.0
28*3
37.0
17.4
10.9
6.5
65.4
66.8
17.1
14.3
17.1
5-1.4
14.3
37.1
20.0
17.1
8.6
2.9
64.7
54.5
2.1
6.1
12.1
24.2
57.6
21.2
54.5
12.1
6.1
6.1
60.4
55
1.7
21.4
9.5
28.6
40.5
40.5
19.0
23.8
11.9
4.8
60.1
59.8
Type of Tan
None
. Light Tan
Average Tan
Deep Tan
Other
Not in Sun
Freckles
Yes
No
10.9
19.6
43.5
21.7
4.3
54,3
45.7
23.5
23.5
41.2
2.9
8.8
71.
28,
9.1
12.1
42.4
36.4
18.2
81.8
9.5
23.8
35.7
21.4
2.4
7.1
42.9
57.1
-------
20
Table 6 (cont'ci.)
Eye Color
Blue 57.8 54.3 51.5 40.5
Brown 17.8 17.1 30.3 26.2
Green 8.9 17.1 12.1 9.5
Other 15.6 11.4 6.1 23.8
Hair Color
Black 8.7 2.9 . 9.1 2.4
Brown 58.7 54.3 72.7 57.1
Red 10.9 5.7 .
Other 21.7 37.1 18.2 40.5
Years Worked on
Primary Occupation
x 28,2 27.6 27.9 21.6
m 27.5 26 28 22
ฐ- 4.6 7.3 1.5 2.0
A
Skin Conditions
Acne 54.3 54.3 42.4 46.3
Psoriasis 4.3 7.1
Chemical Exposure
Yes 37.0 8.6 42.4 19.0
No 63.0 91.4 57.6 . 81.0
Ethnic Background .
Irish 21.7 25.7 18.2 21.4
English 34.8 28.6 18.2 14.3
German 32.6 31.4 54.5 38.1
Scandinavian 26.1 25.7 18.2 47.6
Skin Complexion
Chart
.1 2.2
3 2.2 .
4 .3.4 2.6
5 4.4 2.9 6.9 5.3
6 2.2 5.7 3.4 7.9
7 4.4 2.9 3.4 2.6
8 8.9 5.7 13.8 10.5
9 28.9 28.6 34.5 26.3
10 46.7 54.3 34.5 44.7
Skin Color Complexion
Chart Matches
Exactly 11.4 11.4 3.6 7.9
Closely 72.7 60.0 - 85.7 81.6
Not Closely 15.9 28.6 10.7 10.5
Chart Comparison
with SKIN
Lighter 27.0 35.5 19.2 . 28.1
Darker 73.0 64.5 80.8 71.9
-------
21
o
Irish and English ancestry, however, did appear in greater proportions in
the patient group. A physical characteristic which appears to predominate
among both patient, groups is freckles.
Since the pilot study was not large it offered little likelihood of
detecting small differences between patients and the general population.
Thus the findings of the pilot's patient-general population study are not
unexpected. In contrast to the pilot, the full survey in all areas will
be greatly increased in sizes and therefore has much greater likelihood of
detecting meaningful differences.
-------
22
REFERENCES
1. Scotto, J., Kopf, A.M., and Urbach, F.: Non-melanoma skin cancer among
Caucasians in four areas of the United States. Cancer 34: 1333-1338,
1974.
2. Scotto, J.s Fears, T.R.S and Gori, G.B.: Measurements of ultraviolet
radiation in the United States and comparisons with skin cancer data.
U.S. DHEW, PHS, NIH, NCI, DHEW Pub!. No. (NIH) 76-1029, 1976, 120 pp.
3. Fears, T.R.S Scotto, J., and Schneiderman, M.A.: Mathematical models
of age and ultraviolet effects on the incidence of skin cancer among
whites in the United States. Am. J. Epidemic!. 105: 420-427, 1977.
4. Scotto, J., Fears, T.R., and Gori, G.B.: Ultraviolet exposure patterns.
Environ. Res. 12: 228-237s 1976.
5. Scotto, J. and Fsars, T.R..: Intensity patterns of solar ultraviolet
radiation. Environ. Res. 14: 113-127, 1977.
6. Fluorocarbons and the Environment. Report of Federal Task Force on
Inadvertent Modification of the Stratosphere (IMOS). Council on
Environmental Quality, Federal Council for Science and Technology.
U.S. Govt. Print. Off., Washington, D.C., 1975, 109 pp.
-------
FINAL REPORT
INSTRUMENTATION FOR MEASURING IRRADIANCE IN THE UV-B REGION
J. D. Rowan
K. H. Norris
Instrumentation Research Laboratory
Agricultural Marketing Research Institute
Beltsville Agricultural Research Center
Beltsville, Maryland 20705
EPA-IAG-D6-0168
Project Officer:
R. J. McCracken
Agricultural Research, Science and Education Administration
U.S. Department of Agriculture
Washington, D.C. 20250
Prepared for
Environmental Protection Agency
EAGER Program
Washington, D.C. 20460
-------
Annual Report 1977
EPA Interagency Program on Biological and Climatic Effects (BACER)
Instrumentation for Measuring Irradiance in the UV-B Region
SUMMARY
The responsibility of the Instrumentation Research Laboratory (IRL) in
this program was developing portable instruments for use by biologists to measure
UV-B irradiance in growth chambers, greenhouses, and field plots. A simple
UV-B radiometer and two UV-B spectroradiometers have been designed, constructed,
tested, and put into use in the UV-B research program. Each of these instru-
ments is now being manufactured by commercial firms.
The two spectroradiometers differ only in the monochromators: one has a
single holographic grating; the other, two holographic gratings for greater
stray light rejection. The spectroradiometers automatically scan the 250-400 nm
region in less than 5 minutes, printing a tape of the corrected irradiance as
a function of wavelength. The input is cosine corrected by a specially designed
teflon-bubble diffuser coupled to the input slits. The output of the monochro-
mator is measured with a multiplier-type phototube and a logarithmic response
amplifier. The amplifier output is digitized with a digital voltmeter, and the
digital output is interfaced with a desk-top programmable calculator.
The design provided for the spectroradiometer system response to be stored
in the calculator so that, as the spectrum is scanned, the calculator corrects
the measured signal for instrument calibration and outputs the true spectral
irradiance of the source being measured.
The programmable calculator controls the operation of the spectroradi-
ometer so that, on command, scanning is initiated and readings are recorded for
each nanometer interval. The calculator prints the wavelength and irradiance
for each wavelength interval and, at the end of the scan, reverses the wave-
length drive and returns the monochromator to the starting wavelength. The
calculator is programmed to print an integral for a programmed action spectrum
at the end of each scan. At the completion of the scan, the data can be stored
on a magnetic tape, if desired, for future analyses.
Provision is included for a precise check of wavelength: the spectrum
of a miniature low-pressure mercury-arc lamp is scanned, and the calculator
computes the position of the 253.7 nm and the 296.7 nm lines to a precision of
i 0.01 nm.
The performance specifications approach the requirements for UV-B measure-
ments stated in NBS publication #20, "Optical Radiation News," dated April 1977.
iii
-------
Annual Report 1977
EPA Interagency Program on Biological and Climatic Effects (BACER)
Instrumentation for Measuring Irradiance in the UV-B Region
Our UV-B measuring instruments developed for this project meet the
requested requirements. The broadband radiometer is small, hand-held,
battery operated, and has fast response for rapid measurements. The spectro-
radiometers have the following features:
(1) Fast, accurate, and reproducible.
> (2) Convenient, automatic single-key-stroke operation.
i
t
V,
I (3) Programmable calculator-controlled scanning, Fig. 1.
r
I (4) Logarithmic amplifier for wide dynamic range.
p (5) Calculates and prints true spectral irradiance for each nm
if wavelength, Fig. 2.
I
i (6) Calculates and prints weighting function (Aฃ9).
j: (7) Programmed-wavelength-calibration check.
I (8) Double monochromator unit has very low stray light.
I
| (9) Bandwidth of 2 nm.
)' (10) Temperature stable.
x (11) Receptor has excellent cosine response.
r
[; - (12) Spectra are stored on magnetic tape for efficient data processing.
(13) Portableoperates on small lab cart.
(14) Minor disturbance of area by the "measuring head."
Our spectroradiometer performance specifications approach the requirements
for UV-B measurements stated in NBS publication #20, "Optical Radiation News,"
dated April 1977. These requirements are cited in attached copy of "Making a
UV Measurement?," Electro-Optical Systems Design, 9(6):17, 1977. (page 7)
Cosine Response for UV-B
The spectroradiometers, as well as other UV-B radiation meters developed
in our program, incorporate a new design for cosine-corrected input optics.
We tested all available cosine-correction schemes, including integrating
spheres, diffusing reflectors, and sintered-quartz diffusers. None of these
gave adequate performance for the 250 to 370 nm region, so we developed the
-------
teflon bubble diffuser. The spectral transmission of this teflon diffuser
is shown in Fig. 3. Fig. 4 is a drawing of the input optics for the Spectro-
radiometers. We supplied a similar receptor to our cooperators that have a
Gamma Scientific monochromator, Fig. 5. The response of this type of diffuser
to radiation from different angles is compared with that of a commercial
instrument in Fig. 6. The teflon bubble receptor provides excellent cosine
correction, and has a stable surface which can be readily cleaned. This
type of diffuser is now used in commercial instruments.
Wavelength Accuracy
To check spectroradiometer wavelength accuracy, a low-pressure mercury-
arc lamp is scanned and the mercury line location computed. The wavelengths
of the 253.7 and the 296.7 nm lines are measured with a readout precision
of +_ 0.01 nm, so that wavelength shifts as low as 0.02 nm are readily detected,
Fig. 7. This field check can be made quickly and routinely between measurements
if desired.
Spectroradiometers
Our UV-B Spectroradiometers are now used routinely by technicians to
measure UV-B irradiance in growth chambers, greenhouses, and field plots.
These instruments are identified by the acronyms: IRLSpec-S for the single
monochromator version; IRLSpec-D for the double monochromator; and IRLSpec-SO
for the commercial model single monochromator. Numbers shown on the graphs
for example, 1770815.09identify instrument (first digit), year (next two
digits), month, day, and scan number for that day. The first digit may be
1, 2, or 3 for IRLSpec-S, IRLSpec-D, or IRLSpec-SO, respectively.
Measured performance specifications for the IRLSpec-S spectroradiometer
are as follows:
Wavelength range - 250 to 370 nm
Readout interval - 1 nm
Scanning speed - 0.5 nm/sec.
Spectral bandpass - 2 nm
Wavelength reproducibility - +_ 0.02 nm
Wavelength accuracy - +_ 0.05 nm at 296.7 nm
Radiometric reproducibility - +_ 2%
Radiometric accuracy - +_ 5%
Radiometric range - 0.001 to 2000 mW-m~2-nm~^
Stray light - less than 2 x 10~4 at 285 nm as tested
with a xenon arc lamp filtered with 0.5-mm-thick
cellulose acetate
Cosine correction - within +_ 10%
Output - printed tape direct reading in wavelength
and corrected irradiance at each nanometer interval
from 250 to 370 nm
Size of measuring head - less than 28 x 20 x 10 cm
Temperature stability less than 0.04 nm wavelength shift
for 25ฐC temperature change.
-------
The IRLSpec-D spectroradiometer, with double-grating ir.onochromator,
provides better stray light rejection. This unit incorporates all the
features of the single monochromator unit although the measuring head is,
Of necessity, slightly larger. Observed wavelength instability of the
double monochromator was caused by grating-sync-drive cable temperature
sensitivity. Our technician, George Button, solved this temperature insta-
bility with his cable-stringing technique. The grating-drive cable spring
was eliminated, and the cables were crossed to provide automatic temperature
compensation and wavelength stability.
Measured performance specifications for the IRLSpec-D spectroradiometer
are as follows:
Wavelength range - 250 to 370 nm
Readout interval - 1 nm
Scanning speed - 0.7 nm/sec.
Spectral bandpass - 2 nm
Wavelength reproducibility - +_ 0.02 nm
Wavelength accuracy - +_ 0.05 nm at 296.7 nm
Ra
-------
Measured performance specifications for this commercial IRLSpec-SO
spectroradiometer are as follows:
Wavelength range - 250 to 370 run . ,
Readout interval - 1 run .
Scanning speed - 1.0 nm/sec.
Spectral bandpass - 2 nm
Wavelength reproducibility - +_ 0.02 nm
Wavelength accuracy - +_ 0.05 nm at 296.7 nm
Radiometric reproducibility - +_ 2%
Radiometric accuracy - +_ 5% ,
Radiometric range - 0.001 to 200 mW-m~2-nm
Stray light - less than 1 x 10~4 at 285 nm as tested
with a xenon arc lamp filtered with 0.5-mm-thick
cellulose acetate
Cosine correction - within +_ 10%
Output - printed tape direct reading in wavelength and
corrected irradiance at each nanometer interval from
250 to 370 nm
Size of measuring head - less than 25 x 21 x 13 cm
Temperature stability - less than 0.04 nm shift for
25ฐC temperature change.
Spectroradiometer Sensitivity and Dark Current
The photomultiplier-tube dark current, system correction, and standard-
lamp spectral irradiance are plotted in Fig. 8 for the IRLSpec-S with
Hamamatsu R166 phototube (solar blind response) and Corion SF-1.25 solar
blind filter. The IRLSpec-D photomultiplier dark current (Hamamatsu R212, S-5
response and Corion SF-1.25 solar blind filter), system correction, and
standard lamp spectral irradiance are plotted in Fig. 9. Plots of the IRLSpec-
SO photomultiplier-tube dark current (Hamamatsu R166, solar blind response,
and Corning 7-54 filter), system correction, and standard lamp spectral irradi-
ance are shown in Fig. 10.
The instruments developed in our program are now manufactured. A broad-
band UV-B radiometer is available from Optronic Labs, Inc. Spectroradiometers
similar to ours are advertised in Optical Spectra, 12(1):13,51, and 60, dated
January 1978, and are available from three firms:
EG&G, Inc.
35 Congress Street
Salem, Massachusetts 01970
Gamma Scientific, Inc.
3777 Ruffin Road
San Diego, California 92123
Optronic Labs, Inc.
7676 Fenton Street
Silver Spring, Maryland 20910
Irradiance Spectra
Typical spectra from our IRLSpec-D are plotted in Fig. 11, 12, 13, and 14;
from our IRLSpec-S, Fig. 15 and 16; and from our IRLSpec-SO, Fig. 17.
-------
Irradiances of FS40 and FBZS40 lamps are compared in Fig. 11. A typical
curve for an FS40 lamp with and without a cellulose acetate (CA) filter is
shown in Fig. 12. CA filters with FS40 lamps are compared in Fig. 13. Measured
irradiances of four FS40 lamps used to enhance the sun's UV-B at Snowmass,
Colorado's 2980 m (9777 ft.) site, are plotted in Fig. 14. The fixtures were
two lamps mounted in pairs at a Z of 110 cm and filtered with 0.127-mm (0.005-in.)
CA. The change in irradiance of a FBZS20 lamp after aging 16.5 hr. is shown
in Fig. 15. The irradiance measurements of two Sylvania F15T8 CW lamps at
Z = 8 cm are compared for the IRLSpec-S, Fig. 16; the commercial IRLSpec-SO,
Fig. 17; and the IRLSpec-D, Fig. 18. Irradiance for a 15-watt daylight lamp
is shown in Fig. 19.
Spectral data of the sun at Beltsville, Maryland, during the year are
plotted in Fig. 20 and 21. The sun curve on a very clear day illustrates the
stray light at wavelengths below 289 nm for the IRLSpec-D, Fig. 22. The action
spectra of measured Beltsville sunshine with weighting function 9 are shown
in the region of 294 to 318 nm at the bottom of Fig. 20.
The weighting function plotted on the graphs is the AE9 equation developed
in cooperative research at BARC:
= I 0.25(A/228.178)9'ฐJ exp [ 4-(A/228.178)9-ฐJ
The equation was used to compute UV-B sun equivalent (UVSEB), indicated on the
graphs. Details of its development are presented in the Agricultural Equipment
Laboratory report.
Spectral irradiance data are now plotted automatically with the HP9815A
calculator interfaced directly and controlling a HP9872A plotter. After the
data are recorded on the cassette, selected scans are loaded into the calculator;
and the plotter draws and labels the graph to programmed dimensions, and then
automatically plots the data (Fig. 11, 12, 13, and 14). A curve is plotted from
250 nm to 370 nm in 25 seconds.
Narrow-band Radiometers
Two narrow-band, probe-type radiometers were commercially manufactured to
our specifications. These radiometers have solar-blind photo-multiplier-type
detectors (Mfg. No. R166), a peak response at 294 nm and 299 nm, respectively,
and a half-band width of 5 nm. Irradiance readout is a 3 1/2 digit display
located in the remote electronic unit with switched ranges from 10~^ to 10~H
O 1
watt'cm~^'nm~J-. The small sensor head (5x5x16 cm), with teflon dome receptor,
is attached by 2.75-m cable for easy placement within growth chambers.
Broad-band UV-B Radiometers for Routine Monitoring
A small battery-powered radiometer, the IRLMeter, was designed and con-
structed for measurement of total UV-B radiation from artifical sources. The
instrument has a solar-blind vacuum photo-diode (Hamamatsu R403), an integrated
circuit amplifier, and a microammeter packaged in a meter case. The circuit,
as shown in Fig. 23, provides for four decades of range switching. The relative
wavelength response, as shown in Fig. 24, peaks at 300 nm and is relatively flat
from 280 to 320 nm and the radiometer is not sensitive to radiation longer than
400 nm. Typical comparative data of the IRLMeter with the IRLSpec-S spectro-
radiometer are shown in Fig. 25.
-------
This broad-band UV-B radiometer sensitivity is adjusted to a full scale
reading of 2.0 UV-B sun equivalents, Beltsville (UVSEB) when measuring the
output of FS40 fluorescent lamps filtered with 0.127-mm (0.005 in.) CA.
Correction factors were developed for use with FS40 lamps with 0.254-mm
(0.010 in.) CA filter, and for use with B2S fluorescent lamps.
The IRLSun-meter radiometer is a modification of the IRLMeter. The new
calibration reads directly in sun equivalents, Beltsville (UVSEB. Aฃ9). Cali-
brations for two suns (UVSEB Aฃ9) full scale were developed for four sources:
(1) Sunshine for a clear day (6/8/77), 1:00 p.m. EOT
(2) FS40 lamps filtered with 0.127-mm (0.005 in.) CA
(3) FS40 lamps filtered with 0.254-mm (0.010 in.) CA
(4) FBZS40 CLG lamps (unfiltered)
A commercial version of the IRLMeter has been manufactured, the Optronic
Model 725. These Model 725 radiometers were calibrated by IRL for our coopera-
ting laboratories from IRLSpec-D irradiance values for five sun equivalents
full scale, with FS40 source filtered by 0.127-mm (0.005 in.) CA aged 6 hr.
A calibration scale factor was developed for 0.191-mm (0.0075 in.) CA, 0.254-mm
(0.010.in.) CA, 1.52-mm (0.036 in.) petri dish, and FBZS40 and FBZS20 lamps.
An Optronic Model 725 broad-band radiometer was calibrated from IRLSpec-D
irradiance values for 10 UVSEB full scale with a Rayonet F8T5 RPR3000A (8 watt)
lamp source filtered with 0.127-mm (0.005 in.) CA supplied by the Peoria coop-
erators and then aged 6 hr. This Peoria CA filter absorbed less at shorter
wavelength than Beltsville CA, Fig. 26, 27, and 28, significantly changing the
Aฃ9 action integral.
Rayonet F8T5 RPR3000A (8 watt) lamps have strong energy in the 254 nm
region, the energy approaching the peak value at 313 nm (ratio 1.4). The peak
energy of the FS40 at 313 nm is approximately 700 times the energy at 254 nm.
This 254 nm energy can be removed with CA filter, but it probably accelerates
the aging of the CA filter. The irradiance of the Rayonet RPR3000A with the
CA filter at 20 cm approximates the energy of the FS40 with CA filter at 50 cm
and 75.1 cm, Fig. 29, 30, 31, and 32.
Eye Protectors
Personal eye protectors were evaluated for their UV-B attenuation, and
two FS40 lamps at 50 cm from the IRLSpec-D spectroradiometer receptor were used.
An aperture through black cloth was necessary to prevent significant leakage
between lens and receptor, even though they were placed as close as possible.
Plots of plastic goggles No. 6, 7, and 8 (dashed line) are at the 0.001 irradiance
level in Fig. 33. No. 5 was a pair of plastic goggles, and No. 2, a plastic
face shield. UV-B blocking of prescription eyeglasses (glass and plastic) and
sunglasses are shown in Fig. 34.
-------
MILT.* S. KIVKB
Tin- Engineering Magazine of Electro-Optical and Laser Technology
June, 1977. Vol. 9, No. 6
SKILNYK
& Vice President
Kditnr
WILLIAM S. HUDSPETH
Atxotiate Editor
WILLIAM D. ASHMAN
Lois ESMAIL
Circulation Manager
CKNE BRIKSKE
Marketing Services
EDITORIAL ADVISORY BOARD
L. lieiser, Consulting Physicist
I). Belforte, Aveo Everett Research Lab I
R. Buzzard, Lexel
D. Casasent, Carnegie-Mellon Univ.
W. Hunter, Naval Research Lab
G. Klauminzer, Molectron
H. Lavin, General Electric
D. Lockie, Watkins-Johnson
M. Pasture!, Coherent Radiation
W. Ruderman. Interactive Radiation
R. Watson, ฃGiซn.. |ni.. Published mimlhly by Miltnn S.
Kivpr Publications, Inc. Printed in U.S.A. Control- t
lซi circulation paid at Pontiac. Illinois and Chicago,'
llhnoi. 60607. Subscrintion price: $3 per copy, $20
pf year. POSTMASTER: Send Form 3679 to,
tl.ECTRO-OPTlCAL SYSTEMS DESIGN, 222 West
Adims. Chicago. III. G060S. (312) 263-1866.
OFFICIAL PUBLICATION
JUNE. 1977
4
4
6
6
6
Q
12
12
12
13
14
17
17
Laser "Writes" in Liquid Crystal
And Now: The Free Electron Laser
The Latest in Wrist Wear
Start Clicking for Nikon's Photo Contest
Heat-Generated Ultrasonic Waves
c..f..-n O,I~Kป~~<. ซ~. i /irvซ.
Quality Control Session to be Featured
Coherent Laser Radiation Shrinks to 38 nm
Unconventional Form for CRT Terminal
Need a Laser Diode?
Instant Movie Camera Debuts
BRH OKs First Laser Safety Standard Variance
Making a UV Measurement?
taking a UV Measurement?
Washington, D. C. The increas-
ing interest in ultraviolet optical
measurements has spurred the
National Bureau of Standards' Op-
tical Radiation and Radiometric
Physics groups to look into the
matter. (A special chapter of the
NBS Self-study Manual on Optical
Radiation Measurements will be
devoted to making state-of-the art
accuracy UV measurements and is
planned for later this year.) Over
the past few months, NBS has in-
vestigated and characterized sev-
eral currently available spec-
troradiometers suited for use in
the UV, in a program sponsored by
the EPA.
To summarize their findings (see
Optical Radiation News, April
1977), the current state-of-the-art
for spectral irradiance measure-
ments in the 250-350 nm range var-
ies widely. The uncertainty for a
highly controlled laboratory mea-
surement of radiation of a simple
character is estimated at about 3%.
At the other extreme, the mea-
surement of very complex radia-
tion under very unfavorable field
conditions could hit 26%. Typical
uncertainties for lab and field are
estimated to be about 6% and 15%,
respectively.
What can be done to improve
these "best" UV measurements?
NBS believes that what is needed
most is a field-portable spec-
troradiometer with some impres-
sive characteristics. This instru-
ment would not only have negligi-
ble out-of-band leakage (=* 10~8) and
high wavelength accuracy ( 0.03
nm) but would also be much less
sensitive to temperature and
mechanical shock, be hermetically
sealed and have an easily replace-
able window on its entrance port.
Difficult field measurements could
be made to within 10% uncertainty
with such an instrument. NBS also
suggests microprocessor control
for this device.
(Ed. note: The technology for such
a device exists. But such a spec-
troradiometer would necessarily be
a "big ticket" item even if we disre-
gard the mechanics of just how the
development expenses are amor-
tized. There is some question of
whether the potential market is big
enough to be able to absorb the de-
velopment costs.)O
L
-47
-------
LIST OF FIGURES
Fig. 1 - Spectroradiometer components
Fig. 2 - Typical UV spectra
Fig. 3 - Teflon spectral transmission
Fig. 4 - IRLSpec-S receptor optics
Fig. 5 - IRL receptor optics for gamma
Fig. 6 - Receptor cosine response
Fig. 7 - Hg line wavelength check - IRLSpec-D
Fig. 8 - IRLSpec-S sensitivity and dark current
Fig. 9 - IRLSpec-D sensitivity and dark current
Fig. 10 - IRLSpec-SO sensitivity and dark current
Fig. 11 - Lamp spectra (unfiltered) FS40, FBZS40CLG1076 and weighting
function 9
Fig. 12 - Lamp spectra FS40 and cellulose acetate 6.5 hr. Z = 30 cm,
Z = 50 cm
Fig. 13 - Lamp spectra FS40 and 0.127 mm, 0.191 nan, 0.254 mm cellulose
acetate
Fig. 14 - Lamp spectra, Snowmass, Colorado
Fig. 15 - Lamp aging - BZS20CLG0377
Fig. 16 - Sylvania F15T8 CW lamp spectra - IRLSpec-S
Fig. 17 - Sylvania F15T8 CW lamp spectra - IRLSpec-SO
Fig. 18 - Sylvania F15T8 CW lamp spectra - IRLSpec-D
Fig. 19 - GE F15T8D lamp spectra - IRLSpec-D
Fig. 20 - Sun spectra - Beltsville, MD
Fig. 21 - Sun spectra - Beltsville, MD - IRLSpec-S
Fig. 22 - Sun - Beltsville, MD - IRLSpec-D
Fig. 23 - UV-B IRLMeter circuit
Fig. 24 - IRLMeter spectral response
Fig. 25 - Regression IRLSpec-S vs. IRLMeter
-------
Fig, 26 - Rayonet F8T5 RPR 3000Aฐ spectra new cellulose acetate, Z = 16.3 cm
Fig. 27 - Rayonet F8T5 RPR 3000Aฐ spectra cellulose acetate aged 6 hr., Z =
16.3 cm
Fig- 28 - Rayonet F8T5 RPR 3000Aฐ spectra cellulose acetate aged 6 hr., Z =
20 cm
Fig. 29 - Rayonet F8T5 RPR 3000Aฐ and sun (Beltsville) spectra
Fig. 30 - Spectra FS40 and Rayonet F8T5 RPR 3000Aฐ
Fig. 31 - Spectra FS40 and cellulose acetate aged 0.5 hr., Z =.50 cm
Fig. 32 - Spectra FS40 and cellulose acetate aged 6 hr., Z = 50 cm
Fig. 33 - Spectra - eye protective device
Fig. 34 - Spectra prescription eyeglasses and sunglasses
-------
r
TEFLON RECEPTOR
SINGLE
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UV-B IRLSpec-S
3-31-77
Fig. 1. Spectroradiometer components
r
Jy1E_ASURINฃ
-TEFLON RECEPTOR
DOUBLE
MONO-
CHROMATOR
^
MOTOR
IP M T |
j PHOTOMETERr""^ j
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UV-B IRLSpec-D
3-31-77
-------
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250
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SUM FROM
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TO
369
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3428.0341
SUM FROM
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TO
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SUM FROM
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320
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Fig. 2, Typical UV spectra
-------
1.4-
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300
400
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500
600
MONOCHROMATOR
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FRONT
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MIRROR
rig. 4. !RLSpec-S RECEPTOR OPTICS
3-31-77
12
-------
FRONT
SURFACE
MIRROR
0,020" TEFLON
GAMMA MONOCHROMATOR
rig. 5. IRL RECEPTOR OPTICS FOR GAMMA
3-31-77
13
-------
TRUE COSINE
CURVE
DATA POINTS () ARE
IRL COSINE RESPONSE
TEFLON (0.020ฐ) BUBBLE RECEPTOR*
80 SO 40 20 0 20 40 60 80
ANGLE - DEG.
Fig. 6. Receptor cosine response
11-3-76
14
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Fig. 7. Hg line wavelength check - IRLSpec-D
15
-------
IE-SB-
HAVB1OJ na IRL AK?I FR USJA
Fig. 8. IRLSpec-S sensitivity and dark current
16
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Fig. 9. IRLSpec.D sensitivity and dark current
11 7
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1E-02
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= 75.1 cm
filฎf 3 i_ES4Q_unfiU:eied
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WAVELENGTH n*> IRL AMRI ARS USDA
Fig. 11 Lamp spectra (unfiltered) FS40, FBZS40CLG1076 and weighting function 9
*.. 19
-------
IRL AM FR (HJA
Fig. 12 Lanp spectra FS40 and cellulose acetate 6.5 hr. Z ป 30cm, Z = 5Cbm
. 20 <
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WAVELENGTH nm
IRL AMRI ARS USDA
Fig. 13 Lamp spectra FS40 and 0.127 mm, 0.191 mm, 0.254 mm cellulose acetate
21
-------
Snowmass, Colorado
, .
Z(cm)
0.001
C53 SJ (S3
CM CM CO CO CO CO
WAVELENGTH nm IRL AMRI FR USDA
Fig. 14. Lamp spectra, Snowmass, Colorado
22 ,
CO
-------
I i
Fig. 15. Laiap aging - BZS 20 CLG0377
-------
2-3
LJ - .'
a a -:
N < >
J >?rn
Fig. 16 Sylvania F15T8 CW lamp spectra - IRbSpec-S
24
-------
.00/1
Fig. 17. Sylvania F15T8 CW lamp spectra IRLSpec-SO
25
-------
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Fig. 18. Sylvania F15T8 CW lamp spectra IRLSpec-D
26
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Fig. 19. GE F15T8D lamp spectra IRLSpec-D
27
-------
2770630. BSfflf i ladL 76*J3 (Int. 9)_18
5*_0(Int_91_L
flCTnfc fl)
0. 001 ,
. WAVELENGTH nm
Fig. 20. Sun spectra - Beltsville, MD
28
IRL AMRI.ARS USDA
-------
'e&e
Fig. 31. Sun spectra - Baltsville, MD - IRLSpec-S
-------
R I i ;:;_i
'M444-l-H-i
V. L L Ei. M b
IRLSpec-D UV-B
Z Y\ m B w
IRL-AMRI-ARS-USDA
77P630-
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2.0 30 40 356 go
Fig. B2. Sun - Beltsville, MD - IRI-Spec-D
30
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2-7-77
RESPONSE OF UV~B IRL METER
PT-R403 " 0-0625 TEFLON DISK
t 1
I I I
t I I _ 1 i I
220 240 260 280
300 320 340 360
Xnrn
380 400
Fig. 24- IRLMeter spectral response
31
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Fig. 27. Rayonet F8T5 RPR 3000Aฐ spectra cellulose acetate aged 6 hr.,
Z = 16.3 cm
34ซ
-------
AE9 Zfcnd
2Z11103J5. FILE 0-3z
2221103.20. EILEJht3L
0.601
oa
in ID
CD CO CO
WAVELENGTH nm IRL AMRI FR USDA
Fig. 28. Rayonet F8T5 RPR 3000Aฐ spectra cellulose acetate aged 6 hr.
Z = 20 nm
35
-------
S3.
IS
5.67
= 95.1 Zฐ20
2778330,09 fila? 76 S
11 filo# 14 1_
77B9S2o IS f iiofl 15
filฎ# 21
1:15 EDT
01d_Rayonet //I Unfiltered
tfl +
CO CO
' WAVELENGTH nm IRL AMRI ARS USCA
Fig. 29. Rayonet FOT5 RPR 3000Aฐ and sun (Beltsville) spectra
36
-------
ฎ 1
lฃj j[
E
Ul
1 .
4-
277S923.B8
77S^2.11 filel 14 U
77B9S2.10-file# 15 l_oid
2770630.09 filฉ^ 76
^ 4 rsmj umiltered
Old Rayonet #1 unfiltered
4*8.1 !Z~^ 7^5.1 c
95.1 Z = 20 cm ,
h + O.OOSA'n, CA-lOh AE^= 7.43
20 c
1:15 EOT
= 5.67
ID CD
CM (M
C^-
CM
^
CO
CM
O)
OJ
S5 ~* CM
en en en
ea
en
en
en
in
en
co
en
en
WAVELENGTH nm IRL AMRI ARS USDA
Fig. 30. Spectra FS40 and Rayonet B8T5 RPR 3000Aฐ
37
-------
FS_40+Peo
FS40+Bel
Weiahtin
WAVELENGTH nm IRL AMRI FR USDA
Fig. 31. Spectra FS40 and cellulose acetate aged 0.5 hr. Z = 50 cm
38
-------
AM 7. (am)
FI1F
2H1102JZ. FJLE
22M10SJSL FJLEJliiL.
E
CO
UJ
a:
WAVELENGTH nm IRL AMRI FR USDA
Fig. 32- Spectra FS40 and cellulose acetate aged 6 hr Z = 50 cm
39
-------
IOC
2 ; .
u -
o - :
N < :
j: ^- .
0 C
n L
d l'l
IRLSpec-D
B w = 2 n
; -7 7ซ603-20pHi2->|-l 17. u
: 77dfi03rl7DI
-: Window glajsa: ' : j
AE = 0.800J
770&03-14D
4 1- Mylar (0.005")
I ....:..:.
7 r UV :Goggle |(ajnt>er>) N
iTQYOOSl
8 :- UV Goggle j(yellcwlsh) .
60 7o 8Q To
/a 2o
A "h rn
~44 3^0 60 70 80 W
.*
Fig. 33. Spectra - eye protective device
40
-------
IRLSpec-D UV-B
W = Z
i : :- Source f- FS40
Plabtic lens (Nancy) :
Glabs lens (Don)
: :770B03-11D
3-- Glass lens (George)
: --f v770ioP3-;-lDp
: ~5f r-;Refaectlve
i .:(Randy)
60 70 I 80
Fig. 34. Spectra prescription eyeglasses and sunglasses
41
-------
APPENDIX A
Figure
Ala - Ale - Operate Program, IRLSpec-D
A2 - Calibrate Program, IRLSpec-D
A3a - A3b - Operate Program, IRLSpec-S
A4a - A4d - Operate Program, IRLSpec-SO
Spectral Response Curves
AS - Standard Lamp Plotted Each nm
A6 - FS40, FBZS40CLG1076, Sun, IRLSpec-S
A7 - Filtered FS40 and FBZS40CLG1076, IRLSpec-S
A8 - FS40, FBZS40CLG1076, Z = 73 cm, IRLSpec-D
A9 - FBZS40 WLG, FBZS20 WLG, Vitalite, IRLSpec-D
A10 - F15T8 CW Westinghouse, IRLSpec-D
All - F15TO CW Westinghouse, IRLSpec-SO
A12 - F15T8 CW Sylvania, IRLSpec-D
A13 - F15T8 GRO-LUX Sylvania F15T8 WW GE, IRLSpec-D
A14 - F40R GE, F40IR Westinghouse, F1.5T8R GE, Z - 50 cm, IRLSpec-D
A15 - F40IR Westinghouse, Z = 20 cm, IRLSpec-D
A16 - F15T8BL Sylvania, F15T8BLB GE, IRLSpec-D
A17 - F40 BL + FBZS40 CLG1076, IRLSpec-D
A18 - F20T12/2021 Sample No. 3176-2, IRLSpec-D, IRLSpec-SO
A19 - F20BL Phillips phosphor, F4T5BL conventional phospher,
IRLSpec-S
42
-------
fill TO STflRT
fl-IRR. CflL. SCflH
B-REV
C-CflL. ULGTH
D-SCfiH SPECTRfl
E-3TOP
F-F8RWHRB
G-PP.T. DRTfl
I-flCT. INT. 9
K-WLGT>H. CHECK
L-STD. LflHP EQ
M-SUMS
H-COHT. RE flD
ENTER YR.HO.DflYtt
2 7 8 0 i ฃ y . y 0
RESET
_F I 1 E l^-r '
TYPE @
USED 746
MflX . 1060
0 0 0 0 P R H T *:
080.2 fl
0 0 0 3 -
0 0041
0 0 0 5 R
0 0 0 6 R
0007.
0 0 0 3
0 0 O 9 C
0010 fi
0011 L
0012 .
0013
0014 S
0 0 1 5 C
0016 R
0017 H
0018 B
0019 -
0 fi 2 9 R
0021 E
0022 V
0 0 2 3
0024 LINE
i""j i""j -"i C" i"'
' 0027
0 0 2 8
0 0 2 9
0030
0031
0 0 3 2
0 0 3 3
0 0 3 4
0 8 3 5
0 0 3 6
8 0 3 7
0 0 3 8
0 8 3 9
8 8 4 8
8841
0042
0 8 4 3
0 844
fi Ci J S
U U H -_'
0 8 4 6
0 8 4 7
0048
8 8 4 9
8 0 5 8
0051
0 0 5 2
0 0 5 3
0 0 5 4
0055
0 8 5 6
8 8 5 7
8 8 5 8
8 8 5 9
8 0 6 8
0861
8 8 6 2
8 8 6 3
0 0 6 4
0 8 6 5
8 8 6 6 .
8867
8 8 6 8
8 6 6 9
8 0 7 8
8871
8872
8 6 7 3
8874
C
Fi
L
W
L
G
T
H
LI
n
_
!-
f:
H
H
,-.
p
E
C
T
R
fl
U
E
C;
T
n
p
LI
F
-
F
0
R
N
H .
R
n
LI
G
P
R
T
HE
HE
HE
HE
8 U ii 5 U
8826 -
8 6 7 5
6 8 7 6
8877
8 8 7 8
8 8 7 9
8 8 8 8
8 8 8 1
8832
0 0 8 3
0 8 3 4
0 0 3 5
0 0 3 6
0 8 3 7
8 8 8 8
8 8 8 9
8 8 9 8
8 8 9 1
8 0 9 2
0 0 9 3
' 0894
8 8 9 5
8 6 9 6
8 8 9 7
8 8 9 3
8 8 9 9
8 1 8 8
8181
8 1 8 2
8 1 8 3
8184
0105
0 1 0 6
0107
0 1 8 8
8 1 8 9
8118
8111
8112
8 1 1 3
8114
8115
8116
'8117
8 1 1 8
8119
8 1 2 8
8121
8122
8 1 2 3
8124
.
n
H
T
fl
LI HE
I
-
fl
c
T
.
I
H
T
.
Q
LI HE
K
-
W
L
G
T
H
.
c
H
E
c
K
LI HE
L
-
s
T
D
.
L
fl
M
P
E
Q
Fig. Ala. Operate Program IKLSpec-D (Cassette D4)
43
-------
0125 LINE
0126 H
0127 -
0128 S
0 1 2 9 U
0130 H
0131 S
0179 ttREGS
0180 G U T 0 G 6 9 5
0182 CLERR
0183 6
0184 + ฃ-
0185 ENTER!
0 1 8 6 1
0132 LINE 0137 FRMT 4
0 1 3 3 H ! 0 1 8 9 S T 0 P
0134 - f 019F1 LBL
0135 C C
0136 0 ' 0192 LBL
0137 H . C
0 1 3 8 T ; 0 1 9 4 F R N T <::
0139 . i 0196 N
0140 0197.
0141 R 0198 L
0 1 4 2 E . i 0 1 9 9 .
0 1 4 3 H i 0 2 0 U
0144 B f fi201 f:
0145 LINE 0202 H
0 1 4 6 E I 020 3 L
0147 N 0 2 H 4 .
0143 T
0149 E
0150 R
0151
0152 Y
0153 R
0154 .
0155 M
0 1 5 6 0
i 0157 .
0153 D
0159 fl
0160 V
; 0161 #
i 0162 END*
0163 STOP
0164 FIX 2
0 1 6 6 2
0167 EEX
0:68 6
0169 +
0170 S T 0 R 0 ij 0
0172 PRINT
0173 5
0174 MfiSK 4
0176 1
0177 5
0173 1
0 205 E N H *:
0 2 U 6 5
0 2 070
0203 STO F
0209 SFG 2
0210 LBL
---- D
0212 LBL
D
0214 FIX 2
0 216 .
02170
0213 1
0219 S T 0 + R 0 O 0
0 2 2 1 R C L R 0 U 0
0223 PRINT
0224 FIX 4-
0226 1
'0227 S MJ fl
0228 IF SFG 2
0 2 2 9 G 0 T 0 G' 2 3 5
0231 1
0 2 3 2 2
0 2 3 3 0
0234 STO F
02353
0 2 3 6 9
0 2 3 7 S T 0 G
0233 1
0 2 3 9 E N T E R t
O 2 40 0
0241 + $-
0242 LORD
0243 5
0244 NBYTE
0 246 R E R D
0248 EHTERt
0249 4
0 250 + r -
0251 IF X=Y
0 2 5 2 G 0 T 0
0 2 5 4 G 0 T 0
U 2 5 6 3
0257 WBYTE
O 2 5 9 F 0 R
0 2 6 0 1
0 2 6 1 S T 0
0262 RERD
0264 IF 0
O 2 6 5 L 0 T 0
O 2 6 7 G 0 T 0
0269 X ฃ Y
0270 IF S F G
0271 G 0 T 0
O 2 7 3 S T U
0274 6
0275 WBYTE
0277 FOR
0278 RERD
0 2 3 0 X ^ Y
0 2 3 1 S T 0 +
0232 NEXT
0233 3
0284 WBYTE
0236 RCL
0287 RCL I
0 2 3 9 -
0290 1 0 1 X
. 0291 STO I
0293 CFG
0294 RCL
0 2 9 5 2
0 2 9 6 4
0297 9
0 2 9 8 +
0299 ENTERt
0 3 U 0 1
0301 EEX
0302 3
(-
r
l-
r.
^
r
f
j
r
(
J
t
t
r
E.
4
i_
h
H
4
R
Fig. Alb. Operate Program IRLSpec-D (Cassette D4) (cont'd)
44
-------
.i_-< *^ ...,,'ปa*aฃซ.,-i
0303 *
0 3 04 +
0305 PRINT -
0306 IF CFG 3
0 3 0 7 G 0 T 0 0 3 1 6
0 3 O 9 .
0 3 100
0311 1
0 3 1 2 S T 0 - R O 0 U
0314 G 0 T 0 0317
0316 NEXT R
0317 1
0313 WBYTE 4
0320 CFG 3
0321 RCL fi
0 3 2 2 8
0 3 2 3 U
0324 -
0325 IF +
0326 GOTO 1
0323 IF SFG 2
0329 GOTO K
0331 RETURN
0332 LBL
F
0334 3
0335 WBYTE 4
0337 RETURN
0338 LBL
p
0340 CLEfiR
0341 WBYTE 4
0343- RETURN
0344 LBL
B
0346 1
0347 WBYTE 4
0349 1
0350 RETURN
0351 LBL
I
0353 PRNT*
0 3 5 5 I
0356 N
0357 T
0 3 53.
0 3 5 9 9
0 3 6 0
0361 F
O 3 6 2 F:
0 3 6 3 0
.0364 N
0365 END*
0 3 6 6 2
0 3 6 7 8
0 3 6 3 0
0 3 6 9 S T 0 P
0370 F I X 0
0372 PRINT
0373 ENTERt
0374 2
0375 4
O 3 7 6 9
0 377 -
0 3 7 8 S T 0 R
6 3 7 9 8
0 3 8 O 0
0 3 8 1 S T 0 F
0332 CLERR
O 3 8 3 S TO. I
O :I 3 4 F 0 R fi * F
0C-85 RCL fi
O :I 8 6 2
0 :1 8 7 4
0 '~i 3 3 9
0 3 3 9 +
0 3 9 U 2
0391 2
0 3 9 2 8
0393 .
0 3 9 4 1
0 3 957
O ''- 9 6 8
0 :] 9 7 -
0 :: 9 8 9
U :: 9 9 Y t X
0 4 0 0 S T 0 H
0401 4
O 402 -i-
0 4 0 3 4
0404 Y t X
0 4 0 5 S T 0 . J
0406 RCL H
04074
0 4 0 8 -
0 409 + ฃ -
0410 e t X
0411 R C L J
0412 *
0413 RCL I 'R
0415 *
0416 R C C +
0417 NEXT R
0413 RCL I
0419 FIX 4
0421 PRINT
0422- ENTERt
042 3 3
0424 .
0 4 2 5 0
0 4 2 6 6
0427 r
0423 PRNT*
O 4 3 0 S
0 4 3 1 U
0432 N
0433 3
0434
0435 =
0436 PRINT
0437 END*
0 4 3 8 G 0 T 0 H
0440 LBL
fi
0442 .
044 3 0
0444 1
0445 3 T 0 + R 0 0 O
0447 FIX 2
0449 RCL ' R000
0451 P R I N T
0452 FIX 3
0454 1
0455 STO H
0456 1
0457 5
045 3 0
0 4 5 9 S T 0 F
0460 4
O 4 6 1 0
0462 3 T 0 G
0 4 6 3 1
0464 S T 0 B
0 4 6 5 5
0466 WBYTE 4
0463 REflB 4
0470 ENTERt
0471 4
0472 + i -
0473 IF X=Y
0474 G 0 T 0 047 8
0 4 7 6 G 0 T 0 0
0473 3
0479 WBYTE 4
O 431 F 0 R H -* F
Fig. Ale. Operate Program IRLSpec-D (Cassette D4) (cont'd)
t
45
-------
0432 RERD 4
(3484 IF 0
6 4 3 5 G 0 T 0 0 4 8 9
0 4 3 7 G 0 T 0 0
0 4 8 9 X ฃ V
0490 IF SFG 4
0491 G 0 T 0 0 5 3 0
0 4 9 3 C L. ERR
0494 8 T 0 C
0 4 9 5 6
0496 MBYTE 4
0493 FOR B + G
0 4 9 9 R E R D 4
0501 X ? Y
0502 S T 0 + C
0503 NEXT B
0 5 0 4 3
0505 WBYTE 4
0 5 0 7 1
0 5 0 8 S T 0 B
0 5 0 9 R C L C
0510 STO+ 1 R
0512 + ?-
0513 RCL fl
0514 2
0515 4
' 051 6 9
0517 +
0513 EHTERt
0519 EEX
05204
0 521 #
0522 +
0523 PRINT
0524 IF SFG 8
0 5 2 5 G 0 T d 0 5 4 1
0527 NEXT fl
O 5 2 8 G 0 TO 0541
0530 PRNTo:
0532 0
0533 ','
0534 E
0535 R
0536 L
0537 0
0538 fl
0539 D
0540 EH Do:
0541 1
0542 NBYTE 4
0544 CFG 3
0545 CFG 4
0546 RETURN
0547 LBL
L
0549 LBL
L
0551 CLEfiR
0552 1
0553 L H & G 0
0554 LBL
i/
r\
0556 RCL R047
0553 RCL R043
0 5 6 0 H-
0561 LOG
0562 EHTERt
0 5 6 3 4
0564 *
0565 +?-
O 5 6 6 2
0 5 6 7 9
O 5 6 8 6
0 5 69.
0 5 7 0 5
0571 +
0572 FIX 2
0574 PRHTo:
0576 LINE
0577 L
0578 I
0579 N
0530 E
0581
0 5 3 2 R
0583 T
0534 PRINT
05S5 EN Do:
0536 F I X 4
0583 CFG 2
0539 3TOP
0590 LBL
H
0592 PRNTo:
0594 3
0595 U
0596 H
0597
0598 F
O 5 9 9 R
0 6 0 0 0
0 6 O 1 M
0 6 0 2
0603 EH Do:
0 6 0 4 3 T 0 F'
0605 F I X
0607 PRINT
0 6 0 3 2
0 6 0 9 4
0 6 1 O 9
0611 -
0612 S T 0
0613 PRNTo:
0615 T
0 6 1 6 0
0617
0613 EN Do:
0619 STOP
0620 PRINT
0621 2
0 6 2 2 4
0 6 2 3 9
0624 -
0 6 2 5 S T 0
0626 CLERR
0627 S T G
O 6 2 3 F 0 R
0629 RCL
O 6 3 0 2
0631 4 .
0 6 3 2 9
0 6 3 3 -
0634 RCL I
0 6 3 6 S T 0 +
0637 NEXT
0638 FIX
O 6 4 0 R C L
0641 P R H T o:
0643 M
0644 I
0645 L
0646 L
0647 I
0643 W
0649 R
0 6 5 0 T
0651 T
0652 3
0653 /
0654 M
U 6 5 5 S
0 6 5 6 Q
0657 PRINT
0658 ENDo:
Fig. Aid. Operate Program IRLSpec-D (Cassette D4) (cont'd)
46 ' '
\'
Fi
F
i
h
R
H
I
H
1
1
i
r ~
-------
6bby
8 6 6 1
8 6 6 3
8 6 6 4
8 6 6 5
8 6 6 6
6 6 6 8
6 6 6 9
8 6 7 8
8671
8672
6673
8674
8 6 7 6
6677
6678
6679
8 6 3 6
6681
6 6 8 2
8 6 8 3
8 6 8 4
8 6 8 6
8 6 8 8
8 6 9 6
8 6 9 1
8692
8693
8 6 9 5
8 6 9 6
8 6 9 8
8 6 9 9
. 8781
ij U T U
LBL '
N
CLEfiR
STO
4
MBYTE
4
6
STO
1
STO
FOR
REfiD
' ^ V
S T 0 +
NEXT
RCL
PfiUSE
PfiUSE
C L E ft R
STO
G U T 0
LBL
U
LBL
u
B
8
i
STO-
CLEfiR
MBYTE
1
MBYTE
PRNTv:
H
c
4
G
B
B*G
4
C
B
c
c
6671
R 0 8 8
4
, 4
o 7 y o
8784
8 7 6 5
6 7 6 6
8787
8788
6 7 8 9
671 1
6713
6715
6717
6719
6 728
6721
8722
6723
6724
6725
872 6
672 3
8729
6731
8 7 3 2
6733
6734
8735
6736
8737
6 7 3 3
8 7 3 9
6746
8741
8742
674 3
6744
8745
R
E
O
E
T
EN Do:
.GOTO 6182
LBL
G
LBL
L,
R C L R 8 6 8
F I X 2
PRINT
1
STO fi
1
ฃ
M
STO F
FIX 4
FOR fi + F
RCL I fi
RCL H
i
4
4
+
ENTERt
1
EEX
.Ji
*
+
PRINT
NEXT fi
STOP
END
Fig. Ale. Operate Program IRLSpec--D (Cassette D4) (cont'd)
47
-------
FILE I
YYPฃ i>
USED 218
f'l fl x s e 0
0880 PRHT'M
0 0 0 2 C
0 0 U 3 fl
0 004 L
0005 I
0 0 0 6 E:
0 0 0 7 R
0 0 0 S fl
0 0 0 9 T
0010 I
0 0 1 i u
0912 N
0013 LINE
0014 C
0 0 5 1
3 0 5 2
0 0 5 3 -
0 0 5 4 0
0 0 5 5
0 0 5 6 fl
0057 H .
0 o 5 s n
0059
0 0 6 0 -
0 061 1
0062 LI HE
0063 LINE
0064 LINE
0 0 6 5
0 0 6 6
U U 6 7 N
U U 6 8 0
0069 .
0 0 7 0
i _ _ .
0015 0 ! 0071 R
t
0 0 1 R [ 0 0 7 2 U
0 0 1 7 R ! 0 0 7 3 N
0013 E
0 0 1 9 C
0020 T
0021 I
0022 0
0023 N
0 0 2 4
0025 W
0 0 2 6 I
0027 L
0 0 2 3 L
0029 LINE
0 0 3 0 B
0031 E
0 0 3 2
0 0 3 3 R
0034 E '
O 0 3 5 C
0 O 3 6 0
0037 R
0 0 3 S B
0039 E
0 0 4 0 L
0041
0 O 4 2 0
0043 H
0044 LINE
0045
0 O 4 6
0047 F
0 0 4 3 I
0 0 4 9 L
0 0 5 0 E
0074 S
0075 =
0 0 7 6 '?
0077 EN Do:
0 0 7 8 3 T 0 p
O 0 7 9 S T 0 E
0080 1
0 0 S 1 S T 0 fl
0 0 8 2 1
0 0 S 3 5
0084 0 I
0 1 0 5 2
.0106 1
0107 4
0 1 0 3 S T 0 L'
0 1 0 9 .
0 1 1 0 2
0111 1
0112 4
0 1 1 3 6
0114 S T 0 H
0115 FOR fl*F
0116 RCL fl
0117 2
01184
0119 9
0 120 +
0121 RCL I
0122 *
0123 RCL .i
0124 +
0125 10tX
0126+?-
0127 RCL B
0123 +
0129 16t X
n 1 :-: w :-; T n f
0131 RCL fl
0 1 3 2 2
0133 4
O 1 3 4 9
0135 +
0136 RCL D
0137 -
0161 ~
0162 FTi
- - " '. L
0163 LC-:
0164 -
0165 3Tf
fi 1 6 7 P p i ,
- * - 1 I T ;
0 1 6 3 HF'
' i L.
0 169 1
0170 c,
0171 0
0172 Etif.
0 1 7 3 1
0174 El.'!''
01750
0176 + ? -
0177 RCIi;-
0 178 1
0 179 5
0 1 8 0 0
0181 EHTE'
0 1 8 2 1
0133 EHTE;
0 1 S 4 1
0135 + ? -
0186 RCBiV
fi 1 8 "' P P N T
- A - i 1 1 . 1 1 !
0 1 8 9 C
0 1 9 0 H
0 1 9 1 L '
0192. .
0 1 9 3
O 1 9 4 C:
0 1 9 S n
0133 . 0196 N
0035 STO Fl 0139 5 019? s
0 0 36. 16146* '101 9 R T
0 0 8 7 0
0 0 8 8 0
0 0 3 9 3
0 0 9 O 5
0 091 + v ~
0092 STO I
0093 1
0 094 .
0 0 9 5 3
0096 .1
0 097 7
0 0 9 3 3 T 0 J
0 0 9 9 2
0 100 .
0101 3
0102 7
U 1 0 3 6
* 1 * 4 3 T U B
0141 1/X
0142 STO- f:
0143 RCL fl
0144 3
0145 1
0146 -
0147 RCL H
0148 *
0149 EHTERt
0150*
0151 + ? -
0152 8 1 X
0 1 5 3 .
0154 1
0155 + ฃ -
0156 *
0157 S T 0 + C
0153 RCL I fl
0160 RCL E
0 1 9 9 .
0 2 0 0
0 201 I
0202 N
0203 LINE
U 2 0 4 1
O 2 0 5
0 2 0 6 T
0 2 O 7 0
0 2 0 8
0 2 0 9 1
O 210 5
0211 0
0212 E H D :
0213 CLEH?
0214 S T 0
0215 CLEfiF
0216 L D i. b i'
0217 END
Fig. A2. Calibrate Program IRLSpec-D
48
(Cassette D2)
-------
DRTE '
E UV SPECTRR
1780210.01
OPERATE PROGRAM IRLSpec-S
0000
0001
0002
0003
OOOU
0005
0006
0007
0003
0009
0010
0011
0012
0013
001U
0015
0016
0017
0018
0019
0020
0021
0022
0023
00 2 U
0025
0026
0027
0028
0029
0030
0031
00^2
0033
OO 3 U
0035
0036
0037
0038
0039
OOUO
OOUl
OOU2
OOU3
00 UU
OOU5
00 U 6
OOU7
OOU8
yyus
0050
CHR
D
R
T
E
CHR
E
+
1
EXP
p,
SM
10
E
, .r-i
bP
8 a
3
"7
0
SM
06
FLG
10
0
SM
00
1
U
0
SM
01
2
5
O
SM
02
f9
106
CHR
2
CHR
102
m
0
1
EM
10
r~iM
RM
10
0051
0052
0053
005U
0055
0056
0057
0058
0059
0060
0061
0062
0063
006U
0065
0066
0067
0068
0069
0070
0071
0072
0073
007U
0075
0076
0077
0078
0079
0080
0081
0082
0083
008U
0085
0086
0087
0088
0089
0090
0091
0092
0093
OO 9 U
0095
0096
0097
0093
0099
0100
CHR
IJ
V
.-
P
E
c
T
R
fl
CHR
FIX5
02
LUL
10
CHR
W
L
I
R
R
R
D
W
L
I
R
R
ft
D
CHfi
01
106
CHR
LHH
0101
0102
0103
01 OU
0105
0106
0107
0108
0109
0110
0111
0112
0113
011U
0115
tvcl "1 C
* JL -LC1
0117
0118
0119
0120
0121
0122
0123
012U
0125
0126
0127
0128
0129
013y
0131
0132
0133
013U
0135
0136
0137
0138
0139
01 UO
01U1
01U2
01U3
01UU
01 U5
01U6
01U7
01U8
01U9
-A
101
FLG
fifi
IFER
01
106
101
~
RM
11
L
IND
RM
00
~
iex
L
IND
SM
01
SM
fi-:
102
RM
02
FIX5
fif-l
COL
0-:
RM
03
FIX5
03
COL
08
?
01
1
EM
02
RM
02
FIX5
M0
COL
03
1 06
101
0151
0152
0153
015U
0155
0156
0157
0158
0159
0160
0161
0162
0163
016U
0165
yit't
0167
0168
0169
0170
0171
0172
0173
017U
0175
0176
0177
0178
0179
0180
0181
0182
0133
013U
0185
0186
0187
0188
0189
0190
0191
0192
0193
019U
0195
0196
0197
0198
0199
-'
RM
11
R
IND
RM
00
=
10"
R
IND
SM
01
102
FIX5
y '
COL '
08
>
01
1
EM
OM
EM
01
EM
02
RM
02
RM
06
IF-
00
FLG
01
106
CHR
0
CHR
102
LF
RM
02
RM
07
ซ
T P -4-
1 r ~
0201
0202
0203
02 OU
0205
0206
0207
0208
0209
0210
0211
0212
0213
021U
0215
0216
0217
0218
0219
0220
0221
0222
0223
022U
0225
0226
0227
0228
0229
0230
0231
0232
0233
023U
0236
PP-17
0238
0239
02UO
02U1
02U2
02U3
02UU
02U5
02U6
02U7
02U8
02U9
0250
OU
SP
8b
71
R
RM
Ul
L
RM
U2
LOG
x
1
=
SC:
7O
RM
1&
71
. i j.
'f 1X5
02
COL
07
L
RM
63
R
RM \
63
LOG
;..-
^
_
SC
70
RM
1U
_
FIX5
02
COL
07
LF
Fig. A3a
49
-------
OPERATE PROGRAM IRLSpec-S (cont'd)
0251 E
0252 FLG
0253 00
0250 SP
0255 8C
0256 CM
0257 02
0258 CHfl
fi^cjq .-
fi^fil
| T
1 1
13
0262 R
0263 T
flap's
0266
0267
FPF '3
0270
0271
0272
0273
0270
0275
0276
0277
0278
0279
0280
0281
0282
0283
0280
0285
0286
0287
0288
0289
0290
fisqi
0292
0293
0290
0295
0296
0297
0298
0299
0300
T
1
N
T
1
q
H~
1
<~-
CHfl
E
IFE
12
-X.l_
a
8
0
0
FLG
12
1
f
0
2_
^,
2
SM
01
1
0
0
SM
00
0301
0302
0303
0J00
0305
0306
0307
0308
0309
0310
0311
0312
0313
0310
0315
0J16
0317
0318
0319
0326
0321
0322
0323
0320
0325
0326
0327
0328
0329
0330
0331
0332
0333
0330
0335
0JJ6
0337
0338
0339
0300
0301
0302
0303
0300
0305
0306
0307
0308
0309
0350
0351
FLG
02
L
IND
RM
00
','.
L
IND
RM
01
=
EM
02
R
IND
RM
00
X
R
IND
RM
01
=
EM
02
1
EM
00
EM
01
RM
01
-
7
9
=
IF-
02
RM
02
FIX5
0J
COL
08
CHfl
s
l.l
N
9
' 0352 CHfl
0353 -
0350 RM
0355 15
0356 =
0357 FIX5
0358 05
0359
0360
0361
0362
0363
0360
0365
0366
0367
0368
0369
0370
0371
0372
0^73
0370
0375
0376
0377
0378
0379
0380
0381
0382
0383
0380
0385
0386
0387
0388
0389
0390
0391
0392
0393
0390
0395
0396
0397
0398
0399
0000
COL
16
LF
SP
Sd
FLG
07
CHfl
S
IJ
M
F
R
o
M
CHfi
E
SM
03
CHfl
T
o
CHfl
E
SM
00
CM
02
RM
03
-r
2
+
1
5
:=
SM
03
FLG
03
L
0001
0002
0003
0000
0005
0006
0007
0008
0009
0010
0011
0012
0013
0010
0015
0016
0017
0018
0019
0020
0021
0022
0023
0020
0025
0026
0027
0028
0029
0030
0031
0032
0033
0030
0035
0036
0037
00^8
0039
0000
0001
0002
0003
0000
0005
0006
0007
0008
0009
0050
IND
RM
03
EM
02
R
IND
RM
03
EM
02
1
SM
03
RM
03
1
5
zz
v
2
_
RM
00
:=
IF-
03
LF
RM
02
FIX5
03
?}
GT
07
SP
8e
3
0
0
SM
06
GT
10
CHfl
M
E
M
0051
0052
0053
0050
0055
0056
0057
... | f- f.
U058
0059
0060
0061
0062
0063
0060
0065
0066
0067
0068
0069
0070
0071
0072
0073
0070
0075 '
0076
0077
0078
0079
0080
0081
0082
0083
0080
0085
0086
0087
0088
0089
0090
0091
0092
0093
0090
0095
0096
0097
0098
0099
0500
F
R
0
M
CHH
LF
E
SM
08
CHH
T
Q
CHH
E
LF
SM
09
FLG
11
L
IND
RM
08
FIX5.
03
COL
12
R
IND
RM
08
FIX5
03
COL
10
LF
1
EM
08
RM
08
RM
09
=
IF-
11
EP
00
Fig. A3b
50
-------
flUTO STflRT
H SPECTRfl
B REVERSE
C S L CflL.
E STOP
F FQRWfiRD
G PRINT DflTfl
I INTEGRflL
J RUN WLGTH
K WLGTH EQUfiT.
L STB LHMF EQUfi
M SUN
0 OPERflTE
' ENTER YR.MQ.DRYtt
3780120. 00
REflBY
.HIT fl C OR J
FILE 0
.- TYPE U
'USED 720
HflX 300
0000 PRNTo:
0002 fl
0 0 O 3
0 O 0 4 S
0 0 0 5 P
0 0 0 6 F
r_' V V ' ' U_
0 U 0 7 C
0003 T
0 0 0 9 R
0010 fl
0011 LINE
0012 B
0013
0014 R
0015 E
0016 V
0017 E
0013 R
0019 S
0 O 2 0 E
0021
0022 LINE
0 0 2 3 C
0 0 2 4
0 0 2 5 S
0026
U027 L
0028
0029 C
: 0 0 3 0 fl
0 O 3 1 L
O 0 3 2 .
0 0 3 3 LINE
0 0 3 4 E
0035
0 0 3 6 S
0 0 3 7 T
0 O 3 8 0
0039 P
0040 LINE
0041 F :
0 0 4 2
0 O 4 3 F
0044 0
0045 R
0 0 4 6 W
0047 fl
0 0 4 8 R
0 0 4 9 B
0050 LINE
0 0 5 1 G
0052
0 0 5 3 P
0054 R
0055 I
0 0 5 6 N
0057 T
0 0 5 8
O 0 5 9 B
0060 fl
0061 T
0062 fl
0063 LINE
0064 I
0 0 6 5
0 0 6 6 I
0067 N
0068 T
0 0 69 E
0 0 7 M G
0071 R
0072 fl
0073 L
0074 LINE
0 0 7 5 J
0 0 7 6
0 077 R
0073 U
0079 N
0 0 3 0
0031 W
0 0 8 2 L
0 0 8 3 G
O O 8 4 T
O 0 8 5 H
0 O 3 6 LINE
O 0 3 7 K
U 0 g 8
fi o K '3 u
W %' ' ป' H
0 0 9 U L
0 0 9 1 G
0 092 T
0093 H
L~1 L"1 Q 4.
U U y T"
0095 E
O O 9 6 Q
0097 U
O 0 9 8 fl
O 0 9 9 T
0 100 .
0101 LINE
0102 L
0 1 0 3
M 1 pi 4 '-:
v 4 v i '
0105 T
0106 B
0107
0103 L
0109 fl
0110 H
0111 P
0112
0 1 1 3 E
0114 Q
0115 U
0 116 fl
0117 LINE
0 1 1 8 M
0119
0120 S
0121 H
0122 M
0123 LINE
0124 0
,-> 4 .-, cr
0 1 ฃ. -'
0126 0
0127 P
0123 E
0129 R
0130 fl
0131 T
0132 E
0133
0134 LINE
0135 E
0136 N
0137 T
0138 E
0139 R
0140
0141 Y
0142 R
0 143 .
13144 M
0145 0
0146 .
0147 B
0143 fl
0149 Y
0150 tt
0151 LINE
0152 END*
015:': STOP
0154 3
0155 EEX
0156 6
0157 +
0153 FIX
0160 PRINT
0161 STO
0163 2
0164 5
0165 4
0 1 6 6 0
0167 S T U
0168 2
0169 5
0170 1
0171 O
0172 STO
. 0173 FIX
Fig. A4a. Operate Program IRLSpec-SO
51 '
(Cassette B)
R 0 0 0
-------
6175 1
i~i 4 ~~* "* "~
0 1 r' b D
0177 1
0173 #REGS
0179 LBL
0
0131 LBL
0
0 1 3 3 C F G 3
9134 CFG 3
0185 F'RHTo:
0137 R
0133 E
0139 R
0 1 9 0 D
0191 Y
0192 LINE
.- '. 4 ,"! . | 1 1
u i y J H
0194 I
0195 T
0 1 9 6
0 197 ft
0 1 9 3
0 1 9 9 C
0 2 0 0
0 2 0 1 0
0202 R
0203
0204 J
0205 LINE
0206 EN Do:
0 2 0 7 S T 0 p
0203 LBL
0210 LBL
ft
,-j .-, J -, 4
tlili 1
0213 ENTERt
0214 HNDSK 4
U 2 1 1" 2
0217 WBYTE 4
0219 1
0220 ENTERt
0'"' O 1 ivf
_ ii- tZ. 1 U
0222 +r-
0 2 2 3 L 0 H D
0224 FIX 2
0226 .
0227 0
0223 1
U 2 2 9 S T 0 +
0231 RCL
0233 PRINT
0234 FIX
0236 3
0237 WBYTE
0 2 3 9 1
0 2 4 0 S T U
0241 IF SFG
0242 G 0 T 0
0244 1
0 2 4 5 2
0246 0
0247 3 TO
024 3 F 0 R
0249 STRRT
0251 REflD
0253 1
0254 EEX
0255 3
0256 +
0257 STO- I
0259 RCL I
0261 + *-
0262 10tX
0263 STO I
0265 RCL
0 266 -
0267 IF -
' 0263 CLEflR
0 2 6 9 R C L
0 2 7 0 2
0271 4
0 2 729
0273 +
0274 ENTERt
0275 1
0276 EEX
0277 3
0 2 7 3 *
0279 +
0230 PRINT
0231 IF SFG
0232 GOTO
0234 NEXT
0285 IF SFG
0 2 3 6 G 0 T 0
0288 G OS UB
R00t
R 0 0 t
4
4
H
4
024
F
fi + F
4
4
ft
ft
H
E
fl
.-.
B
fl
4
K
B '
I
I
-1 0290 LBL
j i
0292 LBL
j
0294 PRNTo:
0296 I
0297 N
0293 T
0 2 9 9 .
0 3 0 0
0 3 U 1 F
0 3 0 2 R
0 3 0 3 0
0 3 0 4 M
U 3 O 5
0 3 0 6 ?
0307 EN Do:
O 3 0 8 2
0 3 0 9 3
0 3 100
0 3 11 S T U P
0312 FIX fi
0314 PRINT
0315 FIX 4
0317 ENTERt
0 3 1 3 2
0 3 1 9 4
0 3 2 0 9
fi "'. '' 1 -
V '-' 1 1
0322 STO ft
fl 3 '"' '! ft
'-' _' L. _' '_'
H '"; v 4 C\
T ' tl_ j '_>
0325 STO- F
0326 CLEflR
0 3 2 7 S T 0 I
0328 FOR ft*
0329 RCL ft
0 3 3 0 2
0331 4
0 3 3 2 9
0 3 3 3 +
fl ':; -; 4 ?
1J -' _' *T i^
0 3 3 5 2
O 3 3 6 8
0 3 3 7
0 3 3 ft 1
'-' -' -' '.' i
0 3 3 9 7
0 3 4 0 3
0 341 -H
0342 --.
0343 V,
0344 XT-
0345 4
0346 r
0347 4
0343 rr
0349 STi<
0350 pfL"
0351 4
O 3 5 2 -
0353 + = -
0354 ei:1
0355 RCL
0 3 56 *
0357 RCL
0 3 5 9 *
0360 ft CO
0361 NEXI
0362 RCL
0363 PRUT
0365 LINE
0 3 6 6 I
0367 N
0363 T
O 3 6 9 E
0370 G
0371 R
0372 ft
0373 L
0374
0375 9
0376
0 3 77 =
0378 PR HIT
F 0379 LINE
0330 LIME
0381 END'-
0332 EHTEP
0383 3-
0384 .
O 3 8 5 0
0 3 8 6 6
0 3 8 7 +
0338 PRNT*
0390 S
0 3 9 1 U
0392 N
0 3 9 3 S
Fig. A4b. Operate Program IRLSpec-SO (Cassette B) (cont'd)
52
-------
1
I
8394
8 3 9 5
8 3 9 6
8 3 9 7
0393
0399
0 4 0 0
6401
0 4 0 3
0405
8487
8 4 8 9
041 1
0412
0 4 1 3
0414
8415
8416
6417
0418
8419
0420
0421
0422
0423
0424
0425
0427
8428
0 4 3 0
0432
0433
0434
0435
8 4 3 6
8437
0433
8439
8 448
6441
8442
644 3
8444
6445
8446
6447
6449
=
PRINT
LINE
LINE
EHD*
0 4 5 0
U 4 5 2
0454
'0 4 5 6
'0 4 5 8
GOTO H 6459
LBL
K
LBL
K
R C L R 6 6 4
RCL R685
-^
L 0 G
+ i-
ENTERt
~i
.
4
*
-i
5
~i
t
5
+
F I X 2
PRINT
RCL R647
RCL R648
r
LOG
+ ฃ-
ENTERt
~i
p
IT
*
2
C|
0 4 6 0
0461
0 4 6 2
0464
0 4 6 6
0467
0 469
0476
0472
0473
8475
8476
6473
8 4 7 9
8431
6 4 8 2
8 4 8 4
8 4 8 6
6 4 8 7
6 4 8 3
8 4 8 9
8491
8 4 9 3
6 4 9 5
8 4 9 6
6497
0 4 9 8
0 4 9 9
0 5 8 8
6561
6 ; 0502
0503
5 '0504
+ 0 5 0 5
PRINT
FIX 4
CFG 4
8 5 8 7
0 5 0 3
0510
GOSUB
G 0 T 0
LBL
..1
LBL
.J
5
0
S T 0
SFG
G 0 T 0
LBL
F
2
WBYTE
.-J
WBYTE
B
0
F
4
R
4
8511
8512
. 0513
0514
0 5 1 6
0517
0513
0 5 1 9
0 5 2 0
0522
052 3
0524
0525
0 5 2 6
0527
052 3
4 i 8 5 2 9
RET U R H i 0 5 3 0
LBL
E
C.
WBYTE
C L E R R
WBYTE
0531
i ' 0532
', 0 5 3 3
4 ; 0534
O 5 '3 6
4 8 5 3 7
RETURN 0538
LBL
C
LBL
f:
.
8
1
STO +
RCL
F I X
PRINT
1
STO
1
5
6
S T 0
SFG
1
ENTER
HHDSK
~,
WBYTE
PflUSE
! 8 5 4 8
6541
:
i 854 3
i
i 6545
6546
R 6 0 6 0547
R 0 U 0
2
R
F
~i
T
4
4
054 9
0 5 5 0
0552
055 3
0554
0 5 5 6
0 5 5 7
0 5 5 8
6 5 6 6
0 5 6 1
0 5 6 2
0 5 6 4
0 5 6 6
0563
0 5 6 9
0 5 7 0
PflUSE
PflUSE
F 0 R
RERD
1
EEX
3
-H
STO+ I
+ ฃ-
RCL
L.
4
9
+
ENTERt
1
E E X
-Ji
*
+
F I X
F' R I H T
IF SFG
G 0 T 0
NEXT
LBL '
B
LBL
B
1
E H T E R T-
HHDSK
ฃ
WBYTE
PflUSE
0
W E: Y T E
PR USE
1
WBYTE
PflUSE
5
WBYT'E
REflD
REflD
RCL
+
IF -
fl
4
R
R
4
8
B
R
4
4
4
4
4
4
4
B
Fig. A4c. Operate Program IRLSpec-SO (Cassette B) (cont'd)
53
-------
0 5 7 1 G 0 T 0 6 5 6 6
'"1 5 7 ::! 1
'. '..' T '
'.' '. '.
y i ,' <: K E R Ii 4
0573 RCL D
0579 +
0580 IF -
0531 GOTO 0576
0583 0
0534 MBYTE 4
0536 IF SFG 3
0587 STOP
0588 IF SFG S
0 5 3 9 G 0 T 0 0712
0591 RETURN
0592 LBL
M
0594 LBL
M
0596 PRNTo:
0598 R
0599 E
0 6 0 0 C
0601 0
0602 R
0603 D
0604 ? :
0 6 0 5
0 6 0 6 0
0607 R
0603 LINE
0 6 0 9 S
0610 U
06 if M
0612
0613 F
0614 R
0 6 1 5 0
0616 M
.: 0617 EN Bo:
0613 CLEflR
0619 STOP
0620 IF 0
' 0 6 2 1 G 0 T 0 0
0623 FIX O
0625 PRINT
0626 2 f 0K76 LBL
062? 4 ; - -,
f f~ ' "' ' '
--*-
G 6 2 9 -
0 6 3 0 S T H R
0631 PRNTtf
' 0633 T
0 6 3 4 0
0635 EN Bo:
0636 STOP
0637 PRINT
0638 2
0639 4
0 6 4 y 9
0641 -
0.642 STO F
0643 CLEflR
0644 STO I
0645 FOR fl+F
0646 RCL I fl
0 6 4 8 S TO-*- I
0649 NEXT R
0650 FIX 4
0652 RCL I
0653 PRNTo:
0655 M
0656 I
0657 L
0658 L
0659 I
0660 N
0661 fl
O 6 6 2 T
0 6 6 3 T
O 6 6 4 S
O 6 6 5 /
0666 M
O 6 6 7 S
f} j-r ".- ;.-.; i r-
cT
0680 1
0681 STO
0682 1
0-': 3 5
0V.'-! O
0 6 0 ':< S T 0
0636 FIX
0 6 3 3 F 0 R
0689 RCL I
0 6 9 1 R C L
0692 2
O 6 9 3 4
0694 9
0 695 +
0696 ENTER!
0697 1
O 6 98 E E X
0 6 '" "
0 7 0 y *
0701 + .
0732 PRINT
0703 NEXT
0704 S T 0 P
0705 LBL
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FINAL REPORT
BIOLOGICAL EFFECTS OF ULTRAVIOLET RADIATION
ON. PLANT GROWTH AND FUNCTION
M. N. Christiansen
Plant Stress Laboratory
Plant Physiology Institute
Beltsville Agricultural Research Center
Beltsville, Maryland 20705
3t c
ป ^
EPA-IAG-D6-0168
Project Officer:
R. J. McCracken
Agricultural Research, Science and Education Administration
U.S. Department of Agriculture
Washington, D.C. 20250
Prepared for
Environmental Protection Agency
BACER Program
Washington, D.C. 20460
-------
FINAL REPORT
DIFFERENTIAL SENSITIVITY OF TWO CULTIVARS OF
CUCUMBER (CUCUMIS SATIVUS L.) TO INCREASED
UV-B IRRADIANCE:
I. DOSE-RESPONSE STUDIES
D. T. Krizek
Plant Stress Laboratory
Plant Physiology Institute
Beltsville Agricultural Research Center
Beltsville, Maryland 20705
EPA-IAG-D6-0168
Project Officer:
R. J. McCracken
Agricultural Research, Science and Education Administration
U.S. Department of Agriculture
Washington, D.C. 20250
Prepared for
Environmental Protection Agency
BACER Program
Washington, D.C. 20460
-------
CONTENTS
Abstract iii
Tables iv
Figures v
Acknowledgments vii
Introduction 1
Materials and Methods 3
Plant Material 3
Cultural Conditions 3
UV Source 3
UV Measurements 4
Harvest and Data Analyses 5
Results and Discussion 7
Comparative Phytotoxic Effects of UV-B
Irradiation 7
Comparative Influence of UV-B Irradiation
on Vegetative Growth 8
Conclusions 1.1
Literature Cited 12
-------
ABSTRACT
Dose-response studies were conducted at Beltsville, Maryland, on
two cultivars of cucumber (Cucumis sativus L.) exposed to a UV-B irradiance
gradient representing an increase of 40% to 770% in biologically
effective UV (BUV) radiation over normal sunlight. Plants were irradiated
in a fiberglass greenhouse in April employing FS-40 fluorescent sunlamps
filtered with 0.127 mm Mylar (UV-A) or 0.127 mm cellulose acetate
(UV A&B). UV treatment was given 6 hours per day (from 1000 to 1600)
for 19 days from the time of seeding. Marked differences in UV-B
sensitivity were observed between 'Poinsett1 (extremely sensitive) and
'Ashley' (slightly sensitive). Increasing the UV-B level induced
chlorosis of the leaves, inhibited leaf and shoot growth, and reduced
biomass. These effects were especially marked in 'Poinsett'. 'Ashley'
plants required approximately twice the level of BUV as 'Poinsett' to
exhibit a 20-25% reduction in dry weight or leaf area. Based on linear
regression analysis of the 'Poinsett' data, it x^as estimated that a
maximum proposed decrease in stratospheric ozone content of 20% (or a
40% increase in BUV) would cause a 10% reduction in dry matter production
with a 15% decrease in leaf area. A 100% increase in BUV or greater
t
was needed to cause pronounced chlorosis of the leaves and a marked
*
reduction in dry matter production. Such increases would be far in
excess of the projected BUV levels expected as a result of stratospheric
ozone reduction caused by chlorofluoromethane emissions.
iii
-------
TABLES ' Page
Table 1. Weighted and unweighted UV spectral irradiance under each of 16
four UV set-ups in the greenhouse. Each set-up contained
eight FS-40 fluorescent sunlamps filtered with 0.127 mn Mylar
(M-5) or cellulose acetate (CA-5). Data shown are mean
values with their standard errors and ranges for 48 pot
locations per cultivar per set-up.
Table 2. Criteria for scoring chlorosis in cucumber leaves. 17
Table 3. Influence of increased UV-B irradiation on index of leaf 18
injury, fresh weight and percent dry weight of tops in 'Poinsett*
(P) and 'Ashley' (A) cucumber plants irradiated for 19 days in
the greenhouse (April 1-20, 1977). Plants exposed to a UV
gradient provided by FS-40 fluorescent sunlamps filtered with
0.127 mm Mylar (M-5) or cellulose acetate (CA-5) at various
distances above the canopy. Data shown are riean values with
their standard errors and ranges for 48 plants of each cultivar
per treatment.
IV
-------
FIGURES
Page
Figure 1. 'Poinsett' (green label) and 'Ashley' (yellow label) cucumber 20
plants grown for 19 days from seeding in the greenhouse under
FS-40 fluorescent sunlamps filtered with 0.127 mm Mylar.
Plants received UV-A irradiation but no supplemental UV-B
irradiation. Note absence of chlorosis.
Figure 2. 'Poinsett' (green label) and 'Ashley' (yellow label) cucumber 22
plants after 19 days of UV-B irradiation (6.73 mean weighted
_2
mW-m of BUV) provided by FS-40 fluorescent sunlamps filtered
with 0.127 mm cellulose acetate.
Figure 3. 'Poinsett' (green label) and 'Ashley' (yellow label) cucumber 24
plants after 19 days of UV-B irradiation (11.02 mean weighted
-2
mW-m of BUV) provided by FS-40 fluorescent sunlamps filtered
with 0.127 mm cellulose acetate.
Figure 4. 'Poinsett' (green label) and 'Ashley' (yellow label) cucumber 26
plants after 19 days of UV-B irradiation (15.30 mean weighted
_2
mW-m of BUV) provided by FS-40 fluorescent sunlamps filtered
with 0.127 mm cellulose acetate.
Figure 5. Comparative sensitivity of 'Poinsett' (top row) and 'Ashley'
v
(bottom row) cucumber plants to increased UV-B irradiation.
Plants irradiated for 19 days from time of seeding (17 days
from emergence) in the greenhouse under FS-40 fluorescent
sunlamps filtered with 0.127 mm Mylar (M-5) or 0.127 mm
cellulose acetate (CA-5). The latter plants received a mean
level of biologically effective UV (BUV) irradiance of 6.73,
-2
11.02, or 15.30 weighted mW-rn , respectively.
28
-------
Page
Figure 6. Linear regression of the index of injury for 'Poinsett' 29
cucumber leaves vs. biologically effective UV (BUV) radiation .
-2
in weighted mW'ra . Plants were irradiated in the greenhouse
for 19 days from seeding under eight FS-40 lamps filtered
with 0.127 mm cellulose acetate under a UV gradient ranging
_?
from 4.9 to 26.6 iaW-m BUV (1.6 to 8.7 UV-B sun equivalents).
_7
(One UV-B sun equivalent = 3.06 weighted mW-m of BUV).
Figure 7. UV dose-response relationship under greenhouse conditions. 30
Comparative leaf areas of 'Poinsett1 and 'Ashley' cucumber plants
expressed as percentages of Mylar controls. Plants irradiated
for 19 days from seeding in the greenhouse under eight FS-40
lamps filtered with 0.127 mm cellulose acetate. Lamps mounted
in separate set-ups at 1.43, 0.92, and 0.54 m above the plants.
Means and standard errors are shown for 48 plants within each
-2
set-up for 6.7, 11.0 or 15.3 mW m BUV (2.2, 3.6, or
5.0 UV-B sun equivalents) respectively.
Figure 8. UV dose-response relationship under greenhouse conditions. 31
Comparative dry weights of tops of 'Poinsett' and 'Ashley'
cucumber plants expressed as percentages of Mylar controls.
Plants exposed ten the UV-B irradiation gradients described
in Fig. 7. ;
Figure 9. Linear regression of dry weight of tops of 'Poinsett' cucumber 32
plants vs. exposure to biologically effective UV (BUV) radiation
-2
in mV?-m . See Fig. 6 legend.
Figure 10. Linear regression of total leaf area of 'Poinsett' cucumber 33
_2
plants vs. biologically effective UV (BUV) radiation in mWm
See Fig. 6 legend.
vx
-------
ACKNOWLEDGMENTS
Appreciation is extended to Richard Griffin, Nancy Maher, Scott
Ravitz, and Randy Rowland for their excellent technical assistance;
to E. James Koch and Helen Herlich for their help in obtaining
statistical analysis of the data; and to Jesse Bennett, Sterling
B. Hendricks, E. James Koch, and Olga v.H. Owens for their critical
reviews of the manuscript.
Yii
-------
DIFFERENTIAL SENSITIVITY OF TWO CULTIVAKS OF CUCUMBER
(CUCUMIS SATIVUS L.) TO INCREASED UV-B IRRADIANCE:
I. DOSE-RESPONSE STUDIES
I/ 21 3/
Donald T. Krizek- ' - ' -
INTRODUCTION
The influence of stratospheric ozone reduction and the attendant
increase in solar ultraviolet-B irradiation (UV-B, 280-320 nm) on the
biosphere have been of recent concern (Molina and Rowland, 1974).
During the Climatic Impact Assessment Program (CIAP) in 1972-1975,
numerous studies were conducted on the response of higher plants to
increased UV-B irradiation (Ambler et al., 1975; Basiouny et al., 1978;
Biggs, 1975; Biggs and Basiouny, 1975; Brandle et al., 1977; Caldwell,
1977; Krizek, 1975b; Nachtwey, 1975; Sisson and Caldwell, 1976, 1977;
Van and Garrard, 1975; and Van et al., 1976). Despite the wealth of
Plant Physiologist, USDA, Science and Education Administration,
Agricultural Research, Plant Physiology Institute, Plant Stress
Laboratory, Beltsville, Md. 20705.
2/
Research supported in part^ by the U.S. Environmental Protection Agency
under interagency agreement EPA-IAG-D6-0168.
3/
Abbreviations: PAR: photosynthetically active radiation (400-700 nm);
UV-B: 280-320 nm region; BUV: biologically effective UV irradiance in
-2 -2
weighted mW m ; UV-B SE: UV-B sun equivalent = 3.06 weighted mW'm
BUV; FS-40 lamps: Westinghouse FS-40 fluorescent sunlamps; M-5:
0.127 mm (0.005 in. = 5 mil) Mylar; CA-5: 0.127 mm (0.005 in. = 5 mil)
cellulose acetate.
-------
data collected during this period, relatively little is known about
UV-B dose-response relationships for higher plants. Such information is
critical in order to assess the biological impact of increased UY-B
irradiation caused by stratospheric ozone reduction (Anon., 1977; Krizek,
1975a, 1976, 1977a).
In the course of screening a range of selected species for
comparative sensitivity or resistance to broad-band UV-B irradiation, two
cultivars of cucumber, 'Poinsett' and 'Ashley, were discovered that
differed markedly in their response to increased levels of UV-B
irradiation (Krizek, unpublished results, 1976). The present study was
conducted to establish dose-response relationships for these cultivars
and to determine threshold levels of biologically effective UV (BUV)
irradiation required to induce UV-B damage in these two cultivars.
Regression equations are presented to provide a means for assessing
the potential biological impact of a projected increase in UV-B irradiance
on leaf growth and dry matter production using 'Poinsett1 cucumber as a
model of a highly sensitive plant.
-------
MATERIALS AND METHODS .
Plant Material. 'Poinsett1 and 'Ashley' cucumbers were investigated:
1) because of their differences in UV-B sensitivity observed in an
earlier screening program; 2) their rapid growth rate; 3) their uniformity
in size; and 4) their prostrate habit for the first 2-3 weeks. Experiments
were repeated three times (February, April and June). Data reported
here (obtained during April 1-19, 1977) are representative of the dose-
response relationships obtained.
Cultural Conditions. Plants were grown for 19 days from seed in 12.5 cm
4/
dia. white plastic pots containing a peat-vermiculite mix (Jiffy Mix) .
Five seeds were planted in each pot, using a special template. After
7 days, the seedlings were thinned to one per pot. UV-B irradiation was
begun at the time of planting the seed. Minimum night temperatures in
the greenhouse did not go below 20 C; day temperatures did not exceed
35 C. Natural daylight and photoperiod were used. The plants were
fertilized daily with a 1/4 strength ASMS Hoagland solution (Hammer
et al., 1978).
UV Source. In order to provide a gradient in UV-B irradiance, four
set-ups were constructed in a fiberglass-covered greenhouse (Table 1).
Each set-up contained four fixtures, each containing two FS-40 lamps.
One set-up contained eight FS-40 lamps filtered wฑth Mylar (M-5) as a
4;
Mention of a trademark, proprietary product, or vendor does not
constitute a guarantee or warranty of the product by the USDA and does
not imply its approval to the exclusion of other products or vendors
that may also be suitable.
-------
UV-B control with the center four lamps maintained at a distance of
1 m and the outer four lamps at a distance of 0.75 m above the plants.
Each of the other three set-ups contained eight FS-40 lamps filtered
with cellulose acetate (CA-5) with the center four lamps kept at a
distance of 1.43, 0.92, or 0.54 m and the outer four lamps kept at a
distance of 1.22, 0.70, or 0.54 m respectively above the plants.
The pots were arranged in 8 rows x 12 columns per set-up, each
2
covering an area of 1.73 m (see Cams et al., 1977). This arrangement
was divided, into four quadrants (replicates) and the pots were positioned
equidistantly from the center of each set-up. Pots of 'Poinsett' and
'Ashley' were alternated at 0.15 m intervals along the x and y axes
(corresponding to rows and columns) in order to obtain comparable levels
of UV-B irradiance for each cultivar (Table 2).
The experiment was conducted according to a' standard protocol
described by Krizek (1977b). The lamps were aged 100 hours and the
CA filters were aged for 6. hours prior to use. The CA filters were
changed twice weekly (because of degradation by the short wave UV).
The heights of lamps above canopies were adjusted as the plants grew
\
to maintain the specified levels of UV-B irradiance.
Measurements. Broad-bandi UV-B irradiances were determined at every
', \ot location at the beginning and end of. each experiment by means of a
\ (
', mad-band radiometer developed by the Instrumentation Research
I boratory (IRL UV Meter) (Morris, 1977; Rowan and Morris, 1978). Mean
values for each of the four set-ups are given in Table 1.
The IRL UV Meter was used to obtain the readings presented in
Table 1. The instrument consists of a solar-blind vacuum photo-diode
' (Hamamatsu R403), an integrated circuit amplifier, and a microammeter
-------
packaged in a meter case (Rowan and Korris, 1978; Norris, 1977). The
spectral sensitivity of the IRL Meter in the UV-B .region is relatively
flat, with maximum sensitivity at 300 nm. The detector is insensitive
at wavelengths longer than 400 nm.
Narrow-band UV irradiances were determined at selected pot locations
for every nm wavelength from 250 to 369 nm with an automated spectro-
radiometer developed.by IRL (Norris, 1977; Rowan and Norris, 1978) and
commercially available from Optronic Laboratories, Inc., Silver Spring,
Md. .
-2
BUV weighted irradiances are reported as inW-m , the biologically
effective UV irradiances derived from the AE9 weighting function
described by Thimijan et al., 1978 and Cams et al., 1977. Since UV
irradiation employed in this study was obtained by filtering FS-40
lamps with CA-5, BUV was essentially confined to the UV-B region. Mean
BUV values for each set-up are given in Table 1.
Unweighted spectral irradiances in the UV-B region (Table 1) were
obtained by summing measured values at each nanometer from 280-320 nm
or using the regression equations developed to estimate the UV-B
_2
exposures (Krizek and Koch, 1978). Dividing the mW-m of biologically
v -2 -2
effective UV irradiance by 3.06 raW-m [the raW-m BUV of one Beltsville
control sunshine; i.e., 1 UV-B sun equivalent (SE)] provides the
fraction of BUV relative to that of 1 SE (Thimijan et al., 1978). .
Harvest and Data Analyses. Plants were harvested after 19 days of
UV-B irradiation. An index of injury scale was developed for scoring
the extent of leaf chlorosis (Table 2). Leaf areas were then measured
with a Lambda LI-COR leaf area meter. Fresh and dry weights of the
shoots were taken; the latter were recorded after drying the samples in
-------
a forced draft oven at 80 C for 48 hr. Data on plant height and node
number were also taken but are not reported since .they showed little or
no differences.
Means, standard deviations, standard errors of the mean, and
linear and quadratic regressions on weighted BUV were calculated for
all parameters. Since quadratic regression of the data yielded no
significant improvement of the correlation coefficient (r values), only
linear regression data are presented. Data were analyzed separately by
cultivar and set-up. .
Since the steepest UV gradient was obtained under set-up 4
_2
(Table l)-i.e., that having a mean BUV level of 15.3 mW-m (or 5.0 UV
SE)-data obtained for this set-up were used in calculating the regression
equations presented.
-------
. ป'.'..,T.K
RESULTS AND DISCUSSION
Comparative Phytotoxic Effects of UV-B Irradiation. Leaves of 'Poinsett'
cucumber plants irradiated for 19 days under CA-5 filtered FS-40 lamps
developed marked interveinal and marginal chlorosis with crinkling
distortion evident at the tip and along the margins of the leaves (Fig.
1-5). UV damage was observed within 1-2 days after seedling emergence
and increased in severity as the leaves expanded and UV-B irradiance
increased (Fig. 5). The index of injury under a maximum of 6.7,
-2
11.0, or 15.3 mW-m BUV (2.8, 5.4, or 8.7 UV-B SE) was 3, 5, or 9,
respectively, (Table 3, Fig. 6). These values represented about 15, 25,
or 45% chlorosis, respectively (Table 1). Leaves of 'Ashley' cucumber
plants, on the other hand, never reached an index of injury above 3
_2
(15% chlorosis) even when exposed to 15.3 mW'm BUV (Table 3). For
corresponding mean UV-B doses, 'Ashley' plants exhibited one-third
to one-half as much chlorosis as those of the cultivar 'Poinsett1
(Table 3). .
Under a 40% increase in BUV or (1.4 UV-B SE) (Table 1) which
corresponds to a 20% decrease in 0 content in the stratosphere, leaves
of 'Poinsett' cucumber plants showed an injury scale of 1 (5% or less
chlorosis, while 'Ashley' plants showed an injury scale of 0 (no
chlorosis) (Table 3). The negative regression of injury index for the
'Poinsett' cultivar, as measured by leaf chlorosis and weighted BUV
_2
irradiance in mW- m , is shown in Fig. 6. The r value (i.e., 0.81)
indicates that the regression equation described may be used to estimate
leaf injury expected at a given BUV irradiance level (Krizek and Koch,
1978). Since no significant regression could be established for leaf
injury on BUV for the cultivar 'Ashley' (r = 0.35), the regression is
not shown.
-------
The threshold level for chlorosis in 'Poinsett' cucumber leaves
.varied with season, increasing in the spring and summer and decreasing
in the fall and winter, suggesting a difference in photorepair capability
with season and amount of PAR.
Influence of UV-B Irradiation on Vegetative Growth. At a mean BUV level
of 6.73 mWm~2 (a 120% increase in BUV, or 2.2 UV-B SE), (Table 1)
and 'Ashley' cucumber plants showed approximately equal (6-10%) reduction
in fresh weight of tops (Table 3) and total leaf area (Fig. 7) as
compared to the Mylar control plants. Mean dry weight loss, however,
at this BUV level was greater in the case of 'Poinsett' (5.8%) than for
'Ashley1 (1.4%) when compared to their Mylar controls (Fig. 8).
-2
UV-B irradiances in excess of 6.73 weighted mW'rn . caused greater
reductions in leaf area and dry weight of tops of 'Poinsett1 plants than
for 'Ashley' plants (Figs. 7, 8). The 'Ashley1 cultivar required about
twice as much BUV as 'Poinsett' to produce a 20-25% reduction in leaf
area or dry weight (Figs. 7, 8). Leaf size was reduced by increased UV
irradiation to a greater extent that was dry weight of tops. At 11.02
-2
mWm BUV or greater, vegetative growth as measured by fresh weight
of tops (Table 3), total leaf area (Fig. 7), and dry weight of tops
(Fig. 8), was markedly impaired in both cultivars.
When dry weight data for the 48 'Poinsett' plants under the UV
_2
set-up with the widest exposure range-4.9 to 26.6 mW- m of BUV
(1.6 to 8.7 UV-B SE) and a mean of 15.3 mW-nf2 BUV (5.0 UV-B SE)-were
subjected to linear regression analysis, a correlation coefficient of
0.77 was obtained (Fig. 9). For each unit increase in BUV, the loss in
dry weight would be predicted to be 30.60 mg. On the basis of one SE,
-2
a 37% increase in BUV (4.2 mU-m BUV or 1.37 UV-B SE) would be expected
-------
_o
to reduce dry weight of 'Poinsett1 cucumber plants by 10%; 10.5 mW-m
BUV (or 3.43 SE) would reduce it by 25%; and 21.0 mW'm~2 BUV (or 6.86
UV-B SE) to reduce it by .50%. Since actual reductions in dry weight
obtained at the higher BUV levels were less than this prediction and
since relatively few data points were collected for UV irradiances in
-2
excess of 15.3 mWm BUV (5.0 UV-B SE) , the equation is of greatest
value below this point.
Assuming a 20% decrease in stratospheric 0 reduction caused by
chlorofluoromethanes (CFM's) with a 40% corresponding increase in surface
level BUV [actually likely to be higher than this since the curve is non-
linear above 10% 0 reduction (IMOS, 1975)], one finds on the basis of
this regression curve an approximate 10% decrease in dry weight for a
highly sensitive cucumber cultivar such as 'Poinsett1.
Linear regression analysis of the leaf area data for 48 'Poinsett1
cucumber means on BUV resulted in a correlation coefficient of 0.81
(Fig. 10). For each unit increase in BUV, the decrease in leaf area
2 -2
would be predicted to be 8.84 cm . On the basis of one SE 2.8 mW'm
(0.90 UV-B SE) or 90% of present BUV levels can reduce leaf area in
'Poinsett' cucumber under the conditions of the experiment by 10%;
6.7 inW-in BUV (2.2 UV-B SE) ,* a 120% increase, would be needed to reduce
leaf area by 25%; 13.8 mWir~ BUV (4.5 UV-B SE), a 350% increase, would
be required to reduce leaf area by 50%. Again, assuming a 20% maximum
decrease in stratospheric ozone reduction from CFM's, one could predict
an approximate 15% decrease in total leaf area for this plant.
The inhibitory effects of high UV-B irradiance on leaf growth in
cucumber are consistent with the findings of Sisson and Caldwell (1976,
1977) and Dickson and Caldwell (1978) for Rumex patientia L., previous
-------
work in our laboratory on cotton (Ambler et al., 1975); Alaska pea
(Krizek et al., 1975b) and a number of bedding plants (Krizek and
Semeniuk, 1975); and early work reviewed by Caldwell (1968, 1971).
Preliminary measurements of stomatal resistance did not indicate
a significant difference between Mylar control plants and the green
portions of those exposed to increased UV-B, or between 'Ashley1 and
'Poinsett' leaves. Moisture content of the tops was slightly, but not
significantly, higher in 'Ashley1 than 'Poinsett', suggesting that
differences in turgor were not responsible for the differences in
leaf growth observed (Figs. 8, 10).
Measurements of carbon dioxide exchange rates (CER) made on selected
'Poinsett' cucumber plants at increasing BUV levels indicated a significant
reduction in CER which was related to the amount of chlorosis observed.
BUV irradiances in excess of 40% enhancement levels expected to result
from CFM-catalyzed destruction of stratospheric ozone content were
required to obtain statistically significant differences (Bennett, 1978).
Additional studies are underway to determine the anatomical,
physiological, and biochemical bases for the differences in UV-B
sensitivity observed between the two cucumber cultivars. Possible
explanations to account for these differences might include differences
in optical properties of the leaves with 'different degrees of screening
of the responding sites; differences in photoreactivation; differences
in biochemical make-up including peroxidase activity; and differences
in growth regulator activity.
10
-------
CONCLUSIONS'
Significant differences in UV-B sensitivity were found between two
cucumber cultivars; 'Poinsett' was extremely sensitive and 'Ashley' was
slightly sensitive. Evidence was obtained for UV-B induction of: 1) leaf
chlorosis; 2) inhibition of leaf growth; and 3) reduction in fresh and
dry weight (biomass). These effects were most pronounced under conditions
of low PAR and high UV-B irradiation.
Based on regression analysis of plant data obtained under a range
_2
of UV irradiances from 4.6 to 26.6 weighted mW'm of biologically
effective UV (BUV), it was estimated that a maximum decrease in
stratospheric 0 content of 20% could cause a 10% reduction in dry matter
and a 15% decrease in leaf area in the highly sensitive 'Poinsett'
cucumber cultivar. Whether reductions in growth of this magnitude could
even be detected in nature is questionable.
_2
Increasing the BUV level by at least one SE (i.e., from 3.06 mW-m
-2
to 6.12 mW-m or greater) was required to obtain pronounced chlorosis
of the leaves (> 10% chlorosis) with comparable reductions in biomass;
these levels would be far in excess, however, of the projected levels of
biologically effective UV-B irradiances occurring from CFM-catalyzed
reduction of stratospheric ozbne.
Further work is needed to elucidate the site and mechanisms of UV-B
induced injury in these cucumber cultivars. It is clear that the choice
of plant material is a critical factor to the environmental decision-
maker in assessing the biological implications of stratospheric ozone
reduction and the attendant increase in UV-B irradiation.
11
-------
LITERATURE CITED
Ambler, J. E., D. T. Krizek, and P. Semeniuk. 1975. Influence of UV-B
radiation on early seedling growth and translocation of Zn from
cotyledons in cotton. Physiol. Plant 34(3):177-181.
Anonymous. 1977. United States investigations to evaluate the potential
threat of stratospheric ozone diminution. Presented at United Nations
Environment Programme (UNEP) International Meeting on the Ozone
Layer. Washington, D.C. 1-7, 1977.
Basiouny, F. M., T. K. Van, and R. H. Biggs. 1978. Some morphological
and biochemical characteristics of C_ and C. plants irradiated rath
UV-B. Physiol. Plant 42:29-32.
Bennett, J. H. 1978. Effects of UV-B radiation on photosynthesis and
growth of selected agricultural crops. Final EAGER Report submitted
to the Environmental Protection Agency. 14 pp.
Biggs, R. H. 1975. Effects on plants of increased UV-B radiation.
p. 62-65. In Fourth Conference on CIAP. U.S. Department of Transpor-
tation (T. M. Hard and A. J. Broderick, ed.) Washington, D.C.
Biggs, R. H., and F. M. Basiouny. 1975. Plant growth responses to
elevated UV irradiation under growth chamber, greenhouse, and solarium
conditions. In Impacts of^Climatic Changes on the Biosphere. I.
Ultraviolet radiation effects. Climatic Impact Assessment Program
Monograph 5:195-248.
Brandle, J. R., W. F. Campbell, W. B. Sisson, and M. M. Caldwell. 1977.
Net photosynthesis, electron transport capacity, and ultrastructure of
Pisum sativum L. exposed to ultraviolet-B radiation. Plant Physiol.
60:165-169.
12
-------
Caldwell, M. M. 1968. Solar ultraviolet radiation as an ecological factor
for alpine plants. Ecol. Monogr. 38:243-268.
Caldv.-ell, M. M. 1971. Solar UV irradiation and the growth and
development of higher plants. In (A. C. Giese, ed.) Phytophysiology,
Acad. Press, New York. 6:131-177.
Caldwell, M. M. 1977. The effects of solar UV-B radiation (280-315 ran)
on higher plants: Implications of stratospheric ozone reduction. p.
597-607. _Tn Research in Photobiology (A. Castellani, ed.) Plenum Press,
New York. 726 p.
Cams, K. R., R. Thimijan, and J. M. Clark. 1977. Outline of irradiance
distribution of fluorescent lamps and combinations. Paper 5.6. Presented
at Symposium on Ultraviolet Radiation Measurements for Environmental
Protection and Public Safety. National Bureau of Standards, June 8-9,
1977. p. 74-76. In Final Program and Abstracts.
Dickson, J. G. and M. M. Caldwell. 1978. Leaf development of Rumex
patientia L. (Polygonaceae) exposed to UV irradiation (280-320 nm),
Amer. Jour. Bot. 65(8):857-863.
Hammer, P. A., T. W. Tibbitts, R. W. Langhans, and J. C. McFarlane.
1978. Base-line growth studies of 'Grand Rapids' lettuce in controlled
environments. J. Amer. Soc.^Hort. Sci. 103(5):649-655.
Interagency Task Force on Inadvertant Modification of the Stratosphere
(IMOS). 1975. Fluorocarbons and the Environment. White House Council
on Environmental Quality and Federal Council for Science and Technology.
Washington, D.C.,.NSF 75-403. 109 p.
Kriiiek, D. T. (Chairman), IMOS Subcommittee, on Biological and Climatic
Effects Research. 1975a. A proposed federal program to determine the
biological and climatic effects of stratospheric ozone reduction.
Federal Council for Science and Technology, Washington, D.C. Draft Report.
93 p. .
13
-------
Krizek, D. T. 1975b. Influence of ultraviolet radiation on germination
and early seedling growth. Physiol. Plant 34(3):182-186.
Krizek, D. T. 1976. Influence of increased UV-B radiation on agricul-
tural production, p. 336-365. In Stratospheric Ozone Research and
Effects. Hearings before the subcommittee on the Upper Atmosphere of
the Committee on Aeronautical and Space Sciences. U.S. Senate, 94th
Congress, 2nd session.
Krizek, D. T. 1977a. Biological and climatic effects of stratospheric
ozone reduction: A progress report on the EAGER program. Presented
at Amer. Society for Photobiology Meetings. San Juan, Puerto Rico,
May 15, 1977.
Krizek, D. T. 1977b. Current UV measurement methodology and future needs
in photobiological research. Paper No. 5.2. Presented at Symposium
on Ultraviolet Radiation Measurements for Environmental Protection and
Public Safety. National Bureau of Standards. June 8-9, 1977. p. 49-52.
In Final Program and Abstracts.
Krizek, D. T. and E. J. Koch. 1978. Use of regression analysis in
obtaining estimates of UV spectral irradiance under FS-40 fluorescent
sunlamps filtered with cellulose acetate. Final BACER Report submitted
to the Environmental Protection Agency. 24 p.
Krizek, D. T., R. L. Schaefer, and R. A.'Rowland. 1975. Influence of
UV-B radiation on vegetative growth of Pisum sativum L. 'Alaska'.
HortScience 11(3).-22. (Abstract).
Krizek, D. T. and P. Semeniuk. 1975. Comparative sensitivity of
bedding plants to UV-B radiation. HortScience 10(3):323. (Abstract).
Molina, M. J. and F. S. Rowland. 1974. Stratospheric sink for
chlorofluromethanes: chlorine atom-catalyzed destruction of ozone.
Nature 249:810-812.
14
-------
Nachtwey, S. L. (ed.). 1975. Climatic Impact Assessment Program
(CIAP). Monograph 5, Impacts of Climatic Change on the Biosphere.
Part 1. Ultraviolet Radiation Effects. U.S. Department of .Transpor-
tation. Washington, D.C.
Norris, K. H. 1977. Development of a portable, automated UV-B spectro- '
radiometer. Paper No. 5.5. Presented at Symposium on Ultraviolet
Radiation Measurements for Environmental Protection and Public Safety.
National Bureau of Standards. June 8-9, 1977. p. 72-73. In Final
Program and Abstracts..
Rowan, J. D. and K. H. Norris. 1978. Instrumentation for measuring
irradiance in the UV-B region. Final BACER Report submitted to the
Environmental Protection Agency. 69 p.
Sisson, W. B. and M. M. Caldwell. 1976. Photosynthesis, dark respiration
and growth of Rumex patientia L. exposed to ultraviolet irradiance
(288 to 315 nanometers) simulating a reduced atmospheric ozone column.
Plant Physiol. 58:563-568.
Sisson, W. B. and M. M. Caldwell. 1977. Atmospheric ozone depletion:
reduction of photosynthesis and growth of a sensitive higher plant
exposed to enhanced UV-B radiation. Jour. Expt. Bot. 28:691-705.
\
Thimijan, R. W., H. R. Cams, and L. E. Campbell. 1978. Radiation sources
and related environmental control for biological and climatic effects
UV research (BACER). Final BACER Report submitted to the Environmental
Protection Agency. 78 p.
Van, T. K. and L. A. Garrard. 1975. Effect of UV-B radiation on net
photosynthesis on some Cซ and C crop plants. Proc. Soil and Crop
Sci. Soc. Fla. 35:1-3.
Van, T. K., L. A. Garrard, and S. 11. West. 1976. Effects of UV-B
radiation on net photosynthesis of some crop plants. Crop Sci. 16:715-718,
15
-------
1. Weighted and unweighted. UV spectral irradiance under each of four UV set-ups
in the greenhouse. Each set-up contained eight FS-40 fluorescent sunlamps filtered with
0.127 mm (0.005 in. = 5 mil) Mylar (M-5) or cellulose acetate (CA-5). Data shown are
mean values with their standard errors (SE) and ranges of 48 pot locations per cultivar
per set-up.
Set-Up UV Treatment Cv
1
2
3
4
Filter Max Ht.
Above
Canopy
cm
M-5 97 'Poinsett'
'Ashley'
CA-5 143 'Poinsett'
'Ashley'
CA-5 92 'Poinsett1
'Ashley'
CA-5 54 'Poinsett'
'Ashley'
IRL UV Meter
Reading
scale
3.1
3.1
17.6
17.6
29.6
29.6
41.0
40.9
+ 0.1
+ 0.1
+ 0.5
+ 0.5
+ 1.3
+ 1.3
+ 2.6
+ 2.6
UV-B
Mean + SE
2.2
2.2
3.6
3.6
5.0
5.0
-
+ 0.1
+ 0.1
+ 0.2
+ C.2
+ 0.3
+ 0.3
SE
Range
_
-
1.4-2.8
1.4-2.8
1.7-5.4
1.7-5.4
1.6-8.7
1.5-8.7
BUV-/
Irradiance
Weighted
mW-m
Mean + SE
_
-
6.73 +
6.73 +
11.02 +
11.02 +
15.30 +
15.30 +
0.
0.
0.
0.
0.
0.
3
3
6
6
9
9
UV-B
Spectral
Irradiance
Unweighted
mW m
_
-
548.
548.
920.
920.
1273.
1270.
37
37
32
32
68
58
- BUY = 3.06 x UV-B SE
-------
Table 2. Criteria for Scoring Chlorosis
in Cucumber Leaves
Index of
Injury
0
0.2
1.0
2.0
3.0
4.0
5.0
6.0
7.0
8.0
9.0
10.0
Percent
Chlorosis
None
Trace
5%
10.0%
15.0%
20.0%
25.0% .
30.0%
35 . 0% .
40.0%
45.0%
50.0%
17
-------
Table 3. Influence of increased UV-B irradiation on index of leaf injury, fresh weight and
percent dry weight of tops in 'Poinsett' (P) and 'Ashley' (A) cucumber plants
irradiated for 19 days in the greenhouse (April 1-20, 1977). Plants exposed to a
UV gradient provided by FS-40 fluorescent sunlamps filtered with 0.127 mm (0.005 in.
= 5 mil) Mylar (M-5) or cellulose acetate (CA-5) at various distances above the
canopy. Data shown are mean values with their standard errors and ranges of 48
plants of each cultivar per treatment.
oo
Set-Up UV Treatment
1
2
3
4
Filter Max Ht.
Above
Canopy
cm
M-5 97
CA-5 143
*
CA-5 92
CA-5 54
Cv
..
'Poinsett'
'Ashley'
'Poinsett'
'Ashley'
'Poinsett'
'Ashley'
'Poinsett'
'Ashley'
Index of Injury
Mean + SE
0
0
1.5 + 0.1
0.7 + 0.1
2.8 + 0.2
0.9 + 0.1
4.5 + 0.3
1.5 + 0.2
Range
0
0
1-3
0-3
1-5
0-3
1-9
0-3
Mean
12.0
12.7
11.1
11.7
9.2
11.3
8.6
10.2
Fresh weight %
of tops (g)
+ SE
+ 0.3
+ 0.3
+ 0.2
+ 0.2
+ 0.3
+ 0.3
+ 0.4
+ 0.3
8.
8.
8.
8.
4.
6.
4.
6.
Range
5-17.6
3-19.3
4-15.2
9-14.7
4-13.2
7-15.6
0-13.2
6-15.6
Mean
9.2
9.3
9.5
10.0
9.4
10.0
9.5
10.1
Dry weight
of tops
+ SE
+ 0.9
+ 0.1
+ 0.1
+ 0.1
+ 0.1
+ 0.1
+ 0.2
+ 0.1
Range
7.5-11.6
7.0-10.5
8.4-11.0
8.9-11.4
8.5-11.1
9.2-11.3
6.3-15.7
9.4-11.0
-------
Fig. 1 Appearance of 'Poinsett' (green label) and 'Ashley' (yellow
label) cucumber plants grown for 19 days from seeding in the
greenhouse under FS-40 fluorescent sunlamps filtered with 0.127
mm (0.005 in.) Mylar. Plants received UV-A irradiation but
no supplemental UV-B irradiation. Note absence of chlorosis.
19
-------
w ^
-
> .-. ? ->, --.-
^-'' <! - ? <& ' -; *
,.ซ^
"-"-N
. ^-ซ ซ*ปซปป
.
.-
,,.,., ซป
'
.
;
j
fc^-- -
i
^'ซ*feซi^* ปซ
^
f ""." -- *r*i ' -i *-"x
^smnif''*"'' j,*, ***'*-*
-
.- -'
'
''
"-ปป..
,.
-AX
^^y
'
..^_ซ^ซ^ซ3,*ป- --, . ..^-^ซ*ซ~-..~~ซ~~ - ..-, . - .-
,-
,
f
.
Kswessess-
Mr^t*-*r-?TJj
i
,
t -
-------
Fig. 2 Appearance of 'PoinseLt1 (green label) and 'Ashley' (yellow
label) cucumber plants after 19 days of enhanced UV-B irradiation
_2
(6.73 mean weighted mW-m of BUV) provided by FS-40 fluorescent
sunlamps filtered with 0.127 mm (0.005 in.) cellulose acetate.
21
-------
XUaJfc,
...... - : ,* . 11 f_^~-
-, .. -^K - ซ"4ป vT/, -^
..I."; . . ..-.' ; '-V,..
3 *-J., '. ^^^ +&&
-^H^^-- ! f
'>.. ^f
, - ^-,
,;r " \f -v- ^'-^.-^ ., . '
-
,
;^;-
-.^"v.,^- _
ii^fcvv/1^.^^ *'-
-tr"'
.
>ป ซ. **
"- , , <*ป -.,
..- S~^
<-*v-ซ -
v
/
-.^.
,
...
,:" :*"
r
S
i
K ซ = --^ -^-ป - ^ -*- - -Ji:: -
\, j > .. v
:-X X S '"^
, rf - -***^^ "*I*O
I ; ." !
\
-
.
-------
Fig. 3 Appearance of 'Poinsett1 (green label) and 'Ashley' (yellow
label) cucumber plants after 19 days of enhanced UV-B irradiation
_2
(11.02 mean weighted mW'm of BUV) provided by FS-40 fluorescent
sunlamps filtered with 0.127 mm (0.005 in.) cellulose acetate.
23
-------
r
**" -ซ * "*
,
.
ซ"ป.
^VL_. - .-,
j ^;:^-,< - j . -
' - >-v - ".
f.ll -- 4 -U s
-^ & si 5 ?ซ nfi^
5 "<'-
, v 5 H *.
-^^U^ < 2 | -.,
. -- ^ป \^ *&**;- . ,-, ; .
- ^-.f _ -> / ^ - ^.
%. .,
-'
-^ :
^s?-'
P .. ^ ? \ซ s^-
- --'/ ' x*-*^- ' "
jV*4. : /J*s*fr^
* :.fC ' ^ ^ '""
.
4
' X"ซ-
f.-r- !^J
-*- x ^r
,
.
.
-"? **"'k-, i"^.
>-*^
;
,: ' . ซ%ป
"
*'
'
j
j
i
!
.
(i
,
.
. . -
' ' ' .^Mf-*"""'
-^ci-v -
*fiirt rhaป^ i --^^arttijji'ji ii_
|
-------
Fig. 4 Appearance of 'Poinsett' (green label) and 'Ashley* (yellow
label) cucumber plants after 19 days of enhanced UV-B
-2
irradiation (15.30 mean weighted mW-m of BUV) provided by
FS-40 fluorescent sunlaraps filtered with 0.127 ram (0.005 in.)
cellulose acetate.
25
-------
fc^x.
^tia
_..-__-,
^
...- >,, ' - ,'*.
p :5-^_,,-^
?M
" *2s' i .< 'if
.,.-- &.,. -v - -
; :
ซgs
"-,
- '
-
-<-- ,
I
.
^i'
. . .^^.^ ,- '-
." -^
,
. .
. . * --r-^^saa--*^ y"-*1, . ~ป; ^-^
- ^ ,,..ซ,.^K _^^
/"-,$"'^>~~ ' ,-'''! - -'T-^' ' .
'' ; : . -$ฃ?*
- -
y
5- &* , ,
-
"i - j, /**-
>",.:>&<
,^-:_
~
:^* '
-
N
/-
-
'.
~~
\
X
,
'v
**
J
-------
Fig. 5 Comparative sensitivity of 'Poinsett1 (top row) and 'Ashley'
(bottom row) cucumber plants to increased UV-B irradiation.
Plants irradiated for 19 days from time of seeding in the
greenhouse under FS-40 fluorescent sunlamps filtered with
0.127 mm Mylar (M-5) or 0.127 mm cellulose acetate (CA-5).
The latter plants received a mean level of biologically
effective UV (BUV) irradiance of 6.73, 11.02, or 15.30 weighted
-2 . .
row-in , respectively.
2 7
-------
P
'
17 DAYS OlD L
T R T 17 DAYS I
C f N T E R f
r
t
*-*ซ*;.
NJ
CO
'
:
1*^
5"
V.
.>
CA-Sl
nij Ji"inVi i
. -i~. ... . ^
-------
10
3 8
z
o 6
X
S 4
Z
FS40 & CAS
CUCUMBER
POINSETT
Y =0.9018 + 0.2324XXป.
r = 0.80 8 9 .*****
10 15 20 25
BUV (WEIGHTED mWm'2)
30
Figure 6. Linear regression of the index of injury for 'Poinsett'
cucumber leaves vs. biologically effective UV (BUV) radiation
in weighted mW-nT . Plants were irradiated in the greenhouse
for 19 days from seeding under eight FS-40 lamps filtered
with 0.127 mm cellulose acetate under a UV gradient ranging
from A.9 to 26.6 mW-m BUV (1.6 to 8.7 UV-B sun equivalents).
(One UV-B sun equivalent = 3.06 weighted mW'nT2 of BUV).
29
-------
cs
<
ki-
ll
O
100
ง90
ง 80
o 70
60
50
FS40 & CAS
ASHLEY CUCUMBER
X,
\. \
\x sfi
U POINSETT %(S V
" *ซ4>
"^^
n J^
1 1? * I_ _J___ ^co)
6 9 12
BUV (WEIGHTED mWm'2)
15
Figure 7. UV dose-response relationship under greenhouse conditions.
Comparative leaf areas of 'Poinsett* and 'Ashley' cucumber
plants expressed as percentages of Mylar controls. Plants
irradiated for 19 days from seeding in the greenhouse under
eight FS-40 lamps filtered with 0.127 mm cellulose acetate.
Lamps mounted in separate set-ups at 1.43, 0.92, and 0.54
m above the plants. Means and standard errors are shown __
for 48 plants within each set-up for 6.7, 11.0 or 15.3 mW-m
BUV (2.2, 3.6, or 5.0 UV-B sun equivalents) respectively.
30
-------
to
a. cr
O o
-ฃ
u. z
n ฐ
W U
H- a:
O >^
UJ ^
5 o
o
100
90
80
70
60
50
FS40 & C&5
wb xv^v n ASHLEY CUCUMBER
****** ;ป. ^ ^ ^ ^ V X r-*T-l
***/^ XxLJ \
\ XVVVXV -r
\ . . XVD
*%
*Xr
POINSETT CX
*Vป T*
** JL
****aป - ^^
^
-
0 B S I 9
0 36 9 12 15
-2,
BUY (WEIGHTED mWm'z)
Figure 8. UV dose-response relationship under greenhouse conditions.
Comparative dry weights of tops of 'Poinsett' and 'Ashley1
cucumber plants expressed as percentages of Mylar controls,
Plants exposed to the UV-B irradiation gradients described
in Fig. 7.
31
-------
o
ง
CA
W 4
D.
o
H-
UL.
0
I
o
UJ
ฃ
Qi
Q
1200
1000
800
600
400
200
FS40 & CAS CUCUMBER
POINSETT
'*X V ~ 1285.09-30.5980X
**X r - 0.7662
-
\
\
" ******
\
M
J 8 B I B |
0 5 10 15 20 25 30
BUV (WEIGHTED
Figure 9. Linear regression of dry weight of tops of 'Poinsett' cucumber
plants vs. exposure to biologically effective UV (BUV) radiation
in mW-rn . See Fig. 6 legend.
32
-------
-------
FINAL REPORT
USE OF REGRESSION ANAYLSIS IN OBTAINING ESTIMATES
OF UV SPECTRAL IRRADIANCE UNDER FS-40 FLUORESCENT
SUNLAMPS FILTERED WITH CELLULOSE ACETATE
D. T. Krizek
E. J. Koch
Plant Stress Laboratory
Plant Physiology Institute
Beltsville Agricultural Research Center
Beltsville, Maryland 20705
and
Northeastern Region
EPA-IAG-D6-0168
Project Officer:
R. J. McCracken
Agricultural Research, Science and Education Administration
U.S. Department of Agriculture
Washington, D.C. 20250
Prepared for:
Environmental Protection Agency
EAGER Program
Washington, D.C. 20460.
-------
CONTENTS
Abstract iii
Figures and Tables iv
Acknowledgments vi
1. Introduction 1
2. Materials and Methods 3
UV Source 3
UV Instrumentation 3
UV Measurements 4
Regression Analysis 5
3. Results and Discussion 6
Weighted and Unweighted Measurements of
UV-B Irradiance ' 6
Relationship Between IRL-UV Meter Readings
and UV-B Spectral Irradiance 7
Relationship Between UV-B Sun Equivalent and
UV-B Spectral Irradiance 8
4. Literature Cited 10
-------
. ABSTRACT
Weighted and unweighted UV spectral measurements x^ere obtained under
two Westinghouse FS-40 fluorescent sunlamps filtered with 0.127 mm (0.005 inch
or 5 mil) cellulose acetate using newly developed broad-band UV radiometers
and an automated UV spectroradiometer in 10 cm increments from 20 cm to
110 cm. Correlations were determined between sets of data obtained x^ith
the broad-band radiometer and the UV spectroradiometer.
Linear regression analyses were performed on the weighted and unweighted
spectral data to obtain regression equations for predicting UV-B irradiance
-2 -2
(unweighted mW-rn ), biologically effective UV (BUV) (weighted mW.m ) in
the 280-320 nm (UV-B) region, UV-B sun equivalents, and incident UV flux
-2 21
in the UV-B region in photons-m x 10 integrated over a 6-hour exposure.
Examination of the correlation coefficients (r values) indicated
excellent agreement between measured and predicted values for all comparisons
(r values of 0.9972 to 0.9998).
Use of the regression equations should permit accurate and rapid
estimates of both weighted and unweighted UV irradiances at any location in
an experimental set-up and provide a useful means of making interlaboratory
comparisons of spectral measurements.
iii
-------
List of Figures and Tables Page
Figure 1. Regression of UV spectral irradiance in the 280-320 nra 14
(UV-B) region versus IRL UV-meter (10 scale) reading.
Table 1. Weighted and unweighted measurements of UV-B radiation 15
obtained on Nov. 8, 1977, under two Westinghouse FS-40
fluorescent sunlamps filtered with 0.127 mm (0.005 in.)
cellulose acetate (aged 6 hours). Measurements taken
at 10 cm intervals from 20 cm to 110 cm.
Table 2. UV sun equivalents obtained at various distances under 16
Westinghouse FS-40 fluorescent sunlamps filtered with
0.127 mm (0.005 in.) cellulose acetate (aged 6 hours).
Table 3. Biologically effective UV (BUY) radiation in the 250-279, 17
-2
280-330, and 250-369 nm region (weighted mW-en ) under two
Westinghouse FS-40 fluorescent sunlamps filtered with 0.127
mm (0.005 in.) cellulose acetate (aged 6 hours).
_2
Table 4. Total UV irradiance (unweighted mW-m ) under two Westinghouse 18
FS-40 fluorescent sunlamps filtered with 0.127 mm (0.005 in.)
cellulose acetate (aged 6 hours). Measurements taken every
nm from 250 to 369 nm with an Optronic UV spectroradiometer.
Table 5. Actual and predicted relationship between IRL UV-meter reading 19
(10 scale) and unweighted UV spectral irradiance in the 280-320
nm (UV-B) region. Two FS-40 sunlamps plus 0.127 mm (0.005
in.) cellulose acetate (aged 6 hours).
Table 6. Actual and predicted relationship between IRL UV-meter reading 20
(10 scale) and weighted UV spectral irradiance in the 280-320
nm (UV-B) region. Two FS-40 sunlamps plus 0.127 mm (0.005 in.)
cellulose acetate (aged 6 hours).
IV
-------
Page
Table 7. Actual and predicted relationship between UV-B sun 21
equivalent as measured with an Optronic Model 725 broad-band
radiometer (0-10 scale and 0-5 scale) and unweighted UV spectral
_2
irradiance (mW-m ) in the 280-320 nm region. Two FS-40
sunlamps plus 0.127 mm (0.005 in.) cellulose acetate (aged 6
hours).
Table 8. Actual and predicted relationship between UV-B sun equivalents 22
as measured with an Optronic Model 725 broad-band radiometer
(0-10 scale and 0-5 scale) and biologically effective UV
(BUV) radiation in the 280-320 nm region. Two FS-40 sunlamps
plus 0.127 mm (0.005 in.) cellulose acetate (aged 6 hours).
Table 9. Actual and predicted relationship between UV-B sun equivalent 23
as measured with an Optronic Model 725 broad-band radiometer
(D-10 scale) and incident UV flux in the 280-320 nm (UV-B) region
-2 21
measured in photons'm x 10 integrated over a 6 hour day.
Two FS-40 sunlamps plus 0.127 mm (0.005 in.) cellulose acetate.
Table 10. Actual and predicted relationship between UV-B sun equivalent 24
as measured with an Optronic Model 725 broad-band radiometer
(0-5 scale) and incident flux in the 280-320 nm (UV-B) region
^
-2 21
measured in photons-m x 10 integrated over a 6 hour day.
Two FS-40 sunlamps plus 0.127 mm (0;.0.0.5 :in.) cellulose acetate.
v
-------
ACKNOWLEDGMENTS
We gratefully acknowledge Karl H. Norris, James D. Rowan, and
George F. Buttons, Instrumentation Research Laboratory, for their
outstanding cooperation in developing the instrumentation used in this
study and assisting in the calibration of these instruments; we thank
Scott J. Ravitz for his competent technical assistance in obtaining
these measurements. We also appreciate the assistance of Lowell E. Campbell,
Stanley Holliday, and Richard W. Thimijan, Agricultural Equipment
Laboratory, in constructing the UV irradiation set-ups.
vi
-------
USE OF REGRESSION ANSYLSIS IN OBTAINING ESTIMATES OF UV SPECTRAL IRRADIANCE
UNDER FS-40 FLUORESCENT SUNLAMPS FILTERED WITH CELLULOSE ACETATE
I/ 21
Donald T. Krizek- and E. James Koch-
INTRODUCTION
Broad-band studies on the influence of ultraviolet radiation in the
280-320 nm (UV-B) region on plant growth and development conducted since
1972 during the Climatic Impact Assessment Program (CIAP) have demonstrated
the urgent need for improved UV sources and instrumentation (Ambler et al.,
1975; Biggs, 1975; Brandle et al., 1977; Krizek, 1975a, b; 1977a, b;
Caldwell, 1971, 1972, 1977; Ormrod and Krizek, 1978; Sisson and Caldwell,
1975, 1976, 1977; Skelly et al., 1978; Anon;, 1977; Nachtwey, 1975; Van
and Garrard, 1975, 1976.
With the recent development of broad-band radiometers and an automated
UV spectroradiometer (Norris, 1977; Rowan and Norris, 1978; Cams et al.,
1977) and improvements in spectroradiometers used in CIAP (Kostkowski
and Saunders, 1977) the researcher now has the means of obtaining greatly
improved UV measurements.
The objective of the present study was to provide the investigator
involved in the EPA Interagency Biological and Climatic Effects Research (EAGER)
program a means of obtaining Estimates of weighted and unweighted spectral
irradiance in the UV-B region. The use of regression analysis of spectral
data obtained with both broad-band radiometers and an automated UV spectro-
radiometer is described.
Plant Physiologist, Plant Stress Laboratory, Agricultural Research, Science
and Education Administration, USDA, Beltsville, Maryland 20705.
2/
Biometrician, Agricultural Research, Science and Education Administration,
USDA, Beltsville, Maryland 20705.
-1-
-------
A series of regression equations is described for relating weighted
and unweighted spectral data in the UV-B region. Use of these equations
should facilitate comparison of spectral measurements obtained in
different laboratories.
-------
MATERIALS AND METHODS
UV Source
3/
UV radiation was provided by Westinghouse FS-40 fluorescent sunlamps
that had been aged at least 100 hours according to a standard protocol
(Krizek, 1977b). The lamps were mounted in a single 1.2 meter (A foot)
fluorescent fixture without a special reflector (Thimijan et al., 1978)
and covered with 0.127 mm (0.005 in. = 5 mil) cellulose acetate (CA).
The CA filters were aged for 6 hours on a specially designed lamp rack
before being used in the study (Thimijan et al., 1978).
UV Instrumentation
Broad-band UV-B irradiance levels were determined by means of three
instruments developed by Norris and his associates in the USDA Instrumentation
Research Laboratory (IRL) at Beltsville, Maryland, or based on his specifications
(Norris, 1977; Rowan and Norris, 1978): (a) an IRL Meter UV-B radiometer
(IRL UV meter); (b) an Optronic Laboratories, Inc. Model 725 UV-B radiometer
_2
calibrated to read from 0 to 5 UV sun equivalents [or 0 to 15.3 mW'm of
biologically effective UV (BUV) radiation (Cams et al., 1977)]; and (c)
the same instrument calibrated to read from 0 to 10 UV sun equivalents (or
-2
0 to 30.6 mW-m of BUV radiation).
The IRL meter was used to obtain unweighted UV-B measurements. The
3/
instrument consists of a solar-blind vacuum photo-diode (Hamamatsu R403) ,
an integrated circuit amplifier, and a microammeter packaged in a meter case
(Rowan and Norris, 1978; Norris, 1977). The circuit provides for four decades
3/
Mention of a trademark, proprietary product, or vendor does not constitute
a guarantee or warranty of the product by the USDA and does not imply its
approval to the exclusion of other products or vendors that may also be
suitable.
3
-------
of range switching, referred to as 10 , 10 , 10 , and 10 . In the present
7 8
study, the 10 and 10 scales were used. The spectral sensitivity of the
IRL meter in the 280 to 320 nm region is relatively flat with maximum
sensitivity at 300 nm. The detector is insensitive at x^avelengths longer
than 400 nm (Rowan and Norris, 1978).
UV spectral irradiances were determined every nm from 250 to 369 nm
with an automated spectroradiometer (with a 2 nm bandwidth) developed by
IRL and commercially available from Optronic Laboratories, Inc. Specifications
for this instrument are described by Norris, 1977, and Rowan and Norris, 1978.
UV Measurements
UV measurements were taken with each of these three broad-band radiometers
and the automated spectroradiometer described at 10 cm intervals from 10 cm
to 110 cm. The sensor was placed under the center of the lamp fixture and
adjusted by means of a standard laboratory jack. All measurements were
taken in a darkened room with only UV lamps on. Air temperature was maintained
at 25ฐC.
_2
Weighted irradiance levels are reported as mW'm BUV, the biologically
effective UV radiation derived from the weighting function (AZ9) described
by Thimijan et al., 1978, and Cams et al., 1977.
Since UV irradiation used in this study was obtained by filtering FS-40
lamps with CA, BUV was essentially confined to the UV-B region. Unweighted
irradiances in the UV region were obtained by summing measured or calculated
values at each nanometer from 250-279 nm (UV-C), 280-320 nm (UV-B) , a.nd
321-369 nm (UV-A). Although spectral data were taken previously at every
nm from 321-400 nm, no data were taken beyond 369 nm in the present study,
since the UV-B portion of the spectrum was the major region of concern
in the BACER program.
-------
-2 -2
Dividing the mW-ra BUV by 3.06 (the mW'm BUV of the Beltsville
control sunshine) provides the fraction of BUV measured at each location
relative to that of one Beltsville control sunshine.
Regression Analysis
Linear regression analyses were performed on the weighted and unweighted
spectral data to obtain regression equations for predicting UV-B irrsdiance
-2 -2
(mW*m ), BUV (weighted mW'm ) in the 280-320 nm region, UV-B sun equivalents,
-2 21
and incident UV flux in the UV-B region in photons-m " x 10 integrated
over a 6-hour day exposure.
5
-------
RESULTS AND DISCUSSION
Weighted and Unweighted Measurements of UV-B Irradiance
Weighted and unweighted spectral measurements obtained at various
distances from a pair of FS-40 fluorescent sunlamps filtered with CA are
shown in Table 1. In general, there was good agreement between the values
obtained for UV-B sun equivalents obtained on the Optronic radiometers and
those calculated from unweighted measurements obtained on the IRL UV
meter using the conversion factors provided by Thimijan et al. 1978.
The UV-B sun equivalents obtained by summing values every nm from
280-320 nm (Table 1) or 250-329 nm (Table 2) on the automated spectroradio-
meter by means of computer calculation also agreed within 5 to 10% of
those obtained with the broad-band radiometers (Tables 1, 2).
By using a CA filter over the FS-40 sunlamps, little UV radiation
in the 250-279 nm region was transmitted (Table 2). The level of BUV
radiation transmitted in the 250-279 nm (UV-C); 280-330 nra and 250-330
nm regions under FS-40 lamps filtered with CA are shown in Table 3. Since
UV-C radiation contributed virtually no measurable BUV, the total amount
of BUV obtained in the 250-330 nm region was approximately the same as
that obtained by summing the BUV values in the 280-330 nm region alone
*
(Table 3).
*
The total unweighted UV irradiance obtained at each 10 nm interval in
the 250-279 nm (UV-A), 280-320 nm (UV-B), 321-369 nm (UV-C) and 250-369 nm
regions is shown in Table 4. About 50% of the total UV irradiance
transmitted in the region of 250-369 nm was at 280-320 nm and 50% at
321-369 nm (Table 4).
-6-
-------
Relationship Between IRL-UV Meter Readings and UV-B Spectral Irradiance
The actual and predicted relationship between broad-band radiometer
readings with an IRL UV meter and unweighted spectral irradiance in the
280-320 nm (UV-B) region under two FS-40 sunlamps filtered with CA is
shown in Table 5. A plot of these data (Fig. 1) and evaluation of the
correlation coefficient (r = 0.9995) indicate that the relationship between
UV spectral irradiance obtained on the UV spectroradiometer and that predicted
by the regression equation from the IRL meter readings agrees very well.
The relationship between IRL UV meter reading and incident UV flux
2 21
at 280-320 nm measured in photons-m x 10 integrated over a 6-hour
period is also shown (Table 5). Since most studies in the EAGER program on
simulation of ozone depletion were based on 6 hours of UV irradiation per
day, the calculations for incident flux are based on this duration of exposure,
rather than on the basis of seconds or minutes. The correlation coefficient
(r) of 0.9976 indicates that there is good agreement between the calculated
and predicted incident UV flux in the 280-320 nm region (Table 5). This
equation may, therefore, be used to describe the UV-B irradiance in terms
used by the photobiologist (see e.g., Seliger, 1978; Rupert, 1978; Rupert
and Latarjet, 1978; Caldwell, 1972).
\
The relationship between IRL meter reading and BUV radiation in the
280-320 nm region is shown in Table 6. The r values of 0.9992 obtained
indicates that the agreement between predicted and actual BUV is nearly
perfect. The relationship between IRL meter readings and UV-B sun equiva-
lents is also shown (Table 6). The r of 0.9991 obtained indicates that
there is excellent agreement between predicted and determined UV sun
equivalents.
7
-------
As rule of thumb, instantaneous meter readings on the UV-B broad-
band radiometer (IRL Meter) may be converted to UV-B spectral irradiance
in photons-m~2 for a 6-hr exposure by multiplying the reading by 10 .
Relationship Between UV-B Sun Equivalent and UV-B Spectral Irradiance
The relationship between UV-B sun equivalent as measured with an
Optronic Model 725 broad-band radiometer (either the 0-10 scale or the
0-5 scale) and the unweighted UV spectral irradiance in the 238-320 nm
region is shown in Table 7. The r's obtained, namely, 0.9996, 0.9995,
respectively, indicate that the regression equations described (Table
7) may be used to accurately estimate the total UV-B irradiance obtained
at any of the cooperating laboratories participating in the BACER ter-
restrial effects program that were sent these instruments.
Since all of the Optronics Model 725 radiometers were calibrated by
IRL under a pair of FS-40 fluorescent sunlamps filtered with 0.127 mm
(0.005 in.) CA aged 6 hours, it should be possible to obtain an intra-
laboratory and interlaboratory comparison of weighted and unweighted
spectral irradiance used at each pot location in any particular study by
use of these regression equations provided that no filters were used over
the sensor.
*
The relationship between UV-B sun equivalent as measured with an
Optronic Model 725 broad-band radiometer (either the 0-10 scale or the
0-5 scale) and the BUV radiation in the 280-320 nm region is shown in
Table 8. The r values obtained, namely, 0.9998 and 0.9996, respectively,
indicate that the regression equations obtained may be accurately used to
estimate the level of BUV obtained at any location in the experimental
set-up.
-------
The relationship between UV-B sun equivalent as measured with an
Optronic Model 725 broad-band radiometer (either the 0-10 scale or the
_2
0-5 scale) and the incident flux in the 280-320 nm region in photons-m
x 1021 per 6 hr day of UV irradiation is shown in Tables 9 and 10. The
r values obtained (Tables 9 and 10), namely, 0.9972 and 0.9974, respectively,
indicate that the regression equations obtained may be accurately used
? 21
to estimate the incident UV flux in photons-ra x 10 received by a
plant during a single 6-hr period of UV exposure.
Use of the regression equations described in this report should enable
the investigator to make accurate and rapid estimates of both weighted
and unweighted UV irradiance at any location in an experimental UV set-up,
provided that FS-40 fluorescent sunlamps and 0.127 mm (0.005 in.) cellulose
acetate filters are used. By doing so, countless hours can be saved by
not having to make spectroradiometrie measurements at more than a few
selected locations.
Thimijan et al. (1978) have described a method of calculating the
spectral power output under both filtered and unfiltered FS-40 lamps as
well as Westinghouse BZS lamps by summing values in 5 nm increments from
270 to 320 nm and adding the power output of the Hg lines at 253.6, 289.4,
296.7, 302.2, and 313 nm. This procedure^ however, is more time-consuming
than the present method of using regression equations.
-9-
-------
LITERATURE CITED
\
Ambler, J. E., D. T. Krizek, and P. Semeniuk. 1975. Influence of UV-B
radiation on early seedling growth and translocation of ฐ-*Zn frOm
cotyledons in cotton. Physiol. Plant 34(3):177-181.
Anonymous. 1977. United States investigations to evaluate the poten-
tial threat of stratospheric ozone diminution. Presented at United
Nations Environment Programme (UNEP) International Meeting on the
Ozone Layer. Washington, D.C. March 1-7, 1977.
Biggs, R. H. 1975. Effects on plants of increased UV-B radiation. Pp.
62-65. In Fourth Conference on CIAP. U.S. Dept. of Transportation
(T. M. Hard and A. J. Broderick, ed.) Washington, D.C.
Brandle, J. R., W. F. Campbell, W. B. Sisson, and M. M. Caldwell. 1977.
Net photosynthesis, electron transport capacity, and ultrastructure
of Pisum sativum L. exposed to ultraviolet-B radiation. Plant Physiol.
60:165-169.
Caldwell, M. M. 1971. Solar UV irradiation and the growth and development
of higher plants. In_ A. C. Giese, ed. Photophysiology 6:131-177.
Acad. Press, New York.
\
Caldwell, M. M. 1972. Biologically effective solar ultraviolet irradiation
in the Arctic. Arctic and Alpine Research 4(1):39-43.
Caldwell, M. M. 1977. The effects of solar UV-B radiation (280-315 nm)
on higher plants: Implications of stratospheric ozone reduction.
Pp. 597-607. In_ A. Castellani, ed. Research in Photobiology.
Plenum Press, New York. 726 p.
-10-
-------
Cams, H. R., R. Thimijan, and J. M. Clark. Outline of irradiance distri-
bution of fluorescent lamps and combinations.. Paper 5.6. Presented
at Symposium on Ultraviolet Radiation Measurements for Environmental
Protection and Public Safety. National Bureau of Standards, June 8-9,
1977. Pp. 74-76. In Final Program and Abstracts.
Kostkowski, H. J. and R. Saunders. 1977. Second Quarterly BACER Report
to the Environmental Protection Agency. National Bureau of Standards,
Washington, D.C.
Krizek, D. T. (Chairman), IMOS Subcommittee on Biological and Climatic
Effects Research. 1975a. A proposed federal program to determine
the biological and climatic effects of stratospheric ozone reduction.
Federal Council for Science and Technology, Washington, D.C. Draft
Report.
Krizek, D. T. 1975b. Influence of ultraviolet radiation on germination
and early seedling growth. Physiol. Plant 34(3):182-186.
Krizek, D. T. 1977a. Biological and climatic effects of stratospheric
ozone reduction: A progress report on the BACER program. Presented
at Amer. Society for Photobiology Meetings. San Juan, Puerto Rico.
May 15, 1977.
Krizek, D. T. 1977b. Current UV measurement methodology and future needs
in photobiological research. Paper No. 5.2. Presented at Symposium
on Ultraviolet Radiation Measurements for Environmental Protection
and Public Safety. National Bureau of Standards. June 8-9, 1977.
Pp. 49-52. In Final Program and Abstracts.
Nachtway, D. S. 1975. Climatic Impact Assessment Program (CIAP).
Monograph 5, Impacts of Climatic Change on the Biosphere. Part 1
Ultraviolet Radiation Effects. U.S. Department of Transportation.
Washington, D.C.
-11-
-------
Norris, K. H. 1977. Development of a portable, automated UV-B spectro-
radiometer. Paper No. 5.5. Presented at Symposium on Ultraviolet
Radiation Measurements for Environmental Protection and Public Safety'.'
National Bureau of Standards. June 8-9, 1977. Pp. 72-73. In Final
Program and Abstracts.
Ormrod, D. P. and D. T. Krizek. 1978. Plant stress studies in controlled
environments. HortScience 13(4):453-456.
Rowan, J. D. and K. H. Norris. 1978. Instrumentation for measuring
irradiance in the UV-B region. Final BACER Report submitted to the
Environmental Protection Agency.
'Rupert, C. S. 1978. Uniform terminology for radiations. Pliotochem.
Photobiol. 28:1.
Rupert, C. S. and R. Latarjet. 1978. Toward a nomenclature and dosimetric
scheme applicable to all radiations. Photochem. Photobiol. 28:3-5.
Seliger, H. H. 1978. Environmental photobiology. Chapter 6. Pp. 143-
173. Iii The Science of Photobiology (K. C. Smith, ed.). Plenum
Press, New York.
Sisson, W. B. and M. M. Caldwell. 1976. Photosynthesis, dark respiration,
and growth of Rumex patientia L. exposed to ultraviolet irradiance
v
(288 to 315 nanometers) simulating a reduced atmospheric ozone column.
Plant Physiol. 58:563-568.
Sisson, W. B. and M. M. Caldwell. 1977. Atmospheric ozone depletion:
Reduction of photosynthesis and growth of a sensitive higher plant
exposed to enhanced UV-B radiation. Jour. Expt. Bot. 28:691-705.
Sisson, W. B. and M. M. Caldwell. 1975. Lamp/filter systems for simula-
tion of solar UV irradiance under reduced atmospheric ozone.
Photochem. Photobiol. 21:453-456.
-12-
-------
Skelly, J. M., M. F. George, H. E. Heggestad, and D. T. Krizek. 1978.
Air pollution and radiation stresses. Chapter 2.5. In ASAE Mono-
graph. Modification of the Aerial Environments of Plants. In press.
Thiraijan, R. W., H. R. Cams, and L. E. Campbell. 1978. Radiation
sources and related environmental control for biological and climatic
effects UV research (BACER). Final EAGER Report submitted to the
Environmental Protection Agency.
Van, T. K. and L. A. Garrard. 1975. Effect of UV-B radiation on net
photosynthesis of some C^ and C, crop plants. Proc. Soil Crop.
Sci. Soc. Fla. 35:1-3.
Van, T. K., L. A. Garrard and S. H. West. 1976. Effects of UV-B radiation
on net photosynthesis of some crop plants. Crop Sci. 16:715-718.
-13-
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JMJL Ol 01 X
-------
Weighted and J^k*eighted measurements of UV-B radiation obtained on November 8, 1977 in the laboratory under two Wj^fcing- *~r~~
house FS-40 ^^Prescent sunlamps filtered with 0.127 mm cellulose acetate (aged 6 hours) mounted in a single fixtul^ without
reflector and room lights off. Detector heads placed under the center of the fixture in a horizontal position.
Distance IRL
from Meter
sensor Reading
(cm) (107 scale)
a/ b_/
110 6.90
100 7.95
90 9.35
80 10 . 0 .
70 12.0
60 15.0
K 50 19.0
i
40 24.0
30 33.0
20 48.0
UV-B sun
Equiv.
calculated
from IRL meter
UV-B sun
equiv.
measured
on Optronic
Radiometer
UV~B sun
equiv.
measured
on Optronic
Radiometer
UV-B sun
equiv.
measured
on spectro-
radiometer
BUV mW m 2
280-320nm
based on
spectro-
radiometer
UV-B spectral
irradiance
(280-32j)nm)
mW' m
c/ d/ ฃ/ f/ ฃ/ W
(0 to 5 scale) (0 to 10 scale)
0.8
0.9
1.1
1.2
1.5
1.9
2.3
3.0
4.1
5.9
i
0.9
1
1.1
1-2
, 1-5
1.7
2.1
2.6
3.3
4.4
off scale
0.8
1.0
1.1
1.4
1.6
2.0
2.5
3.2
4.3 .
6.1
0.7806
0.8861
1.0331
1.2317
1.4758
1.7811
2.2037
2.8514
3.7911
5.4484
2.3885
2.7116
3.1613
3.7689
4.5161
5.4503
6.7433
8.7252
. 11.6008
16.6722
206.7507
234.6552
274.8037
324.2766
392.0929
475.1478
591.2049
763.8646
1024.9800
1481.1394..
a/ distance measured from outer wall of lamp to detector head
b_/ measurements at 90, 100, and 110 cm read at 10 ฐ scale and converted to 10' scale
cj based on conversion factor provided by R. Thimijar. of IRL meter reading x 0.123
d/ set to a maximum of 5 UV-B sun equivalents, (15.3 mW-m B.UV)
ej set to a maximum of 10 UV-B sun equivalents, (30.6 mW-m ^BUV)
, f/ weighted portion indicated for 280-320nm region alone _2
ฃ/ BUV = biologically effective UV radiation from 280-320nm (weighted mW-m" )
BUV of 3.06 mWm = one UV-B sun equivalent
h/ actual unweighted flux obtained on an Optronic spectroradiometer; UV-B irradiance measured every nm from
250-369nm but summed from 280-320nm
-------
Table 2.
UV sun equivalents obtained at various distances under two Westinghouse
FS-40 fluorescent sunlamps filtered with 0.127 mm (0.005 in.) cellulose acetate
(aged 6 hrs.). Measurements taken with an Optronic UV spectroradiometer in
the laboratory with room lights off.
Distance
from
sensor
110
100
90
80
70
60
50
40
30
20
UV Sun
Equiv.
250-279 nm
.0057
.0066
.0075
.0092
.0110
.0136
.0172
.0215
.0288
.0431
UV Sun
Equiv.
280-329 nm
0.7806
0.8861
1.0331
1.2317
1.4758
1.7811
2.2037
2.8514
3.7911
5.4484
UV Sun
Equiv.
250-329 nm
0.7863
0.8927
1.0406
1.2409
1.4868
1.7947
2.2209
2.8729
3.8199
5.4915
-------
Table 3.
Biologically effectively (BUV) radiation in the 250-279nm, 280-330nm and
250-330nm region (mWra )under 2 Westinghouse FS-40 fluorescent sunlamps
filtered with 0.127 mm (0.005 in.) cellulose acetate (aged 6 hrs.).
Values based on UV spectroradiometer measurements taken every nm from
250-369nm.
Distance'
from
sensor
(cm)
110
100
90
80
70
60
50
40
30
20
Total
BUV
250-279
weigh t-fi$
mW -m
.0177
.0202
.0229
.0283
.0335
.0415
.0526
.0659
.0882
.1319
Total
BUV
280-330
weighted
mW ' ni
2.3885
2.7116
3.1613
3.7689
4.5161
5.4503
6.7433
8.7252
11.6008
36.6722
Total
BUV
250-330
weighteji,
mW 'ra
2.4062
2.7318
3.1842
3.7972
4.5496
5.4918
6.7959
8.7911
11.6890
16.8041
-17-
-------
Table 4.
Total UV irradiance (unweighted mVJ-m") under two Westinghouse FS-40
fluorescent sunlamps filtered with 0.127 mm (0.005 in.) cellulose acetate
(aged 6 hours). Measurements taken with an Optronic UV spectroradiometer
every nm from 250-369nm, and total irradiance determined for the UV-A,
B, and C regions.
Distance
from
sensor
(cm)
110
100
90
80
70
60
50
40
30
20
Total
Irradiance
UV-C
250-279nm
mW 'ra
0.0223
0.0258
0.0294
0.0361
0.0428
0.0533
0.0668
0.0850
0.1127
0.1691
Total
Irradiance
UV-B
280-320nm
_2
mW -m
206.7507
234.6552
274.8037
324.2766
392.0929
475.1478
591.2049
763.8646
1,024.9800
1,481.1394
Total
Irradiance
UV-A
321-369nm
_2
mW -m
195.8107
226.1185
263.9468
314.7281
381.6198
467.3057
583.7436
760.5765
1,027.8479
1,489.0918
Total
Irradiance
250-369_i^n
mW m
402.5837
460.7995
538.7799
639.0408
773.7555
942.5068
1,175.0173
1,524.5261
2,052.9406
2,970.4003
-18-
-------
Table 5. Actual and predicted relationship between IRL UV meter reading and unweighted UV spectral
irradiance in the 280-320 nm (UV-B) region. YI = UV-B irradiance in mWm and Y- = incide
flux in the 280-320 nm region measured in photons-m~2xlO" per 6 hour day.
UV-B Irradiance
in unweighted
roW -m~z
Distance
from
sensor
cm
110
100
90
80
70
60
50
40
30
20
IRL UV Meter Reading
(10 scale)
X
6.90
7.95
9.35
10.00
12.00
15.00
19.00
24.00
33.00
48.00
Instantaneous
Measurement
Yl
206.7507
234.6552
274.8037
324.2766
392.0929
475.1478
591.2049
763.8546
1024.9800
1481.1394
r = 0.995
Error of
Prediction
A
Yr*i
-9.9667
-14.6079
-17.8537
11.4719
17.2964
7.3636
-.5629
17.1073
-.7304
-9.5095
X = IRL UV meter reading
Incident UV-B
in photonsปm
Integrated for
6 hr day
Y2 '
6.7459
7.6564
8.9664
10.5306
' < 12.7934
**r 15.5033
19.2901
24.9237
30.7494
48.3272
r = 0.9976
X = IRL.UV meter
flux in
-2 in21
xlO
Error of
Prediction
A
Y2-y2
-.3446
-.4698
-.5407
.4323
.6723
.4230
.2642
.9658
-2.0861
.6957
reading
(107 scale)
y = predicted
^\
y, = 2.8457 + 30.9959X
UV-B irradiance
(10 scale)
A
y_ = predicted incident UV-B flux
in photons-m~2 xlO21 per 6 hr day
vn = 0.2843 + 0.9864X
-------
I
ro
O
table 6. Actual and predicted relationship between IRL UV meter reading (10 scale) and weighted
UV spectral irradiance in the 280-320 nm (UV-B) region. Yn = UV-B sun equivalent and Y,
biologically effective UV (BUV) radiation in weighted mW*ra
Distance
from
sensor
cm
110
100
90
80
70
60
50
40
30
20
IRL UV Meter
Reading
(107 scale)
X
6.90
7.95
9.35
10.00
12.00
15.00
19.00
24.00
33.00
48.00
r
X
UV-B Sun
Determined
Yl
0.7806
0.8861
1.0331
1.2317
1.4758
1.7811
2.2037
2.8514
3.7911
5.4484
= 0.9991
Equivalent
Error of
Prediction
V*i
-.0478
-.0624
-.0755
.0487
.0640
.0261
-.0086
.0668
-.0231
-.0818
= IRL-jUV meter reading
Biologically Effective UV in
weighted mU-m~~
Actual
Y2
2.3885
2.7116
3.1613
3.7689
4.5161
5.4503
6.7433
8.7252
11.6008
16.6722
r = 0.9992
X = IRL UV meter reading
Error of
Prediction
Y2-y2
-.1491
-.1906
-.2272
.1547
.2073
.0996
.0034
.2488
-.0013
-.1394
'
(10 scale)
= predicted UV-B sun
equivalent
= 0.3900 + 0.1144X
(10 scale)
yป = predicted BUV (weighted mW-m~ )
y2 = 0.1412 + 0.3473X
-------
Table 7. Actual and predicted relationship between UV-B sun equivalent as measured with an Optronic
Model 725 broad band radiometer (0-10 scale and 0-5 scale) and unweighted UV spectral
irradiance (mW -m ) in the 280-320 nm (UV-B) region.
Distance
from
sensor
cm
110
100
90
80
70
60
50
40
30
20
UV-B Sun Equiv.
(0-10 scale)
Measured on
Optronic 725
Radiometer
Xl
0.8
1.0
1.1
1.4
1.6
2.0
2.5
*
3.2
4.3
6.1
UV-B Irradiance 2
in unweighted mW-m
Measured on
UV Spectro-
radiometer
Yl
206.7507
234.6552
274.8037
*ป
324.2766
392.0929
475.1478
591.2049
763.8546
1024.9800
1481.1394
Error of
Prediction
V*i
15.2720
-4.9999
11.0603
-11.7315
7.9083
-5.3898
-9.7739
-5.7420
-9.5873
12.9837
UV-B Sun Equiv.
(0-5 scale)
Measured on
Optronic 725
Radiometer
x2
0.9
1.1
1.2
1.5
1.7
2.1
2.6
3.3
4.4
off scale
UV-B Irradiance _ฃ
in unweighted mW- m
Measured on
UV Spectro-
radiometer
Y2
206.7507
234.6552
274.8037
324.2766
392.0929
. 475.1478
591.2049
763.8646
1024.9800
1481.1394
Error of
Prediction
Vy2
10.8111
-8.4680
8.0887
-13.2137
7.4191
-3.8931
-5.7949
1.7224
3.3283
r = 0.9996
X = UV-B sun equivalent
y = Predicted UV-B irradiance
y =-1.2273 + 240.8825X
r = 0.9995
X->= UV-B sun equivalent
-'
y? = Predicted UV-B. irradiance
y2 = -16.3863 + 235.9177X
-------
NJ
Table 8. Actual and predicted relationship between UV-B sun equivalent on Optronic Model 725
radiometer (0-10 scale and 0-5 scale) and biologically effective UV (BUV) radiation
in the 280-320 nm region.
Distance
from
sensor
cm
110
100
90
80
70
60
50
40
30
20
UV-B Sun Equiv.
(0-10 scale)
Measured on
Optronic 725
Radiometer
Xl
0.8
1.0
1.1
1.4
1.6
2.0
2.5
3.2
4.3
6.1
Biologically Effective
UV in weighted mW m~2 UV-Sun Equiv.
(0-5 scale)
Measured
Yl
2.3885
2.7116
~ 3.1613
3.7639
4.5161
5.4503
6.7433
8.7252
11.6003
16.6722
r = 0.9998
X = UV-B sun
Error of
Prediction
Yryi
.1348
-.0821
.0976
-.1048
.1024
-.0434
-.1005
-.0086
-.1031
.1082
equivalent
Measured on
Optronic 725
Radiometer
X2
0.9
1.1
1.2
1.5
1.7
2.1
2.6
3.3
4.4
off scale
obtained
\ r- * __
Biologically Effective
UV in weighted mW -m
Determined Error of
Measured
Y2
2.3885
2.7116
3.1613
3.7689
4.5161
5.4503
6.7433
8.7252
11.6008
16.6722
r = 0.9996
X-3 UV-B sun
Prediction
A
Y -y
2 y2
.0976
-.1110
.0728
-.1172
.0933
-.0310
-.0674
.0534
.0045
equivalent obtained
on Optronic Model 725 radiometer
(0 - 10 scale)
= Predicted BUV in the 280-320nm
region
= .0936 + 2.700X
(0-5 scale)
y = Predicted BUV in the 280-320nm
region
v^ = -0.1020 + 2.6587X
-------
I
NJ
V
Table 9. Actual and predicted relationship between UV-B sun equivalent as
measured with an Optronic Model 725 broad band radiometer (0-10
scale and 0-5 scale) and incident flux in the 230-320 nm region
measured in photons-m xl02l integrated over a 6 hour day.
Incident UV-B flux in
photons-nT2 X1021 ner 6 hr dav
Distance
from
sensor
cm
110
100
90
80
70
60
50
40
30
20
UV-B Sun
Equiv.
Optronic 725
(0-10 scale)
X
0.8
1.0
1.1
1.4
1.6
2.0
* 2.5
3.2
4.3
6.1
Total Spectral Calculated Error of
irradiance
280-320 nm
mW * in
206.7507
234.6552
274.8037
324.2766
392.0929
475.1478
591.2049
763.8646
1024.9800
1481.1394
Y
6.7459
7.6564
8.9664
10.5806
12.7934
15.5033
19.2901
24.9237
30.7494
48.3272
r = 0.9972
Prediction
A
Y -y
.4523
-.1697
.3741
.3105
.3699
.0149
-.0296
.2403
-2.3626
1.4229
X = UV-B sun equivalent
y = predicted
incident UV flux at
280-320 nm in photons-nf
xlO per 6 hour day
y = 0.1637 4-7.6624X
-------
I
ro
~i.-abltr-.tfci. ii~i.aal ki-vJ prfe-c^c-ted ^-lati.,-.^-nipU,_-Jweew. /-B LJ eqt__lien,
measured with an Optronic Model 725 broad band radiometer (0-5
scale) and incident flux in the 280-320 nm region measured in
photons-m~ xlO^l integrated over a 6 hour day.
Distance
from
sensor
cm
110
100
90
80
70
60
50
40
30
20
UV-Sun
Equiv.
Optronic 725
(0-5 scale)
X
0.9
1.1
1.2
1.5
1x7
2.1
2.6
3.3
. 4.4
off scale
Total Spectral
irradiance
280-320 nm
mW m
206.7507
234.6552
274.8037
324.2766
392.0929
475.1478
591.2049
763.8646
1024.9800
1481.1394
Incident UV-B"
photons -m xl
Calculated
Y
6.7459
7.6564
8.9664
10.5806
12.7934
15.5033
19.2901
24.9237
30.7494
48.3272
flux in
0 per 6 hr day
Error of
Prediction
Y - y
-.0365
-.3696
.0485
-.4728
.3164
.1790
.4066
1.0574
-.9470
r
X
y
0.9974
UV-B sun equivalent
predicted incident flux at 280-320 nm
in photons-m"2 x 10Z1 per 6 hour day
y = 0.3759 + 7.1183X
-------
FINAL REPORT
MULTIPLE EFFECTS OF UV-B IRRADIATION
ON FUNGAL SPORE GERMINATION
Olga v0 H. Owens
Donald T. Krizek
Plant Stress Laboratory
Plant Physiology Institute
Beltsville Agricultural Research Center
Beltsville, Maryland 20705
EPA-IAG-D6-0168
Project Officer:
i
R. J. McCracken
Agricultural Research, Science and Education Administration
U.S. Department of Agriculture
Washington, D.C. 20250
Prepared for
Environmental Protection Agency
BACER Program.-
Washington, D.C. 20460
-------
CONTENTS
Abstract ....................................... . ...........
Table [[[ iv
Figures ... ................ . ....................... . ........ v
Acknowledgments ................................. . .......... vii
1. Introduction .... ............... ........... . ....... 1
2 . Materials and Methods .... ......................... 2
UV Source ... ........... . ......... . ........... 2
Experimental Material .... .................... 3
3. Results .................. . ............ * ........... 5
Growth of the Spores ......................... 5
Survival Curves .............................. 5
Action Spectrum .............................. 6
Reciprocity .................................. 7
Character of 330 nm Inhibition ............... 7
Delay, Outgrowth Rate and Photoreactivation .. 8
Germ Tube Outgrowth in Relation to Sunshine
and Ozone Reduction ...... . ....... ..... ..... 9
4. Discussion ......... , .............................. 10
-------
ABSTRACT
We investigated the influence of narrow-band UV irradiation in the
265-330 nm region on germination of fungal spores of Cladosporium
cucumerinum Ellis & Arth. using a xenon arc lamp and various filters.
Based on survival curves and action spectra data, we propose that there
are two active regions between 280 and 320 nm (UV-B) that might be
influenced by changes in the stratospheric ozone layer: a short-wave
portion (265-295 nm) and a long-wave portion (300-330 nm). Action spectrum
data obtained with narrow-band interference filters confirmed previous
reports of damage to DNA from UV irradiation at 265-295 nm UV and in
addition demonstrated significant inhibitory effects of UV irradiation
at 300-320 nm. Further studies made of the 300-330 nm portion of the
spectrum using a combination of plastic and glass filters showed that the-
influence of UV irradiation in this region was primarily to produce a
non-photoreactivable delay in germ tube outgrowth. The implications of
these findings are discussed in relation to the possible impact of strato-
spheric ozone reduction.
iii
-------
TABLE
-2 -1
Table 1. Global downward energy fluence in J-m 5 nra for a 6-hr
period (assuming an average zenith angle of 30 )at three ozone
concentrations at standard temperature and pressure.
-------
FIGURES
1, Accumulated percent outgrowth of germ tubes of Cladosporium cucumerinura
fungal spores following inhibition and placement on a water agar plate
as described in the Materials and Methods. The two symbols are
replicate plates, and the vertical lines are one standard deviation.
The curve was drawn by inspection.
2. A, B, and C: Survival curves for 5 nm half-band width UV exposures
of Cladosporium cucumerinum fungal spores at the indicated central
wavelengths. Percent survival was determined at 22 hours and is
the percent of control survival. The fluences were obtained by
varying the time.
3. Action spectra for UV inhibition of fungal spore ge'rmination in
Cladosporium cucumerinum plotted from data of Figure 2 (A, B, and C).
9
Action is expressed as reciprocal of the joulesm~z that gave 90%
(o) and 37% (*) survival. The vertical lines are the 95% confidence
limits.
4. Percent survival of Cladosporium cucumerinum fungal spores following
1 ?
exposure to UV irradiation at 265 nm given at 0.1 J*s ซm (o) and
1 9
1.0 Jปs *m (). Counts were made as in Fig. 2.
t
5. Percent survival of Cladosporium cucumerinum fungal spores following
*
exposure to a broad band UV source centering on 325 nm (inset)
12 19
given at 80 Jปs -m (o) and 800 Jซs -1 ซm~^ (a). Counts were
made as in Fig. 2.
6. Survival curve for exposure of Cladosporium cucumerinum fungal spores
to a broad band UV source centering on 330 nm (inset) given at 400
J*s~-*- m~ , Counts were made at 22 hours as in Fig. 2.
7. Accumulated percent outgrowth of Cladosporium cucumerinum fungal
spores vs duration of exposure to broad band UV irradiation centering
-------
at 330 nra. The indicated values (360, 480, and 600) are kJ-nT2
obtained by duration of exposure to 400 Jซs"~l *m~2 for 15, 20, and
25 rain., respectively. Control spores received no UV irradiation.
8. Time course for outgrowth of Cladosporium cucumerinum fungal spores
showing comparative influence of short-wave and long-wave UV.
_i 2
A. 275 nm narrow-band UV (5 min. 0.57 J*s *m ; SS^, ri).
-1 7
B. 330 nm broad-band UV (5 min. 520 Jซs ซm ,
-------
^ JIUซUB1 I.
ACKNOWLEDGMENTS
The authors gratefully acknowledge Mr. Karl H. Norris for his help
in the use of the spectroradiometer; Mr, Scott J. Ravitz for his
assistance in maintaining and supplying cultures; Mr. William A. Dungey
for help with the xenon arc lamp; Dr. Joseph H. Graham for suggesting
the use of C, cucumerinum; Mr. Richard H. Thimijan for advice on
statistical analysis of the data; and Drs. Sterling B. Hendricks,
Takuma Tanada, and Hugh Sisler for their critical review of the manuscript.
The research was supported by EPA Contract No. EPA-IAG-D6-0168.
vii
-------
Multiple Effects of UV-B Irradiation on Fungal Spore Germination
I/ 21
. Olga v.H. Owens- and Donald T. Krizek-
INTRODUCTION
The possibility of stratospheric ozone reduction has increased concern
about the potentially harmful effects of irradiation in the spectral region
where ozone absorption normally cuts off the sun's ultraviolet radiation.
This region (UV-B) is defined as the 280-320 nm region.
The lethal and mutagenic effects of UV absorption at the shorter
wavelengths, 250-280 nm (UV-C), are well-documented and are associated
with DNA and protein absorption. Previous studies on the effects of UV
irradiation showed that the 320-400 nm (UV-A) region acts on a variety of
metabolites to produce lethal, inhibitory, and delaying effects (l^ 2, 3,
6, 7, 15). Between these two active regions is the UV-B region where there
are few major absorption peaks. Because any effect observed at the peak of
absorption of a photo-chemically active compound would be observed also in
the region of lesser absorption, it seems reasonable to expect that observed
effects of UV-A and UV-C irradiation would overlap in the UV-B region.
i'Formerly Plant Physiologist, Agricultural Research, Science and
Education Administration. Presently Associate Program Manager,
Competitive Research Grants Office, USDA/SEA, 1300 Wilson Blvd.,
Arlington, VA 22209.
Plant Physiologist, Agricultural Research, Science and Education
Administration.
-------
Another major characteristic of the UV-B region is the 5-decade
decrease in the sun's irradiance from 320-280 nm caused by the sharply
increasing absorption coefficient of ozone. It is evident that a reduction
in stratospheric ozone concentration could shift this sharp cut-off to
shorter wave-lengths, and increase UV irradiance on the short wave side
of the cut-off region.
i
In order to assess the biological effects of this increase in UV
irradiance, we need to estimate the effects at those wavelengths where
changes in the ozone concentration will have a major effect on the sun's
fluence at the earth's surface. In this report we show that for germination
of non-proliferating fungal spores (conidia), the action of UV irradiance
from 265-295 nm on survival is closely correlated with nucleic acid
absorption. From 300-330 nm the response resembles that reported for the
near UV region (8, 9) and presumably involves a delay in protein synthesis.
MATERIALS AND METHODS
. . - . 3/ ' "
UV Source; The UV source (Schoeffel) was a 2.5 kW high-pressure xenon
arc lamp equipped with collimating lenses, a 15 cm circulating cooled
water filter and a mirror at a 45 angle to produce a vertical beam. To
provide narrow band irradiation we placed various filters in the parallel
beam: The first filter was a 1 cm deep, 1:1 mixture of saturated
solutions of NiSO -6H 0 and CSO 7H?0 in an open dish with parallel sides
and a flat quartz bottom. In later experiments the mixture was diluted
i
3/
Mention of a trademark, proprietary product, or vendor does not
constitute a guarantee or warranty of the product by the U.S. Department
of Agriculture and does not imply its approval to the exclsuion of other
products or vendors that may also be suitable.
- 2 -
-------
further with dis ti lied water that had no detectable effect on the
transmission characteristics. The dish was kept below 50ฐC with a
cooling coil around the outside. The second filter was either a 5 nm
half-band width interference filter (Corion) of the selected central
wavelength or a combination of plastic and glass filters for the broader
bands: for 325 nm, 2 mm pyrex and 0.13 mm cellulose acetate; for 330 nm
Corning Nos. 0-54 and 7-54.
The beam was directed from the mirror through the NiSO.-CoSo.
44
solution, then through the secondary filter onto a 15 rpm turntable
containing the test organisms as described below. The UV irradiance was
measured with a pyroelectric radiometer (Molectron) that could be moved
into the beam as needed. From just below the mirror the beam was
surrounded by a black box maintained at 21-23 C. A tube surrounded the
beam below the filters, and baffles prevented stray radiation from falling
on the radiometer or the test organisms.
Experimental Material; The test organisms were conidiospores of the fungus,
Cladosporium cucumerinum Ellis & Arth., a leaf pathogen of cucumbers. We
used this species because of its ease in cultivation and handling. Its
spores normally give 95-98% germination and under natural conditions are
exposed in the imbibed state (from dew) to the sun's irradiance during their
germination period on the leaf surface.
Cultures of the fungus were maintained on potato-dextrose agar. Spores
were harvested from 6-day-old colonies by addition of 10 ml of distilled
water to the colony and agitated to obtain a uniform suspension. From the
suspension 30 ul was dropped onto the center of a water agar plate. The
-------
water was absorbed by the agar in about 20 minutes leaving an area of imbibed
spores, with a diameter of about 1 cm, in an even layer on the agar surface.
The spore area was one-third the diameter of the shielded, collimated beam
from the xenon arc. Following absorption of the water the covered plates
were held in darkness at 40ฐC for 15-30 minutes until the start of the
experimental period.
For exposure, the agar plate without a top was placed in the beam on
the turntable and exposed for a period of time that gave the specified photon.
fluence as determined from the power readings with the radiometer. Uncovered
matched control plates were placed in the same box but away from the beam
and shielded by mylar. For longer exposure times, viz. 1-6 hours, covered
control plates were also included in the box. There were no significant
differences in percent outgrowth between covered and uncovered controls.
Following exposure, the plates were covered and incubated in darkness
(unless otherwise specified) at 22 C. Germination of spores results in the
outgrowth of a germ tube. The presence of the germ tube was used as the
indication of survival (percent survival or percent outgrowth).
Observations of germ tube outgrowth were made at 150 x magnification
in a darkened room, and when necessary the microscope light source was
\
filtered through red and yellow colored plastic to prevent photoreactivating
*
light from reaching the spores. For each datum point all spores (about 100)
2
in each of five random areas (each 0.56 mm ) were counted and the proportion
with a distinct germ tube was recorded. Data were expressed as average
percent of spores with germ tube outgrowth ฑ the standard deviation. In
most cases the outgrowth produced on the exposed plates was expressed as
percent of the outgrowth of the control plates. For time course experiments,
the plates were removed at intervals for counts, then returned to the incubator.
-------
The method used for spore handling had several advantages: (1) the
spores were located on a flat surface minimizing scattering errors; (2)
microscopic examination allowed for elimination from the counts of over-
lapping and clustered spores; and (3) no additional nutrients were
necessary for germ tube outgrowth.
RESULTS
Growth of the Spores; The population of fungal spores on the agar plate
produced germ tubes over a period of time. Figure 1 shows the accumulated
percent of spores with germ tubes for replicate control plates determined
at intervals over a period of 25 hours. Outgrowth began after about 4 hours
and reached a maximum after 20 hours. The rate of accumulated outgrowth
increased and reached a maximum at about 9 hours, when 55-65% of the spores
had germ tubes, and then decreased. The accumulated percent outgrowth
superficially resembled a cumulative normal frequency distribution but
deviated significantly from normality in most cases.
Survival Curves; Figure 2 (A, B, and C) shows the influence of UV irradiance
of the indicated 5 nm bandwidths on outgrowth of germ tubes of Cladosporium
cucumerinum fungal spores. Percent of control outgrowth was determined
*
after 22 hours of incubation as described under Materials and Methods. The
-2
dose, expressed as joules'm , was obtained by varying the time of exposure.
The curves for 265-295 nm demonstrate the lack of an exponential relationship
between percent outgrowth and dose at low doses. The presence of the shoulder
can be attributed to an active dark repair mechanism. The curves for
300-320 nm show only a shallow linear relationship indicating that those
wavelengths were relatively ineffective.
-------
Because we are primarily concerned with the effects of solar
irradiation, we have concerned ourselves with the low dose parts of the
curves, located on the shoulders, corresponding to the sun's irradiance
at each wavelength examined.
Demonstration of measurable decreases in germ tube outgrowth at
310-320 nm required long exposure times of up to 6 hours. We could not
show the lower part of the survival curves with the use of the narrow
band UV filters.
Action Spectrum: Figure 3 shows two action spectra constructed from the
data in Figure 2 (A, B, and C). Action is expressed as the reciprocal of
_2
the joules-m . The upper curve shows the joules permitting 90% survival
which is on the shoulder of the survival curves. We assumed that the
survival curves were nearly linear to 70% and calculated a value for 90%
survival from the regression equations developed from the individual points.
This calculation introduced a small error and enlarged the 95% confidence
interval (vertical lines) but allowed us to make a single action spectrum
throughout the 265-320 nm region. The lower curve of Figure 3 shows an
action spectrum constructed from the 1/e values (37% survival) from those
survival curves that could be extended to 37% survival. These values
\
were obtained from curves drawn by inspection. The similarity of the two
ซ
curves in the 265-295 nm region (the short-wave portion) indicates that
the slope of the shoulder was proportional to the final slope and suggests
that the rate of dark repair was nearly proportional to the rate of damage.
The 265-295 nm action spectra closely match published spectra (13,16) and can
be attributed to damage to DNA.
Values on the 300-320 action spectrum are too high in relation to
265 nm to be attributable to DNA damage. We conclude from these results
that there are two active portions in the UV-B region: a short wave portion
-------
(265-295 rim) whose action corresponds to DNA absorption and a long-wave
portion (300-330 nm) whose action corresponds to an unknown component(s).
Reciprocity; Figure 4 shows the effect of 265 nm irradiation at two fluence
-1 -2
rates: 0.1 and 1.0 J-s ซm . For the low rate a neutral density filter
was used and the duration of exposure was increased 10-fold. The figure
shows that at the low fluence rate the UV irradiation was less effective
than at the high rate, indicating a greater efficiency of repair, The ratio
of the fluences at the low and high rates in producing a similar percent
survival was about 1.9. These data indicate that reciprocity was
incomplete.
We could not measure reciprocity at 300-320 nm using the narrow band
interference filters because of the long exposures that would be required
at low irradiances. Therefore we used a broad band filter, centering on 325
nm, described under Materials and Methods, to provide a higher fluence
rate in this spectral region. Figure 5 shows the influence of low (80 Jปs
2 1 ?
m ) vs high (800 J's -m ) fluence rates on spore survival. The inset shows
the transmission character of the filter determined spectrophotometrically.
Reciprocity was nearly complete, indicating that in the 325 nm portion of the
spectrum there was no detectable simultaneous repair mechanisms as that shown
t
for the 265 nm band.
Character of 330 nm Inhibition; In the 325 nm experiment described above,
the filters used allowed significant transmission below 300 nm. We, therefore,
used another filter combination (see Materials and Methods), centering on
330 nm that gave <1% transmission at 300 nm and above 370 nm (inset, Fig. 6).
Figure 6 is a survival curve obtained with the 330 nm broad band filter.
As for exposures at 265 nm, the points represent counts at 22 hours after
-------
exposure. We noticed, however, that several hours later additional spores
had germinatedan effect not observed at any of the other wavelengths,
-1 -2
The influence of 330 irradiation (AGO J-s -m ) on percent outgrowth
was determined at intervals over a period of time for control and irradiated
spores (Fig. 7). Three fluences were chosen to coincide, respectively, with
three points on the survival curve shown in Fig. 6: (1) on the shoulder (360
-2 -2
kJ-rn ); (2) where the curve bends (480 kJ-m ); and (3) on the steep part
_2
of the curve (600 kJ-m ). The figure shows that broad band UV irradiation
centered at 330 nm delayed the start and depressed the rate of outgrowth.
Thus, the data in Fig. 7 indicate that the "survival curve" of Fig. 6 is
an indication of both a delayed and a depressed rate of outgrowth; the shape
of the curve depended on the time at which spores were counted,
Delay, Outgrowth Rate, and Photoreactivation; We compared the effect of
narrow-band irradiation at 275 nm (representing the short-wave portion), to
the effect of broad-band 330 nm irradiation, (representing the long-wave
portion). Both exposures were for 5 min. The fluence rates were 0.57
-2 -1 -2 -1
J-m -s and 520 J-m -s , respectively. Both fluences were on the
shoulders of the percent survival curves. Following exposure, replicate
plates were incubated in darkness or under a bank of 1500 raA cool white
\
fluorescent lamps filtered with 0.127 mm (0.005 in) Mylar (short-wave UV
*
cut-off, 3% T at 320 nm). Controls received no UV exposure. Percent
outgroxtfth, shown in Fig. 8, was determined at the intervals indicated in
Fig. 7.
Figure 8 shows that the incubation light inhibited the rate of outgrowth
of the controls, but did not delay onset of outgrowth. The fluorescent
lamps used for incubation contained, in addition to the visible radiation,
-1 -2 -1 -2
UV irradiation at 365 nm (95 mJ-s -m ) and at 334 nm (9 mJ-s -m ).
-------
This irradiation may have produced a cumulative inhibitory effect during the
incubation period. Figure 8B shows that exposure to broad band 330 nm
followed by either dark or light incubation significantly delayed onset of
outgrowth in comparison with the controls. After the delay the rates of out-
growth in dark equalled those in the light. Delay in outgrowth of spores,
incubated in either light or dark, was greater from the 275- (Fig. 8A) than
from the 330-nm (Fig. 8B) treatment. With spores treated at 330 nm, light
during incubation shortened the delay and increased the rate of outgrowth to
a value equal to that for the light-incubated control. The results of this
experiment and previous ones showed that the effects of the short-wave and
long-wave portions of the UV region studied were similar in that both delayed
the start of outgrowth of spores. Effects differed between the long-wave and
short-wave portions. With the long-wave portion the delay in the start of
outgrowth was not affected by light incubation and UV irradiation had little
effect on the rate of outgrowth except at high doses (Fig. 7:).
Germ Tube Outgrowth in Relation to Sunshine and Ozone Reduction: By use of the
regression equations developed from data in Fig. 2, we determined the effective-
ness of each 5 nm wavelength band of solar irradiation in inhibiting germ tube
outgrowth. The data of Shettle et al. (11) for solar irradiance were used as
a basis for the calculations.v We used values for dowm^ard global flux, 30
zenith angle and, ozone concentrations of. 0.32 atnrcm, 0.28 atm-cm, 0.16 atm-cm.
Assuming that the fungal spores would be exposed to a maximum of 6 hours of
sun in June at the Washington, B.C. latitude, average zenith angle 30ฐ,
2 1
we converted the data to J-m .5 nm for 6 hours. These values are
shown in Table 1. Figure 9 shows the calculated percent inhibition that
would be expected for the irradiance values at the three ozone concentrations
of Table 1. The inhibition assumes no photoreactivation and does not distinguish.
-------
between lethality and temporary delay of spore outgrowth. At 0.32 atnrcm,
an approximation of "normal" ozone, i.e. no reduction, the sun's irradiance
throughout the 300-320 nm region is sufficient to produce a 5% inhibition
at each wavelength, a result also noted by Jagger (5) for the near UV region.
A reduction in stratospheric ozone concentration to 0.28 attn'cm would not
significantly inhibit germ tube outgrowth, but a reduction to 0.16 atra-cm,
a 50% depletion would produce a significant inhibition of nearly 20% at
295-300 nm. The inhibition at 295 nm was somewhat under-estimated because
the fluence value was slightly beyond the survival curve shoulder (Fig. 2, B).
The effectiveness of the 295-300 nm region in inhibiting germ tube outgrowth
of Cladosporium spores agrees with the analysis of Setlow (10) and approxi-
mates that of Elkind et al. (2) for effectiveness of normal sunlight in
skin cancer production.
DISCUSSION
The fungal spore produces the germ tube after a period of germination
that is primarily a period of protein synthesis. In our study we used lack
of appearance of the germ tube as a measure of damage to the protein-
synthesizing system of the spore. Most fungal spores have a complete
system for protein synthesis in which enzymes increase in activity and amount
during the termination period (12). Protein synthesis and germ tube outgrowth
*
are inhibited by cycloheximide indicating involvement of a ribosomal system.
New RNA synthesis occurs, but the extent of the requirement for new RNA has
not been established. DNA replication does not occur until the time of
outgrowth or after (4,13), but our results indicate that DNA must function at
some stage of the germination sequence.
10
-------
We propose that there are two active regions between 280 and 320 nm.
Our results suggest that the short-wave portion primarily affects DNA and
the long-wave portion is primarily related to effects of near UV noted by
other investigators. Because the DNA-absorbing part indicates the presence
of both a dark repair mechanism and photoreactivation, a moderately
increased irradiance from the sun might be ineffective in producing an
immediate inhibition. Enhanced UV irradiance from the sun, however, might
retard the rate of growth, observable even at normal sunlight, that would
not be repaired in either darkness or light. We have shown that the
critical region for major effects, whether lethal or delaying, is at 295-
300 nm, a region that should be thoroughly analyzed.
11
-------
Literature Cited
1. Doyle, R. J. and H. E. Kubitschek. 1976. Near ultraviolet light
inactivation of an energy-independent membrane transport system in
Saccharomyces cerevisiae. Photochera. Photobiol. 24:291-293.
2. Elkind, M. M., Antun Han, and Chin-Mei Chang-Liu. 1978. 'Sunlight'-
induced mammalian cell killing: A comparative study of ultraviolet
and near-ultraviolet inactivation. Photochera. Photobiol. 27:709-715.
3. Fong, F., J. Peters, C. Pauling, and R. L. Heath. 1975. Two mechanisms
of near-ultraviolet lethality in Saccharomyces cerevisiae; A respiratory
capacity-dependent and irreversible inactivation. Biochem. Biophys. Acta
387:451-460.
4. Hollomon, D. W. 1973. Protein synthesis during germination of
Peronospora tabacina conidia: An examination of the events involved
in the initiation of germination. J. Gen. Microbiol. 78:1-13.
5. Jagger, J. 1975. Inhibition by sunlight of the growth of Escherichia
coli B/r. Photochera. Photobiol. 22:67-70.
6. Jagger, J. 1976. Effects of near-ultraviolet radiation on microorganisms.
Photochem. Photobiol. 23:451-454.
7. Koch, A. L., R. J. Doyle, and H. E. Kubitschek. 1976. Inactivation of
membrane transport in Escherichia coli by near-ultraviolet light.
*
J. Bacteriol. 126:140-146.
8. Rambhadran, T. V. 1975. The effects of near-ultraviolet and violet
radiations (313-405 nm) on DNA, RNA, and protein synthesis in E,. coli
B/r: implications for growth delay. Photochem. Photobiol.
22:117-123.
12
-------
9. Ramabhadran, T. V. and J. Jagger. 1975. Evidence against DNA as
the target for 334 nm-induced growth delay in Escherichia coli.
Photochem. Photobiol. 21:227-233.
10. Setlow, R. B. 1974. The wavelengths in sunlight effective in
producing skin cancer: A theoretical analysis. Proc. Nat. Acad. Sci.
U.S. 71:3363-3366.
11. Shettle, E. P., M. L. Nack, and A. E. S. Green. 1975. Multiple
scattering and influence of clouds, haze, and smog on the middle UV
reaching the ground. JEH Nachtwey, D.S. (ed.). Impacts of Climatic
Change on the Biosphere, CIAP Monograph 5, Part 1 - Ultraviolet
Radiation Effects DOT-TST-75-55. pp 2-38 to 2-49.
12. Sussman, A. S. and H. A. Douthit. 1973. Dormancy in microbial
spores. Ann. Rev. Plant Physiol. 24:311-352.
13. Tyrrell, R. M. 1978. Solar dosimetry with repair deficient bacterial
spores: Action spectra, photoproduct measurements and a comparison
with other biological systems. Photochem. Photobiol. 27:571-579.
14. Van Etten, J. L., L. D. Dunkle, and R. H. Knight. 1976. "Nucleic acids
and fungal spore germination" in Weber, D. J. and W. M. Hess (eds.)
The Fungal Spore, Form and Function, Wiley, N.Y. p. 243-299.
15. Webb, R. B. 1977. Lethal and mutagenic effects of near-ultraviolet
radiation, In Photochem. Photobiol. Reviews, Vol. 2 (K. C. Smith,
ed.) P.'Ierium Publishing Corp., N.Y. '.
16. Webb, R. B. and M. S. Brown. 1976. Sensitivity of strains of
Escherichia coli differing in repair capability to far UV, near UV,
and visible radiations. Photochem. Photobiol. 24:425-432.
13
-------
Table 1. Global downward energy fluence in J'm~2ซ5nin for a 6-hour
period (assuming an average zenith angle of 30ฐ) at three
ozone concentrations at standard temperature and pressure.
Values calculated from Shettle et al. (1975).
Wavelength
5 nm band
320
310
300
295
290
280
0.32 atra-cm
3.44 x 104
1.19 x 104
5.62 x 102
1.76 x 101
3.31 x 10~2
2.27 x 10-16
Ozone Concentration
0.28 atm-cm
3.59 x 104
1.37 x 104
9.08 x 102
4.22 x 101
1.67 x 10"1
5.99 x 1Q-14
0.16 atm'cia
4.07 x 104
2.10 x 104
3.81 x 103
5.92 x 102
2.21 x 101
1.09 x 10~6
14
-------
100
10 !5
TIME, MRS.
25
Figure 1. Accumulated percent outgrowth of germ tubes in Cladosporium
;
cucumerinum fungal spores following inhibition and placement
on a water agar plate as described in the Materials and
Methods. The two symbols are replicate plates, and the
vertical lines are one standard deviation. The curve was
drawn by inspection.
15
-------
100
30
w 10
(-
z
Ul
o
CC
w
a 3
I
- 265
100
0246
.M-2 (XIO-2)
0 10 20
J-M-2 (X IO'2)
FLUENCE
100
80
CC.
-------
10
-I
10
-2
5
CO
O
IO
-3
"3
ง ,0-4
O
UJ
oc
10
-5
10
-6
260 280 300 320 340
WAVELENGTH, NM.
Figure 3. Action spectra for UV inhibition of fungal spore
germination in Cladosporium cucumerinum plotted
from data of Figure 2 (A, B, and C). Action is
-2
expressed as reciprocal of the joules-m that
gave 90% (o) and 37% (o) survival. The vertical
lines are the 95% confidence limits.
17
-------
100
90
80
70
6O
UJ
UJ
Q.
50
40
30
A.
r
I I I
0 50 100 150 200 250
FLUENCE, J-M-2
Figure 4. Percent survival of Cladosporium cucumerinum fungal
spores following exposure to UV irradiation at 265
-2
nm given at 0.1 J-s -m (o) and 1.0 J-s
Counts were made as in Fig. 2.
~2
m (ป) .
18
-------
100
90
80
70
CO
h-
550
o
UJ
Q.
40
30
%T
60
40
20
280 320 360
0 100 200 300
FLUENCE, KJ-NT2
Figure 5. Percent survival of Cladosporium cucumerinum fungal
spores following exposure to a broad band UV source
-*1 2
centering on 325 nm (inset) given at 80 Jซs ซm (o)
-1 -2
and 800 J-s -m (e). Counts were made as in Fig. 2.
19
-------
100
80
60
40
CE
3
CO
10
Z 8
Ui
ฃ ^
bJ
0.
%T
40
30
20
10
300 320 340 360
X.NM.
ZOO 400 600
FLUENCE, KJ-M'2
800
2400
Figure 6. Survival curve for exposure of Cladosporium cucumerinum
fungal spores to a broad band UV source centering on 330
1 2
nm (inset) given at 400 J-s -m . Counts were made at
22 hours as in Fig. 2.
20
-------
too
10 15 20 25 30 35 40 46 55
TIME, HRS.
Figure 7. Accumulated percent outgrowth of Cladosporium cucumeririum
fungal spores vs duration of exposure to broad band UV
irradiation centering at 330 nm. The indicated values
-2
(360, 480, and 600), are kJ-m obtained by duration of
-1 -2
exposure to 400 J-s -m for 15, 20, and 25 min.,
t
respectively. Control spores received no UV irradiation.
21
-------
10 15 20 25
15
TIME, MRS.
20 25
Figure 8. Time course for outgrowth of Cladosporium cucumerinum
fungal spores showing comparative influence of short-
wave and long-wave UV. A. 275 nm narrow band UV (5 min.
057 J-s"1- ; ga, O). B. 330 nm broad band UV (5 min. 520
1 2
J'S~ -m ; <ฃ*ป m <ฃ>). Controls (no UV exposure: , o).
*
Plates were incubated in dark (closed symbols) or in
light, (open symbols).
22
-------
H
5
O
O
< ID
-J O
< UJ
O O
85
a:
UJ
G- 75
280
300
320
Figure 9. Influence of stratospheric ozone reduction on percent
germ tube outgrowth of Cladosporium cucumerinum fungal
spores in the UV-B region. Values were calculated from
the data of Fig. 2 and Table 2. The values assigned to
the curves represent "normal" ozone concentration, 0.32
atm-cm; 12% reduction, 0.28 atm-cm; and 50% reduction
0.16 atm-cm.
23
-------
FINAL REPORT
RESPONSE OF SELECTED VEGETABLE AND AGRONOMIC CROPS TO
INCREASED UV-B IRRADIATION UNDER GREENHOUSE CONDITIONS
John E. Ambler
Randy A. Rowland
Nancy K. Maher
Plant Stress Laboratory
Plant Physiology Institute
Beltsville Agricultural Research Center
Beltsville, Maryland 20705
EPA-IAG-D6-0168
Project Officer:
R. J. McCracken
Agricultural Research, Science and Education Administration
U.S. Department of Agriculture
Washington, D. C. 20250
Prepared for
Environmental Protection Agency
BACER Program
Washington, D. C. 20460
-------
ABSTRACT
Biomass measurements for alfalfa, rice, and wheat in greenhouse
experiments showed no reduction in growth of enhanced UV-B treated plants
as compared with the Mylar control. None of these species showed chlorotic
_o
tissue. However, the 'Poinsett1 cucumber cultivar under 13 to 15 mWra"
BUV of UV-B showed chlorosis of about 11 percent of the total leaf tissue,
whereas, 'Ashley* showed chlorosis of about 1 percent of tissue on a dry
weight basis. Leaf area and weight of both cultivars showed about a
7 percent reduction in response to enhanced UV-B as compared to the Mylar
control.
For each species studied, there seemed to be a shade effect due to
the experimental design of the set-up. Biomass yields were higher from
plants at the perimeters than at the centers of the experimental areas.
Mention of a trademark name or a proprietary product does not constitute
a guarantee or warranty of the product by the USDA and does not imply
its approval to the exclusion of other products that may also be available.
-------
INTRODUCTION
Recent greenhouse and growth chamber studies conducted in the Plant
Stress Laboratory (PSL) (1, 3, 5, 6, 9) have demonstrated the inhibitory
effects of high levels of biologically effective UV (BUY) radiation on
plant growth and development of cotton and selected vegetable species.
We conducted our experiments to extend these studies to other
plant species including alfalfa, cucumber, rice, and wheat. We examined
the effects of UV-B on vegetative growth and biomass production in alfalfa,
cucumber, rice, and wheat and on grain production in wheat.
MATERIALS AND METHODS
The UV-B enhancement facilities were developed in cooperation with
the Agricultural Equipment Laboratory (AEL), Beltsville Agricultural
Research Center (BARC). Enhancement studies were conducted according to
the guidelines established for the BACER program (2, 4). Two lamp fixtures
provided UV radiation, each containing two Westinghouse FS-40 fluorescent
sunlamps filtered with 5 mil Mylar (UV-A) or 5 mil cellulose acetate (UV-A,
-B). The UV lamps were kept 62 cm from the plant canopy, and the lamp
fixtures were 60 cm apart. Filters were aged for 6 hours before use and
were changed twice weekly (4). We measured UV at every pot location at the
\
start of the experiments and at selected points periodically thereafter
with either an Optronic Laboratories, Inc., Model 725, UV-B Radiometer or
an Instrumentation Research Laboratory (IRL) UV-B Radiometer (7, 8).
Radiometer readings were verified by spectral irradiance determinations
(250-369 nm) with an automated spectroradiometer (7, 8) at selected locations
in the experimental irradiation areas.
_o
Weighted irradiance levels are reported as mWm BUV, the biologically
effective UV derived from the AE9 weighting function, and unweighted
-------
-2
irradiance as Wra obtained by summing the measured or calculated values
-9 -2
at each nanometer from 280-320 nm. Dividing mWm BUY by 3.06 (the mWm
BUV of the Beltsville control sunshine) provides the fraction of BUV received
by each plant relative to that of one control sunshine (10).
Where UV irradiation was obtained by filtering the FS-40 lamps through
cellulose acetate (CA), BUV was limited to the UV-B region (280-320 nm).
For details concerning average control sunshine, spectral character-
istics of UV fluorescent lamps and filters, and the weighting function, see
the BACER final reports of the AEL and IRL, BARC (8, 10).
Experimental plants receiving UV-B were combined into groups averaging
7 mWnT2 BUV (3.1 Wm~2), 10 mWnT2 BUV (4.5 Wm~2), 13 mWm"2 BUV (5.8 Wm~2)
_2 _9
vand 15 mWm BUV (6.7 Wm ) for comparison of the results.
Cultural Procedure (General)
Plants were seeded in 12.5-cm white plastic pots containing a commercial
peat-vermiculite mix, and they were thinned on emergence as follows: alfalfa,
5 plants; cucumber, 1 plant; rice, 8 plants; and wheat, 8 plants per pot.
Plants were grown in a glass greenhouse with and without supplemental
UVB during the summer-fall 1977. They were irradiated daily from emergence
for 6 hours from 1000 to 1600 hours.
Plant material was dried* in a forced-draft oven at 70ฐ C, weighed and
_2
combined into the four groups receiving the average 7, 10, 13, or 15 inWm
BUV under the CA filter and compared to matched set of control plants placed in
like groups for similar numbered positions grown under Mylar filters.
RESULTS
I. UV-B effects on alfalfa (Medicago sativa L.)
Procedure: Alfalfa cv. 'Williamsburg* was seeded on August 8, 1977.
Biomass was harvested by cutting the plants 5.1 cm above the soil level,
6 weeks, 9 weeks, and 12 weeks from seeding.
-------
Results: The results showed no effect from enhanced UV-B (Table 12).
Table 12. Effects of UV-B on biomass (g) of alfalfa*
7mWm~2 BUV lOmWm"2 BUY 13tnWm~2 BUY
Harvest - wks. Harvest - wks. Harvest - wks.
Filter 6 9 12 6 9 12 6 9 12
Mylar 5.2 2.7 1.6 3.4 1.8 1.4 4.0 2.2 1.5
CA 6.4 3.0 2.2 4.9 2.1 1.2 4.5 1.9 1.4
15mWm~2 BUV
Harvest - wks.
6 9 12
3.2 1.7 1.4
3.8 1.9 1.6
* Mean weight 10 plants.
Discussion: A shade effect was notedbioraass yield was largely influenced
by pot position. Plants grown directly under light fixtures produced less
growth than plants not directly under the fixture.
II. UV-B effects on cucumber (Cucumis sativus L.)
Procedure: Pots were seeded alternately with 'Ashley1 or 'Poinsett*
cultivars on November 28, 1977, and thinned to one plant per pot on
emergence. Above-ground plant parts, excised at the cotyledonary node were
harvested 4 weeks after seeding. Leaf areas were measured and selected
plant sections were placed in a 1:1:18 FAA solution for morphological
examination.
*
Results: The effects of increased UV-B on cucumber are shown in
*
Table 13.
-------
Table 13. Effect of UV-B on leaf weight and leaf area in cucumber
mWra"2 BUV
UV-B* 7
irradiation
10
13
15
UV-B* 7
irradiation
10
13
15
CA
'Ashley1
.343
.244
.235
.209
CA
'Ashley1
194
129
132
114
. Dry weight (g)
filter
'Poinsett1
.345
.295
.244
.199
Leaf
filter
'Poinsett1
200
161
132
119
Mylar
'Ashley'
3.67
3.23
3.02
3.06
area
Mylar
'Ashley'
203
191
170
176
filter
'Poinsett'
4.34
3.84
3.40
4.06
filter
'Poinsett'
242
221
193
216
* Number of samples for 7, 10, 13, and 15 mWm BUV were 12, 16, 22, and 6,
respectively.
Discussion: Dry weights. Three effects were noted. A shade effect
from light fixtures, enhanced UV-B effect and a chlorotic leaf-tissue effect.
Yield of control plants (Mylar) directly under light fixtures was about
90 percent of the yield of plants not directly under light fixtures. This
was due to a shading effect.
Leaf dry weight of plants grown under the CA filters, as compared with
plants at like positions grown under the Mylar filters, were reduced 9, 8,
7, and 5 percent for 'Poinsett' and 9, 8, 8, and 7 percent for 'Ashley1,
i
respectively.
-------
_2
Chlorotic leaf tissue in the 'Poinsett' cultivar under 13-15 mWm
UV-B made up about 11 percent of the leaf tissue on a dry weight basis.
In 'Ashley' for similar BUY levels chlorotic leaf tissue comprised about
1 percent ,of the total leaf tissue on a dry weight basis.
Plants grown under Mylar filters (control) appeared green and normal
for all pot positions.
Leaf Area
Two effects were noted, a shade effect and an enhanced UV-B effect.
Leaf area of control plants grown directly under light fixtures was about
90 percent of the values of plants not grown directly under light fixtures.
Leaf areas of plants grown under the CA filters, as compared with plants in
like positions under Mylar filters, were reduced by 17, 27, 32, and 45 percent
for 'Poinsett' and 4, 32, 22, and 35 percent for 'Ashley1, respectively, with
increasing UV-B demonstrating varietal differences within species.
III. UV-B effects on rice (Oryza sativa L.)
Procedure: Rice was seeded in soil enclosed by a plastic bag within
a waxed container on August 8, 1977. Plants were thinned to eight plants
per pot and flooded 2 weeks after seeding. Flooded conditions were
maintained throughout the experiment. Plants were harvested on December 27,
1977, 19 weeks from seeding. *
Results: The results of increased UV-B in rice are shown in Table 14.
Table 14. Effect of UV-B on biomass (g) of rice*
Filter
Mylar
CA
7mWm~2 BUV
21.2
22.3
lOmWm"2 BUV
14.3
19.0
13mWm 2 BUV
15.7
15.5
15mWm~2 BUV
10.7
10.8
* Mean weight per pot.
-------
Discussion: One effect was noted, a shade effect which directly
affected the plants growing in the center of the quadrant. Plants on the
outer perimeter were not affected by shade and thus were greater in height
and growth. The inner plants were shaded by light fixtures and also shaded
by plants external to their pot positions.
IV. UV-B effects on wheat (Triticum aestivum L.)
Procedure: Wheat cv. 'Pacific Triple Dwarf was seeded in the summer
of 1977 and harvested 12 weeks after seedling emergence.
Results: The effects of increased UV-B on vegetative growth and grain
development were negligible (Table 15). Plant growth appeared to be green
and normal under both Mylar and CA filters.
Table 15. Effect of UV-B on growth and development in Pacific Triple Dwarf
wheat from emergence to age 12 weeks.
Straw
dry weight (g)
per 10 plants
Grain, No. seeds
per 10 plants
Grain weight (g)
per 100 seed
Filter
CA
Mylar
CA
Mylar
CA
Mylar
7mWm"~2 BUV
7.72
8.51
161
209
2.73
* 2.29
lOmton"2 BUV
6.49
7.35
137
161
2.72
2.37
13mWnT2 BUV
7.11
7.04
132
181
2.56
2.32
_2
15raWm BUV
7.27
7.45
140
161
2.69
2.46
-------
LITERATURE CITED
1. Ambler, J. E., D. T. Krizek, and P. Seraeniuk. . 1975. Influence of UV-B
radiation on the early seedling growth and translocation of Zn from
cotyledons in cotton. Physiol. Plant 34:177-181.
2. Cams, H. R., R. Thimijan, and J. M. Clark. 1977. Outline of irradiance
distribution of UV fluorescent lamps and combinations. Symposium on
Ultraviolet Radiation Measurements for Environmental Protection and
Public Safety. June 8-9, 1977. NBS, Gaithersburg, MD (Abstr.).
3. Krizek, D..T. 1975. Influence of ultraviolet radiation on germination
and early seedling growth. Physiol. Plant 34:182-186.
4. Krizek, D. T. 1977. Current UV measurement methodology and future
needs in photobiological research. Symposium on Ultraviolet Radiation .
Measurements for Environmental Protection and Public Safety. June 8-9,
1977, NBS, Gaithersburg, MD (Abstr.).
5. Krizek, D. T. 1978. Response of selected vegetables and agronomic crops
to increased UV-B irradiation under greenhouse and growth chamber conditions.
Final BACER Report submitted to the Environmental Protection Agency.
6. Krizek, D. T. 1978. Differential sensitivity of two cultivars of
Cucumis sativus L. to increased UV-B irradiance. Plant Physiol. Suppl.
61(4):92. (Abstr.). *
7. Norris, K. H. 1977. Development of a portable, automated UV-B
spectroradiometer. Symposium on Ultraviolet Radiation Measurements for
Environmental Protection and Public Safety. June 8-9, 1977. NBS,
Gaithersburg, MD (Abstr.).
8. Rowan, J. D. and K. H. Norris. 1978. Instrumentation for measuring
irradiance in the UV-B region. Final BACER Report submitted to the
Environmental Protection Agency.
-------
9. Skelly, J. M., M. F. George, H. E. Heggestad, and D. T. Krizek. 1978.
Air pollution and radiation stresses. In ASAE Monograph. Modification
of the Aerial Environment of Plants, Chapt. 2.5.
10. Thimijan, R. W., H. R. Cams, and L. E. Campbell. 1978. Radiation
sources and related environmental control for biological and climatic
effects UV research. Final BACER Report submitted to Environmental
Protection Agency.
-------
FINAL REPORT
RESPONSE OF SELECTED VEGETABLE AND AGRONOMIC CROPS TO
INCREASED UV-B IRRADIATION UNDER FIELD CONDITIONS
John E. Ambler
Randy A. Rowland
Nancy K. Maher
Plant Stress Laboratory
Plant Physiology Institute
'Beltsville Agricultural Research Center
Beltsville, Maryland 20705
EPA-IAG-D6-0168
Project Officer:
R. J. McCracken
Agricultural Research, Science and Education Administration
U.S. Department of Agriculture
Washington, D. C. 20250
Prepared for
Environmental Protection Agency
BACER Program
Washington, D. C. 20460
-------
ABSTRACT :
Enhanced UV-B radiation under field conditions on various economic
crops conducted at the Beltsville Agricultural Research Center ranged
from ambient up to eight times ambient. The enhanced radiation was
achieved with either unfiltered BZS-CLG or FS-40 Westinghouse sunlamps.
All crops were affected less by the enhanced radiation than similar
crops grown under greenhouse and growth chamber conditions.
Although variable, these results supported results obtained else-
where in this BACER Program which indicate that higher plants have a high
light energy-requiring system of photorepair or photoprotection. With
all crops investigated, additional research is needed to establish the
levels of UV-B required to injure plants in the field.
Mention of a trademark name or a proprietary product does not constitute
a guarantee or warranty of the product by the USDA and does not imply
its approval to the exclusion of other products that may also be available.
-------
INTRODUCTION
Growth chamber and greenhouse experiments have indicated that several
crop plants displayed reduced growth and chlorosis when subjected to
enhanced levels of UV-B (1, 2, 3, 4). The field experiments reported
here were designed to examine the responses of selected crops to enhanced
UV under field conditions.
MATERIALS AND METHODS
The field selected for these studies was located on the south farm of
the Beltsville Agricultural Research Center, Beltsville, Maryland. The
plots were silt loam, characteristically consisting of recently deposited
materials washed from acid crystallite rock of the capitol Piedmont.
These soils also have a concentration of fine mica, which contributes
to poor drainage. The area was rototilled in early spring of 1977 and
fertilized with a 10-10-10 fertilizer at a rate of 500 pounds/acre 3 weeks
before planting.
The total area was divided into six plots, which were supplied with
separate UV-enhancement assemblies. The lamp assemblies, constructed
by the Agricultural Equipment Laboratory, were designed so that height
adjustment could be easily maintained by a simple pulley system (Fig. 1).
A height of 1.6m was mainta&ied above the plant canopy during the course
of the experiment. :
For the first experiment, lamp banks were designed to obtain a two
dimensional gradient of UV irradiationone parallel to a lamp assembly,
the other at right angles to the lamp assembly. Each assembly consisted
of lamp fixtures placed end to end as follows: A two-lamp fixture at the
high UV-B end; then two single lamp fixtures placed end to end; a 33-cm
-------
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-------
space and another single lamp; a 65-cm space and a final lamp that was
taped to reduce the UV-B irradiance by one half. This positioning of the
lamps provided a uniform gradient of supplemental UV within the experimental
plot area. Each plot consisted of four 11.0-m rows parallel to the UV-B
gradient. Two rows were 0.5 m from the center, line of the lamp assembly,
the other two were 1.5 m from the assembly center line. Supplemental
UV radiation was provided by unfiltered Westinghouse BZS-CLG 40 watt
fluorescent lamps. Exposure time from emergence to harvest was 6 hours/day
from 1100 to 1700 hours. Table 1 shows the irradiance levels obtained
for each meter of row length.
-2
Weighted irradiance levels are reported as mWm BUV, the biologically
effective UV derived from the AZ9 weighting function, and unweighted
2
irradiance as mWm obtained by summing the measured or calculated values
7 2
at each nanometer from 280-320 nm. Dividing mWm * BUV by 3.06 (the mWm
BUV of the Beltsville control sunshine) provides the fraction of BUV
received by each plant relative to that of one control sunshine (6).
Two different crops were chosen for each plot to minimize shading
effects, and they were grown in alternate 0.5- and 1.5-ra rows. The paired
crops were as follows: squash (Curcurbita maxima (L.) cv. 'Early Prolific
Straightneck') and bean (Phaseolus vulgaris (L.) 'Contender Bush'); sweet
corn (Zea mays (L.) cv. 'Golden Cross Bantuin') and sorghum (Sorghum
biocolor (L.) Moench cv. 'R-720'); and soybean (Glycine max (L.) Merr.
cv. 'Amsoy-71') and sugar beet (Beta vulgaris (L.) cv. '7322-0'). Also
included were guard rows of pea (Vigna unguiculata (L.) Walp. cv. 'Cow')
on either side of the 1.5-m rows. Each plot of paired crops was duplicated.
All plants were sown in May unless otherwise specified.
A second experiment was undertaken in mid-summer using unfiltered
Westinghouse FS-40 UV fluorescent lamps. A similar gradient assembly was
-------
Table 1. Spectroradioraeter measurements at 1-m intervals along the 11-m
rows for BZS-CLG sunlamps unweighted (mWm~J and weighted
BUY) and fraction of control sunshine above ambient.
Row
position
(m)
__
mWm
-2
mWra BUV
Control
sunshine
(fraction)
0.5-m Row
0
1
2
3
4
5
6
7
8
9
10
11
9.113
96.131
138.105
117.339
94.996
79.977
63.131
42.018
28.319
7.562
1.768
.662
.146
1.669
2.498
2.131
1.736
1.450
1.137
.898
.518
.124
.026
.009
.05
.55
.82
.70
.57
.47
.37
.29
.17
.04
.01
.00
1.5-m Row
0
1
2
3
4
5
6
7
8
9
10
11
12.109
45.701
65.431
76.497
66.966
53.034
32.011
29.146
19.644
5.246 *
1.227
.459
.193
.790
1.175
1.553
1.146
.956
.547
.491
.283
.068
.014
.004
.06
.26
.38
.51
.47
.31
.18
.16
.09
.02
.00
.00
-------
Table 2. Spectroradioraeter measurements at 1-m intervals along the 14-m
rows for FS-40 sunlamps unweighted (mWm ) and weighted
BUV) and fraction of control sunshine above ambient.
Row
position
(m)
o
mWm *
mWm~2BUV
Control
sunshine
(fraction)
0.5-m Row
0
1
2
3
4
5
6
7
8
9
10
11
12
13
14
88.672
. 450.287
709.471
607.306
512.496
421.542
347.719
232.890
163.677
60.898
17.505
8.025
4.263
1.583
.432
3.227 ,
17.594
28.736
24.280
19.879
17.035
14.271
9.473
6.763
2.333
.636
.292
.155
.058
.015
1.05
5.75
9,39
7.93
6.50
5.57
4.66
3.09
2.20
.76
.21
.09
.05
.02
.00
1.5-m Row
0
1
2
3
4
5
6
7
8
9
10
11
12
13
14
80.56
409.287
310.778
266.025
251.797
207.110
105.920
93.942
80.403
29.915
13.132
6.020
3.198
1.188
.324
3.182
7.566
12.357
11.889
9.734
5.132
4.299
3.617
* 3.296
1.857
.478
.193
.137
.041
.010
1.03
2.47
4.04
3.89
3.18
1.68
1.40
1.18
1.08
.61
.16
.06
,.05,
..01
iOO
-------
constructed with twice as many'lamps, which substantially increased the
supplemental UV irradiation. Rows were 14 ra long. Table 2 shows the
irradiance levels obtained for each meter of row length in the plot area.
Radiation was measured with a single monochrometer spectroradiometer
described in the Instrumentation Research Laboratory final report (5).
Data were obtained at 1-nm intervals from 250 to 369 nm. Experimental
results were subjected to analyses of variance.
RESULTS AND DISCUSSION
Table 3 gives the temperature and percipitation means for the duration
of the experiments. There were no unexpected temperature or rainfall
extremes, but variability from row to row and within rows was considerable
in all experiments, which may account for the lack of statistical significance
in many of the parameters measured. Because of this variability, Probability
Values (P) of 0.3 and lower are shown for all measured responses.
BZS-CLG lamp assemblies used in the first experiment plus sunshine provided
_o
total average irradiance levels as follows; 3.1, 3.7, and 4.2 mWm BUV
for the 1.5-m row and 3.2, 4.2, and 5.1 mWm~2 BUV for the 0.5-m row. These
irradiances provided approximately 1.0, 1.2, and 1.4 times the Beltsville
control sunshine for the 1.5-m row and 1, 1.4, and 1.7 times for the
0.5-m row. The results obtained using these lamps are discussed below on
a crop by crop basis: .
Squash. Squash was selected as a subject species because its related
species, the 'Poinsett' cucumber, was one of the most sensitive crops in
Beltsville greenhouse and growth chamber experiments (4). Table 4 shows
the effects of UV enhancement on dry weight of tops, fresh weight of fruits,
number of fruits, and number of male and female flowers. Only the reductions
in fruit weights and number were significant (P = 0.2 to 0.3). The dry
weight of tops also decreased with each increase in UV-B irradiance, but
-------
Table 3. Temperature and Precipitation Measurements, 1977
Mean high
temperature
(ฐC)
May
June
July
August
September
October
November
25.4
26.9
31.7
30.7
27.2
18.7
12.8
Mean low
temperature
(ฐC)
11.8
14.8
19.5
19.6
16.3
7.3
5.3
Precipitation
(mm)
37.1
60.2
112.5*
42. -9
33.8*
133.9
104.9
* Includes supplemental irrigation
-------
Table 4. Means of squash parameters and standard error for the 0.5-m rows and 1.5-m rows and means and
ranges of weighted mWra~2 for the three levels of biologically effective UV above ambient UV.
Row
Control
0.5
1.5
Low
Dry
weight
of
Range Mean tops
(mWm~2 BUV) (mWrn~2 BUY) (g)
m .009- .518
m .004- .193
.165
.070
585.75
464.89
Fresh
weight
of
Standard fruits
Error (g)
567.82
433.02
Fruits
Standard
Error (No.)
55.4
110.3
18.0
11.0
Flowers
Standard
Error (No.)
1.4 15.
1.9 6.
62
75
Standard
Error
3.5
2.5
Flowers
Standard
(No.) Error
4.63
2.38
2.1
0.8
enhancement
0.5
1.5
m .898-1.450
m .491- .956
1.162
.665
514.63,
357.99
20.6 436.54
44.6 253.95
126.6
84.7
14.3
7.0
1.7 14.
1.1 6.
67
17
3.3
2.7
4.67
1.67
1.0
0.8
High
enchancement
0.5
1.5
0.5
1.5
m 1.669-2.498
m .790-1.553
m
m
2.009
1.166
P =
P -
485.16 .
.396.53
N.S.
N.S. .
37.6 418.59
20.0 258.65
0.3
N.S.
45.5
25.7
13.1
7.9
0.2
0.3
1.7 14.
0.9 6.
N,
N.
50
25
S.
S.
2.2
4.7
6.38
2.50
N.S.
N.S.
2.8
0.8
-------
not significantly. The results suggested that higher irradiance levels
may adversely affect plant growth, and continued investigation under
field conditions are warranted.
Bean. Garden beans were unaffected by UV-B enhancement. The crop
was not visibly damaged during the course of the experiment (Table 5).
Soybeans. Although weight of seed and plant height differed significantly
(P = 0.2 and 0.3, respectively) no consistent trends were observed (Table 6).
As with beans, the plants were not visibly injured but the results warrant
continued investigations.
Sugar beets. This was the most sensitive species tested in the field
experiments. All parameters measured except sucrose content at the lower
irradiance levels were significantly affected (Table 7). Each UV-B
increment reduced dry weight of tops and fresh weight of roots. In contrast,
sugar content increased with increased UV-B at the higher irradiance levels,
probably as a result of lower metabolic activity in leaves which provided .
higher carbohydrate concentrations for transport to the roots.
These data, however, suggest that even in field experiments small
increases of present ambient UV-B may injure this crop.
Sorghum. The higher irradiance levels reduced fresh weight of tops
(P = 0.3) (Table 8). Because the probability level was so low and because
\
plant height was not significantly affected, further research is needed
*
to confirm possible UV-B effects.
Sweet corn. The fresh weight of ears, the number of ears, and the
plant height were reduced (P = 0.2 to 0.3) (Table 9). As with the crops
other than sugar beets, the level of probability was low. However, since
three out of the four parameters measured were significant, continued
evaluation of UV-B effects under field conditions seems warranted.
-------
Table 5. Mean of bean parameters and standard error for the 0.5-m rows and 1.5-m
rows and means and ranges of weighted raKm~2 for the three different
levels of biologically effective UV above ambient UV.
Row
Dry
weight
of
Range Means tops
(mWnf2 BUY) (mWnT2 BUV) (g)
Fresh
weight
of
Standard Fruits Standard
Error (g) Error
Fruits
Standard
(No.) Error
Control
0.5 n
1.5 m
Low
enhancement
0,5 m
1.5 m
High
enhancement
0.5 m
1.5 m
.009- .518
.004- .193
.898-1,450
.491- .956
1.669-2.498
.790-1.533
.165
.070
1.162
.665
2.009
1.166
94.33
98.55
97.29
104.91
84.00
102.25
8.11
4.41
15.70
2.30
6.04
3.00
719.0
759.4
730.1
818.7
634.2
789.0
6.53
6.69
4.28
4.00
3.12
2.92
15.9
15.5
17.0
17.1
12.8
17.9
1.33
2.44
.61
' .40
2.11
.74
Differences were not significant at the 0.3 level according to the P test.
-------
Table 6. Means of soybean parameters and standard error for the 0.5-m rows and
the 1.5-m rows and means and ranges of weighted mWm for the three
different levels of biologically effective UV above ambient.
Row
Control
0.5 m
1.5 m
Low
enhancement
0.5 m
1.5 m
High
enhancement
0.5 m
1.5 m
0.5 m
1.5 m
Range
(raWm"^ BUV)
.009- .518
.004- .193
.898-1.450
.491- .956
1.669-2.498
.790-1.533
Mean
(mWnT2 BUV)
.165
.070
1.162
.665
2.009
1.166
P =
P =
Weight
of seed
(g)
275.88
366.88
267.73
289.30
277.15
303.63
N.S.
0.2
Standard
Error
9.40
36.52
13.2
13.3
15.5
12.7
Height
(cm)
476.98
620.65
470.73
648.57
541.43
518.08
0.3
N.S.
Standard
Error
22.2
53.6
41.5
77.2
6.1
41.1
10
-------
Table 7. Means of sugar beet parameters and standard error for the 0.5-m rows and
the 1.5-ra rows and means and ranges of weighted raWnT^ for the three
different levels of biologically effective UV above ambient.
Range
Row
BUY)
Dry Fresh
weight weight
of of
Means tops Standard roots
r2 BUV) (g) Error (g)
Root
Standard sucrose Standard
Error (%) Error
Control
0.5 m
1.5 m
Low
enhancement
0.5 m
1.5 ra
High
enhancement
0.5 m
1.5 ra
0.5 m
1.5 m
.009- .518
.004- .193
.898-1.450
.491- .956
1.669-2.498
.790-1.533
.165
.070
1.162
.665
2.009
1.166
P =
P = -
470.13
540.33
295.26
428.21
243.14
388.37
0.05
0.3
54.3
67.8
26.7
48.3
22.2
39.8
5705.78
5600.00
3720.27
3844.97
3059.40
3997.55
0.1
0.2
874.7
689.8
482.9
341.5
391.2
518.1
13.98
14.73
16.90
14.43
16.35
15.05
0.05
N.S.
0.5
0.2
0.5
0.0
0.5
0.4
11
-------
Table 8. Means of sorghum parameters and standard error for the 0.5-m rows and the
1.5-m rows and means and ranges of weighted mMn for the three different
levels of biologically effective UV above ambient.
Fresh
weight
of
Rows
Control
0.5 m
1.5 m
Low
enhancement
0.5 m
1.5 m
High
enhancement
0.5 m
1.5 m
0.5 m
1.5 m
Range
(mWm~^ BUY)
.009- .518
.004- .193
.898-1.450
.491- .956
1.669-2.498
.790-1.533
Mean
(mWnT2 BUV)
.165
.070
1.162
.665
2.009
1.166
P =
P =
tops
(8)
2526.68
2243.23
2162.23
2136.63
2144.05
2163.80
0.3
N.S.
Standard
Error
245.9
109.9
7.3
155.5
38.9
100.3
Height
(cm)
1130.15
852.53
1110.30
866.23
1191.40
838.40
N.S.
N.S.
Standard
Error
47.1
55.1
53.5
76.1
79.1
59.1
12
-------
Table 9. Means of sweet corn parameters and standard error for the 0.5-m rows and the 1.5-m rows and means and
ranges of weighted mWnf^ for the three different levels of biologically effective UV above ambient.
Row
Control
0.5 m
1.5 m
Low
enhancement
0.5 m
1.5 m
High
enhancement
0.5 m
1.5 m
0.5 ra
1.5 m
Range
(mWnr2 BUV)
.009- .
.004- .
.898-1.
.491- .
1.669-2.
.79 -1.
518
193
450
956
498
533
Means
(mWm~2 BUV)
.165
.070
1.162
.665
ป
2.009
1.166
P =
P =
Dry
weight
of
tops
(8)
140.06
185.00
*
117.19
196.46
116.63
142.76
N.S.
N.S.
Standard
Error
14.3
35.0
13.2
17.8
11.0
21.9
Fresh
weight
. of
ears
(g)
109.40
137.55
83.42
119.91
60.97
111.16
0.2
N.S.
Standard
Error
14.9
14.5
18.4
17.8
8.0
13.7
Height
(cm)
140.40
144.35
146.73
177.83
128.95
147.15
N.S.
0.3
Standard
Error
28.8
102.1
13.3
274.4
170.6
18.9
Ears
(No.)
4.75
5.75
3.33
5.00
2.75
4.38
0.2
0.3
Standard
Error
.7
.6
.8
.9
.4
.5
-------
For the second experiment, the FS-40 lamp assemblies provided
significantly higher total irradiances than those on the first experiment,
as well as different spectral characteristics. Irradiances levels averaged
3.8, 5.0, and 11.9 mWnT2 BUY for the 1.5-ra row and 3.9, 7.9, and 23.9 raWm"2
BUV for the 0.5-m row. These irradiances provided approximately 1.2,
1.6, and 3.9 times for the 1.5-m row and 1.3, 2.6, and 7.8 times for the
0.5-m row of the control Beltsville sunshine.
Squash. This species was quite resistant to these high irradiance .
levels. Dry weight of tops and fresh weight of fruit decreased significantly
only at the highest irradiances, and then only at P = 0.2 or 0.3 (Table 10).
Broccoli. Broccoli was much more sensitive to UV than squash, providing
P values of 0.05 and 0.01 at the higher irradiance levels (Table 11).
However, irradiance levels two to eight times the value of the Beltsville
control sunshine were required to provide this level of significance.
We conclude from these field experiments that plants are considerably
more resistant to injury from enhanced UV in a field environment, with its
higher visible light energy, than they are when grown in a greenhouse or
a growth chamber. Whether resistance increases enough to preclude UV
damage at projected UV-B increases, however, cannot be determined from
our results. ^
14
-------
Table 10. Means of squash parameters and standard error for the 0.5-ra rows and
1.5-m rows, means and ranges of weighted mWm for the three different
levels of biologically effective UV above ambient provided by unfiltered
FS-40 lamps.
Dry Fresh
weight weight
of of Fruits
Row
Control
0.5
1.5
Low
Range Means tops Standard fruits Standard
(mWm BUV) (mWm~2 BUV) (g) Error (g) Error (No.)
m
m
.106- 3.
.088- 3.
227
182
.805
.743
444.
381.
85
10
53.0
54.7
460.95
396.83
62.0 15.
64.0 15.
6
5
Standard
Error
1.8
2.8
enhancement
0.5
1.5
High
m
m
.636- 9.
.478- 3.
473
296
4.801
1.887
358.
384.
95
09
45.5
38.9
342.41
302.58
22.3 14.
69.2 11.
6
5
2.6
1.6
enhancement
0.5
1.5
0.5
1.5
m 14
m 4
m
m
.271-28.
.229-12.
736
257
20.840
8.797
P =
P =
337.
362.
0.
N.
27
64
3
S.
30.3
55.4
304.69
314.89
0.2
N.S.
35.9 14.
38.6 14.
N.
N.
2
1
S.
S.
1.3
1.6
15
-------
Table 11. Means of broccoli parameters and standard error for the 0.5-m row and
the 1.5-m rows, means and ranges of weighted mWm for the three
different levels of biologically effective UV above ambient provided
by unfiltered FS-40 lamps.
Row
Range
(mWm~2 BUY)
Dry
weight
of
Means tops Standard
(mWnT2 BUY) (g) Error
Fresh
weight
of Fruits
Fruits Standard Standard
(g) Error (No.) Error
Control
0.
1.
Low
5
5
m .106- 3.227
m .098- 3.182
.805
.784
844.48
977.25
79.4
111.9
794.
729.
82 49.1
16 155.4
3.20
2.50
.3
.3
enhancement
0.
1.
5
5
ra .636- 9.473
m .478- 3.296
4.801
1.887
617.33
849.15
45.3
61.2
605.
505.
90 43.9
28 59.6
3.75
2.50
.2
.4
High
enhancement
0.
1.
0.
1.
5
5
5
5
m 14.271-28.736
m 4.299-12.257
m
m
20.840
8.797
P =
P =*
602.70
781.16
0.05
0.3
31.7
21.7
438.
642.
0.
N.
96 53.4
34 51.2
01
S.
2.30
2.20
0.05
N.S.
.3
.3
16
-------
LITERATURE CITED
1. Ambler, J. E., D. T. Krizek, and P. Semeniuk. 1975. Influence of UV-B
radiation on the early seedling growth and translocation of Zn from
cotyledons in cotton. Physiol. Plant 34:177-181.
2. Krizek, D. T. 1975. Influence of ultraviolet radiation on germination
and early seedling growth. Physiol. Plant 34:182-186.
3. Krizek, D. T. 1978. Response of selected vegetables and agronomic crops
to increased UV-B irradiation under greenhouse and growth chamber conditions.
Final BACER Report submitted to the Environmental Protection Agency.
4. Krizek, D. T. 1978. Differential sensitivity of two cultivars of
Cucumis sativus L. to increased UV-B irradiance. Plant Physiol. Suppl.
61(4):92. (Abstr.).
5. Rowan, J. D. and K. H. Norris. 1978. Instrumentation for measuring
irradiance in the UV-B region. Final BACER Report submitted to the
Environmental Protection Agency.
6. Thimijan, R. W., H. R. Cams, and L. E. Campbell. 1978. Radiation
.sources and related environmental control for biological and climatic
effects UV research. Final BACER Report submitted to Environmental
Protection Agency.
17
-------
FINAL REPORT
EFFECTS OF UV-B RADIATION ON PLANT MEMBRANE PERMEABILITY,
RESPIRATION AND OXYGEN EVOLUTION
P. C. Jackson
Plant Stress Laboratory
Plant Physiology Institute
Beltsville Agricultural Research Center
Beltsville, Maryland 20705
EPA-IAG-D6-0168
Project Officer:
R. J. McCracken
Agricultural Research, Science and Education Administration
U. S. Department of Agriculture
Washington, D.C. 20250
Prepared for
Environment Protection Agency
BACER Program
Washington, D.C. 20460
-------
ABSTRACT
Poinsett cucumber plants were grown in a growth chamber under UV-A
_n
and under UV-A + 8-24 mWm~ UV-B (2.8-7.9 Sun equiv.) for 6 hours a day
midway in the light period. Leaves of plants under UV-A + UV-B were
smaller, weighed less, and took up ions, respired and evolved 02 at slower
rates than leaves of plants under UV-A alone. The pattern of appearance
of these effects varied among leaves and with duration of treatment. The
effects of UV-B were much the same whether treatment was started as soon
as the plants emerged or on the eighth day of plant growth. When rates
4*
of Na and Cl uptake, respiration and Oo evolution are expressed on the
basis of amounts per leaf instead of amounts per gram, the inhibitory
effects of UV-B are greater because of the lower weights of the leaves
under UV-B. There is no evidence of an increase in ion permeability in
these plants under conditions of this study. The data are more indicative
of a decrease in permeability insofar as the ion uptake rates were inhibited
and somewhat more K+ was retained by the plants under UV-B. Effects of
respiration, Oo evolution and ion uptake generally preceded appearance of
visual symptoms (chlorotic spots). Thus measurements of either ion uptake
or respiration could provide an early assay for plant sensitivity to UV-B.
-------
INTRODUCTION
Recent awareness that the use of chlorofluorocarbons in aerosol spray
cans and as refrigerants may decrease the protective layer of ozone in the
stratosphere sufficiently to cause ?n increase in ultraviolet radiation in
the region of 280-320 nm (UV-B) raises the question of the effect of increased
UV-B radiation on agricultural crops. Although effects of ultraviolet
radiation in the regions of 100-200 nm (UV-C) and 315-400 nm (UV-A) on plants
are well known, there is a dearth of information on effects of UV-B on plants.
The work presented here was undertaken to provide information on early
effects of UV-B radiation on plants and to assess whether measurements of
respiration, oxygen evolution and permeability to ions can serve as assays
for UV-B effects before symptoms of toxicity are visible.
MATERIALS AND METHODS
Seeds of cucumber (Cucumis sativus var. Poinsett) (a cultivar known to .be
sensitive to UV-B radiation) were germinated on moist paper towels at 30ฐC.
On the second day, each seedling was planted in a 10-cm pot of moistened
vermiculite and placed in a grovrth chamber in the presence or absence of UV-B
radiation (Fig. 1). UV-B radiation was from four Westinghouse FS-40 sunlamps
suspended 30 cm above the tops of the plants and covered either with 5-mil
\
Mylar film (UV-A control) or with cellulose acetate at a thickness of 5 mil
'Mention of a trademark, proprietary product, or vendor does not constitute
a guarantee or warranty of the product by the U.S. Department of Agriculture
and does not imply its approval to the exclusion of other products or
vendors that may also be suitable.
-------
(CA-5) or 10 mil (CA-10). The intensity of the UV-B irradiance as measured
by a Norris Spectroradiometer is shown in Table 1. The plants were maintained
on a 16-hour light period at 30ฐC with f> hours of UV-B exposure midway during
the light period, and an 8-hour dark period at 25ฐC. They were watered daily
with a 1/5 Johnson solution and the pots were rearranged daily to minimize
effects of position. Samples of whole leaves, leaf sections, or cotyledons
were taken immediately after UV-B treatment.
Oxygen uptake was measured in the dark and oxygen evolution was measured
in the light, in water at 25ฐC by means of oxygen electrodes. Permeability
was assayed by measuring fluxes of K , Na , Cl , Ca , and Mg . Samples were
-2 -A
maintained in aerated solutions of 10 M NaCl + 10 M CaSO, at pH 5.5 for
various periods from 1 to 6 hours. Then they were removed, rinsed twice with
demineralized water, blotted gently and weighed. The samples were then ashed
at 480ฐC for 1 hour. The ashed samples were dissolved in 1 N HNO- + 10%
CH-COOH and aliquots of these solutions were taken for analyses of ion content.
+ + 2+ 2+
Na and K were determined by flame photometry, Ca and Mg were determined
by atomic absorption and Cl was measured by potentiometric titration.
. . . RESULTS
Week-Old Cucumber Plants
\
First leaves of plants grown for 7 days before UV-B treatment was initiated
showed little or no difference from controls (under Mylar) in leaf size or
weight during the subsequent 2 weeks under UV-B (Table 2). On about the sixth
day, small chlorotic areas appeared on the edges of the second and third leaves
of the plants under CA-5 filters. Leaves of plants under CA-10 filters had few
or no chlorotic spots. Examination of these areas under a microscope revealed
-------
that they were not only devoid of chlorophyll but almost devoid of chloroplasts,
starch and other structures as well. No necrotic areas or oxidized phenols
/
were apparent in these leaves when they were examined after removal of
chlorophyll.
The second and third leaves of control plants emerged about the third day
.of UV-B treatment, but emergence and expansion of these leaves under UV-B was
delayed initially as shown by relatively low weights at 6 days in Table 2.
The weights caught up somewhat when the control leaves reached full develop-
ment however. Cotyledons tended to decrease in weight sooner under UV-B. The
UV-B irradiance had no significant effect for the first 10 days on K concen-
-2
trations of the first and second leaves whether they were in water or in 10
M NaCl. However, K concentrations were significantly higher in these leaves
at 14 days and at 6 days in the third leaves.
Rates of Na uptake by first leaves under UV-B generally were somewhat
stimulated initially (Table 3) but were not effected after 2 days. Although'
Na uptake by second leaves was inhibited at 6 days, soon after emergence, UV-B
had no effect thereafter and had no effect on Na uptake by third leaves. Both
Na"*~ and Cl~ uptake by cotyledons under CA-5 was inhibited from the first day
of treatment, but UV-B at the lower intensity had no consistent effect. Rates
of Cl~ uptake were more sensitive to UV-B exposure, especially in leaves under
s
CA-5, the higher UV-B intensity. Inhibition was observed as early as 6 days in
2+
second leaves and in all leaves by the fourteenth day. Concentrations of Ca
9+ '
and Mg were affected little by UV-B exposure (Table 4). First leaves had
2+2+
higher Ca and Mg concentrations at 14 days than control leaves and third
2+
leaves had lower Ca concentrations at 6 days. Rates of oxygen uptake and
evolution were also stimulated during the first days of UV-B treatment and
inhibited thereafter (Fig. 2). The inhibitory effects were generally greater
under CA-5.
4
-------
Two-Day-Old Cucumber Plants
Leaves of plants exposed to UV-B from the time of epicotyl emergence
(2 days) generally respired and evolved' 0 at somewhat inhibited rates during
the first few days after leaf emergence, but thereafter, the rates were about
.the same as or slightly higher than those of control leaves on a nmole/min-g
fresh wt. basis (Fig. 3). Leaves of these UV-B-treated plants, however,
weighed less than control leaves (Table 5) and were 30-40% smaller, so that
respiration and 0 evolution on a nmole/plant basis were inhibited. Leaves of
plants under UV-B for 8-10 days also took up Na+ and Cl~ from 10~2 M NaCl at
slower rates than control leaves (Table 6). Rates of uptake by first and
second leaves were less inhibited by the fifteenth day, suggesting some degree
of photorepair. Ion uptake rates by cotyledons were inhibited from the first
and showed no evidence of photorepair. Since weights of leaves in these experi-
ments were less also under UV-B, particularly at the higher UV-B intensity
(CA-5), Na and Cl~ uptake rates were likewise inhibited on a micro-equiv/
plant basis. There was no consistent effect of UV-B on K concentrations
(Table 5). The concentrations in the cotyledons and leaves under UV-B were
sometimes higher than those in control leaves; consequently, leakiness in the
94- 2+
leaves under UV-B treatment was not evident. Concentrations of Ca^" and Mg
in leaves under UV-B were generally lower than concentrations in control
Ot o I
leaves. This probably resulted from inhibi'tion of Ca and Mg~ uptake from
the nutrient solution used to water the plants and not from leakiness. The
74- 2+
Ca and Mg concentrations of control leaves increased with age, but concen-
trations of these ions in UV-B-treated leaves increased very little until about
the fifteenth day.
-------
CONCLUSION
In general, leaves of Poinsett cucumber plants exposed to UV-B radiation
in these experiments were smaller, weighed less, and took up ions, respired,
and evolved oxygen at slower rates than leaves of control plants under UV-A.
The magnitude of the effects reflected the intensity of the UV-B radiation,
but did not reflect the age of the plants. Effects of UV-B were much the same
whether treatment began on the second day or on the eighth day of plant growth.
Weights of the leaves growing under UV-B were lower in both groups of plants,
so that expression of the rates of ion uptake, respiration and 0- evolution
and ion contents on the basis of equivalents per plant instead of equivalents
per gram would greatly intensify the inhibitory effects. Some degree of photo-
repair was suggested insofar as the effects were sometimes less as time
proceeded. .
There is no evidence of an increase in ion permeability in these studies.
The data are more indicative of a permeability decrease since ion uptake rates
were inhibited and K was not lost. Effects on ion uptake, respiration and Oo
evolution generally preceded the appearance of visual symptoms. The onset of
inhibition of Na and Cl uptake occurred about the same time, regardless of
the leaf development, which suggests that the UV-B affected the whole plant.
Generally ion uptake was affected at about the same time as respiration and 0^
evolution. This and similarities in the patterns of the effects suggests that
ion uptake rates reflect respiratory rates. Thus measurements of either
respiration or ion uptake could provide an early assay for plant sensitivity
to UV-B radiation.
-------
Table 1. UV-B Irradiances in the Growth Chamber.
1 t
'Filter '
Mylar
CA 10 Mil.
CA 5 Mil.
Biologically Effective UV '
i
(mWm~2 ) i
0.265
10.954 - 8.604
24.131 -16.733
UV-B Sun
0.09
3.58
7.89
Equivalents
- 2.81
- 5.47
Plants were maintained at 30 cm from the UV source. Pots were
rearranged 5x/wk. The higher figures are the irradiance when the
filters were new and the lower figures are the irradiance a week
later, when the filters were replaced.
-------
Table 2. Effects of UV-B radiation on K concentration and weights of
cucumber leaves
1 I
r i
.'UV-B '
'Treatment '
Days
Cotyledons
1
2
6
Leaf 1
1
2
6
10
14
Leaf 2
6
10
14
Leaf 3
6
10
.14
t
+ ,
K Concentration
Mylar
neq/g
50
56
61
91
91
73
62
57
85
66
63
62
81
73
CA-10
% My
108
107
115
112
103
97
100
117**
92
106 .
119**
138**
96 %
107
CA-5 '
lar
103
112
116
108
97
103
108
112**
93
115
111**
152**
100
113
Mylar
8
0.55
0.60
0.63
0.39
0.62
1.39
1.40
1.35
2.35
2.73
3.24
1.18
3.70
4.46
t
Weight '
CA-10
%
91
108
72
79
102
90
93
120
72*
91
102
103
87
101
CA-5 '
Mylar
108
92
77
91
93
81
101
119
63*
91
88
49*
64
83
Plants were 7 days old when UV-B irradiance was started. The * and **
indicate significant differences from controls (Mylar) at the 5% and 1%
levels, respectively.
-------
Table 3. Effect of UV-B radiation on rates of Na and Cl uptake by
_o
cucumber leaves from solutions of.10 "M NaCl, pH 5.5-
1 t
'UV-B
'Treatment
Days
Cotyledons
1
2
6
Leaf 1
1
2
6
10
14
Leaf 2
6
10
14
Leaf 3
6
10
14
Na Uptake Rates
Mylar
neq/min-g
31
18
35
30
42
36
41
44
45
61
54
51
51
53
CA-10
%
115
55
111
103
117
116
127
97
.78
93
82
99
86
95
CA-5
Mylar
78
53
65
121
136
108
88
82
76
82
88
95
* 101
93
t i
' Cl~ Uptake Rates '
1 Mylar
neq/min-g
19
9
22
18
27
20
19
24
28
21
32
18
23
' '* 25
CA-10
%
102
100
104
77
103
119
107
67
86
95
63
112
85
73
CA-5 '
Mylar
68
72
79
122
99
122
91
71
72
77
61
115
63
59
The plants were 7 days old when UV-B irradiance was started.
-------
2+ 2+
Table 4. Effect of UV-B radiation on Ca and Mg concentration of
cucumber leaves.
1
'UV-B
'Treatment
Days
Cotyledons
1
2
6
Leaf 1
1
2
6
10
14
Leaf 2
6
10
14
Leaf 3
6
10
14
i
2+
' Ca Concentration
' Mylar
neq/g
103
110
190
. 77
83
151
165
140
59
170
187
47
75
141
CA-10
% Mylar
130
85
137
90
88
88
120
160**
81
90
120
45**
110
110
CA-5
113
138
133
75
88
95
134
184**
81
87
105
35**
90
88
i
2+
MS
1 Mylar
neq/g
85
85
133
36
45
84
110
84
56
105
98
47
73
83
i
Concentration '
CA-10
% Mylar
115
86
114
102
106
89
114
126
82
91
105
92
97
106
CA-5 '
100
118
121
86
93
96
98
144*
85
85
104
89
90
88
Plants were 7 days old when UV-B irradiance was started. The * and **
indicate significant differences from controls (Mylar) at the 5% and 1%
levels, respectively.
-------
.OUUUIL-.
Table 5. Effects of UV-B radiation on K+ concentration and weights of
cucumber leaves
1
UV-B '
Treatment '
Days
Cotyledons
1
2
3
4
5
8
10
Leaf 1
5
8
10
12
15
Leaf 2
8
10
12
15
Leaf 3
10
12
15
i
K"*" Concentration '
Mylar
neq/g
61
69
61
76
62
60
51
88
72
71
66
52
91
70
88
51
99
109
57
CA-10
%
137**
102
102
81
98
119
121**
103
141**
117**
83
99
127*
124
75
111
117
75
95
CA-5 '
Mylar
105
100
88
81
108
117
149*
116 .
132*
123**
79
114
106
113
73
121*
123
81
112
Mylar
g
0.196
0.236
0.412
0.458
0.515
0.565
0.492
0.385
0.710
0.812
0.942
0.975
0.762
1.690
2.G65
2.170
0.685
1.137
2.848
i
Weight '
CA-10
%
93
116
61*
98
98
97
115
68
79**
94
89
97
68**
78*
62*
93
83
88
71*
CA-5 '
Mylar
98
110
62*
93
83**
79**
95
53*
63**
80**
72**
72**
48**
62**
51**
49**
51**
52*
59**
Plants were 2 days old when UV-B irradiance was started. The * and **
indicate significant differences from controls (Mylar) at the %5 and 1%
level, respectively.
-------
Table 6. Effect of UV-B radiation on rates of Na+ and Cl~ uptake from
10~2 M NaCl (pH 5.5) by leaves of cucumber seedlings^
1
*UV-3
'Treatment
Days
Cotyledons
1
2
3
4
5
8
10
Leaf 1
5
8
10
12
15
Leaf 2
8
10
12
15
Leaf 3
10
12
15
I
' Na+ Uptake
' Mylar
neq/min-g
49
40
35
33
24
28
18
36
47
50
24
38
50
57
75
103
32
118
132
CA-10
% M
90
66
91
83
100
76
96
132
84
82
88
84
73
89
74
94
108
52
95
Rates
CA-5
ylar
75
58
67
66
84
82
77
104
77
75
77
' -
69
%S7
68
66
80
37
72
t
1 Cl~ Uptake Rates
' Mylar
neq/min-g
9.2
11.7
9.2
9.6
12.5
14 .2
12.3
16
20
18
17
20
17
26
24.
36
13
20
52
i
i
CA-10 CA-5 '
7, Mylar
100
-
86
78
73
82
81
91
105
113
110
81
64
84
88
91
98
88
57
100
79
77
72
97
50
74
109
103
84
64
94
62
78
91
91
72
90
50
4
Plants were 2 days old when UV-B irradiance was started..
-------
2+ 2+
Table 7. Effects of UV-B radiation on Ca and Mg concentrations of
cucumber leaves.
1 1
'UV-B '
'Treatment '
Days
Cotyledons
1
2
3
4
5
8
10
Leaf 1
5
8
10
12
15
Leaf 2
8
10
12
15
20
Leaf 3
10
12
15
20
2+
Ca
Mylar
neq/g
65
135
106
175
123
168
228
203
123
172
260
250
68
122
296
197
316
81
127
169
231
Concentration
CA-10
% Mylar
101
91
94
92
98
107
91
103
87
97
79
81
90 *
73**
71
81
106
CA-5
93
88
88
80*
94
95
95
99
84
90
61
83
72*
78**
51*
92
90
98 106
84
91
114
80
97
90
M,2+
' Mylar
neq/g
76
89
92
104
109
131
164
55
71
97
127
123
51
. 67
87
134
115
52
58
88
75
Concentration
CA-10
% Mylar
96
93
91
100
103
103
94
84
107
99
92
100
73**
80*
96
t
89:
107
93
90
89
111
i
CA-5 '
93
95
92
90
112
100
87
71**
97
93
49*
77
77*
78**
77
86
82
85
96
100
83
Plants were 2 days old when UV-B irradiance was started. The * and **
indicate significant differences from controls (Mylar) at the 5% and 1%
levels, respectively.
-------
FIGURE LEGENDS
Fig. 1. Diagram of the growth chamber showing levels of visible light.
Positions of the UV lamps are indicated by dashed lines. Sheets
of methylmethacrylate, shown by solid lines, divided the chamber
into 3 sections for the UV-B treatments. The treatments are
indicated at the top of the diagram by descriptions of the filters
which covered the UV lamps. Initially, there were about 20 pots
of plants located in the center of each section.
.Fig. 2. Effects of UV-B radiation on rates of Qฃ uptake and evolution by
leaves of cucumber plants. The rates are expressed as percentages
of the control rates (nmoles/min-g fresh wt.). Solid lines are
for treatment under CA-10 filtered UV-B and dashed lines for CA-5
filtered UV-B. The plants were 7 days old "when UV-B irradiance
began.
Fig. 3. Effects of UV-B radiation on rates of (^ uptake (Fig. 3a) and
evolution (Fig. 3b) by leaves of cucumber plants. The rates are
expressed as percen^<ฐes of the control rates (nmoles/min-g fresh
wt.). The plants were 2 days old when UV-B irradiance began.
-------
o 9
e
5 mil CA 10 mil CA
My lar
1
BfflSESC
'
'
300
330
305
i
.WBWWJW
1
,'.
"jgawgggga
310
330
320
i
SWMKBgBrfaaiftaM^MltigJgSAiytMl'
!w;ygซfa
1
'aiK*M!WWป
j
310
i
*
.
.330
310
laiHMiHHBHiimi.uaปimjja^wj
'
;.ซ9MBปJM4
to
H
-------
O2 UPTAKE
O2 EVOLUTION
80
inn
IUU
80
t-
ce.
6
Gฃ
z
O
u
N 80
120
inn
IUU
80
60
THIRD LEAF
P II
1 1 1
i
i
ilt i .1 '
: : I i
! SECOND LEAF [ j
I t ' '1
i i
t P i! ' J
111 if,.
If !
if i
FIRST LEAF
i
1 :
1. ป !
i i
' ! !
i
COTYLEDONS
1234 789 1
17 1234 789
17
DAYS OF UV-B TREATMENT
Fig. 2
-------
i-fluvc 3
0
tft 5-
80
I
JO
r/
Dous
10
UV-
17
-------
v-
. ce
A
-J
loo
loo
7 (o
O C-volurCeo
Utxf
'7
II
*
1
i i rsc uu
0 l ( l
LoKi led>n>.
ir
II
ฃ
7 JO /<-/ //
I >^ปcป, ** 4-
-------
FINAL REPORT
PHYSIOLOGICAL AND BIOCHEMICAL EFFECTS ON UV RADIATION:
CHANGES IN ANTHOCYAN'IN PIGMENTATION IN COLEUS BLUMEI BENTH,
A. L. Fleming
Plant Stress Laboratory
Plant Physiology Institute
Beltsville Agricultural Research Center
Beltsville, Maryland 20705
EPA-IAG-D6-0168
Project Officer:
R. J. McCracken
Agricultural Research, Science and Education Administration
U.S. Department of Agriculture
Washington, D.C. 20250
Prepared for
Enviromental Protection Agency
BACER Program
Washington, D.C. 20460
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ABSTRACT
Rooted cuttings of Coleus blumei Benth. were exposed to enhanced UV
radiation [50 to 800% increase in biologically effective UV (BUV) radiation]
for 1 to 48 hours. The UV radiation was provided by Westinghouse FS-40
fluorescent sunlamps, unfiltered (UV-A, B, C) or filtered with 5, 10 or
20 mil cellulose acetate (UV-A, B) or 5 mil Mylar (UV-A). Exposure to high
levels of unfiltered UV radiation (700 to 800% increase in BUV) resulted
in a significant decrease in the concentration of anthocyanin extracted
from the leaves with methanol and HC1. Degradation of the pigment occur-
red after 12 hours of exposure and was intensified with an increase in
duration of exposure up to 24 hours. Exposure to a 100% increase in BUV
under 5 mil cellulose acetate caused glazing of the leaf surfaces, distortion
of the leaf margins, and inhibition of leaf expansion. At this UV level, a
significant decrease in the anthocyanin content of leaves occurred after
36 hours of exposure (6 days at 6 hours/day).
A leaf injury index was developed to provide a visual evaluation of
the extent of UV injury. This index was useful in rating the severity of
plant responses to UV treatments at increases in BUV of less than 100%.
Spectrophotometric analysis of methanol-extractable materials in coleus
leaves indicated that the components absorbing at 280, 330, 412-434, and
525 nm decreased with UV treatment, whereas those absorbing at 415-425 and
650-660 nm increased.
-------
Recent studies (1, 7, 8, 9, 12, 13) have demonstrated that exposure
to UV radiation can alter levels of anthocyanin and associated plant
pigments. Wellmann (12, 13) showed that UV irradiation induced flavonoid
synthesis and phenylalanine ammonia-lyase (PAL) activity in parsley seedlings.
Ambler et al. (1) induced red pigmentation in cotton seedlings by exposing
them to UV-B radiation. Semeniuk (8) reported that the leaves of 'Supreme
Annette' poinsettia formed a purple red anthocyanin when exposed to a 100
percent or greater increase in biologically effective UV (BUV) radiation.
In contrast, high levels of UV-B radiation significantly decreased the
anthocyanin content of 15 Coleus cultivars (8). Time-lapse photography
of Coleus blumei Benth. plants (unpublished data, this laboratory) revealed
that the anthocyanin pigment broke down within 24 hours of exposure
to broad-band UV radiation from unfiltered Westinghouse FS-AO sunlamps.
The extent of the color change was a function of the physiological age of
the leaves and the total UV irradiance. In addition, no degradation of the
pigment was noted in portions of leaves that were shaded by the plant canopy.
In subsequent experiments, the UV radiation of FS-40 lamps was filtered
with cellulose acetate (CA) to approximate the natural UV spectrum (11). The
spectral cutoff and total UV irradiance were controlled by varying the filter
thickness. ' . ' >
*
The purpose of this study was to describe the time course of the
anthocyanin pigment changes in coleus and to obtain an indication of the
metabolic effects of broad band UV radiation.
"Mention of a trademark, proprietary product, or vendor does not constitute
a guarantee or warranty of the product by the USDA and does not imply its
approval to the exclusion of other products or vendors that may also be
suitable.
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MATERIALS AND METHODS
UV-B enhancement facilities were developed in cooperation with the
Agricultural Equipment Laboratory (AEL), Beltsville Agricultural Research
Center (BARC). Enhancement studies were conducted in accordance with
the guidelines established for the BACER program (2, 3). UV radiation
was provided by one or more Westinghouse FS-40 fluorescent sunlamps
unfiltered or filtered with 6-hour-aged 5 mil Mylar (UV-A) or 5, 10, or
20 mil cellulose acetate (UV-A, B).
UV-B irradiance levels were determined with either an Optronic
Laboratories, Inc. Model 725 UV-B Radiometer or an Instrumentation
Research Laboratory (IRL) UV-B Radiometer (4, 6). Radiometer readings were
verified by spectral irradiance determinations (250-369 nm) with an auto-
mated spectroradiometer (4, 6) at selected locations in the experimental
irradiation areas.
_2
Weighted.irradiance levels are reported as mVm BUV, the biologically
effective UV derived from the AT9 weighting function, and unweighted
-2
irradiance as mWm obtained by summing the measured or calculated values at
2 2
each nanometer from 280-320 nm. Dividing mWm BUV by 3.06 (the mWm BUV
of control sunshine) provides the fraction of BUV received by each plant
relative to that of one control sunshine.
When UV irradiation was obtained by filtering the FS-40 lamps through
cellulose acetate, BUV was limited to the UV-B region (280-320 nm).
For details concerning average control sunshine, spectral character-
istics of UV fluorescent lamps and filters, and the weighting function, see
the BACER final reports of the AEL and IRL, BARC (6, 11).
The first experiment (Expt. 1) was conducted with unfiltered UV
radiation to determine the change of anthocyanin in the leaves of coleus
-------
with time. The subsequent experiments (Expts. 2 and 3) were conducted with
filtered UV radiation to determine the effect of UV radiation on the spectra
of methanol-extractable substances in coleus leaves, and to assess the
reliability of the anthocyanin content as an indicator of UV damage.
EXPERIMENT 1: EFFECT OF UV EXPOSURE ON ANTHOCYANIN PIGMENT OF COLEUS LEAVES
Six-week-old coleus cuttings were exposed to broad-band UV radiation
in a laboratory maintained at 26 + 2ฐC. The light sources were one unfiltered
FS-40 sunlamp and three 40 watt cool-white fluorescent lamps, positioned
-2 1
20 cm above the plants to provide approximately 150-200 yEm s of
photosynthetically active radiation (PAR). Treatments consisted of
continuous exposures of 1, 2, 4, 8, 12, and 24 hours. A UV control was run
concurrently under a section of the same lamp bank fitted with a plexiglass
filter; this filter has a cutoff at approximately 340 nm. Plants (54 cm
in height) were trimmed to two pairs of leaves at the 4th and 5th nodes.
The stem was excised above the 5th node. Each pair of leaves was analyzed
as a separate sample.
Following UV exposure fresh weights were determined and the leaves were
frozen at dry ice temperatures. After 15 minutes, the samples were thawed
.and cut into strips in preparation for extraction (5). Following extraction
\
with 100 ml of a solution of methanol containing HC1 (99:1), the anthocyanin
*
content was measured at 525 nm with a Gilford spectrophotometer. Pigment
concentration was reported as absorbance/gin fresh weight.
To determine the effect of shading, several leaves of a separate set of
plants were partially shielded by cardboard masks. The change in anthocyanin
was observed and recorded photographically.
Because of the large variation in the initial concentration of anthocyanin
the experiment was repeated with changes in the sampling and analytical methods,
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Instead of using entire leaves as analytical samples, leaf disks (11 mm
diameter) were taken from each leaf half. Samples were taken at 0, 12, or 24
hours. These treatments were randomized within each leaf pair and replicated
three times.
Three leaf disks from each leaf were placed in the barrel of a plastic
syringe and extracted for 1 hour with 99:1 methanol-HCl. The disks were
rinsed twice at 20-minute intervals with 5 ml portions of the extracting sol-
ution. The concentration of anthocyanin was determined spectrophotometrically
at 525 nm. The results were reported as absorbance/3 leaf disks.
EXPERIMENT 2: EVALUATION OF UV DOSE - RESPONSE FOR COLEUS
In an effort to further define the UV dose-response relationships in
coleus, 4-week-old coleus cuttings were exposed to broad-band UV radiation
produced by two FS-40 lamps, with appropriate plastic filters. The filters,
5 mil Mylar, 5 mil CA, 10 mil CA, and 20 mil CA were used. The spectral
cutoff characteristics of these filters are illustrated in Fig. 7.
Plants were grown on a 16 hour photoperiod at ca 28/25ฐ day/night
temperature. PAR was provided by four 1500 ma cool white fluorescent
lamps. Plants were exposed to UV irradiation during the last 6 hours of
the photoperiod for 3, 6, and 8 days at UV irradiances and BUV levels shown
*
in Table 1. UV measurements were taken from 250-369 nm with an Optronic
Laboratories Model 725 UV-B spectroradiometer (4,6).
After 3, 6, and 8 days of treatment, the second set of leaves below the
apex was extracted with methanol-HCl (99:1). The anthocyanin concentration
in each leaf was measured spectrophotometrically at 525 nm and reported as
absorbance/gm fresh weight. Plants were scored for visual leaf injury based
on leaf color, shape and size.
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EXPERIMENT 3. DETERMINATION OF SPECTRAL CHANGES OF METHANOL-EXTRACTABLE
SUBSTANCES IN COLEUS LEAVES.
Following exposure to unfiltered and filtered UV radiation, coleus leaves
were extracted with methanol-HCl (99:1). The resulting solution was concen-
trated by flash evaporation to less than 5 ml and diluted to 10 ml with the
extracting solution. UV and visible absorption spectra were determined
with a Perkin-Elmer UV-Vis spectrophotoraeter.
In order to classify the methanol-extractable constituents of coleus
leaves, other samples were extracted with methanol, concentrated by flash
evaporation, and chromatographed on paper (5, 10). Following development with
butanol:acetic acid:water (BAW, 4:1:5, top layer) the chromatographs were
viewed under a UV lamp. Fluorescent spots were eluted with methanol-HCl (99:1)
and diluted to 2 mil. The UV and visible spectra were determined as indicated
above.
RESULTS AND DISCUSSION-
EXPERIMENT 1: EFFECT OF UV EXPOSURE ON ANTHOCYANIN PIGMENT OF COLEUS LEAVES
The exposure of coleus to unfiltered UV radiation resulted in epidermal
damage, glazing and reduction in the anthocyanin pigment content. This damage
was primarly confined to the upper leaf surfaces and initially involved only
those tissues exposed to the radiation. The effect of shading is shown in Fig. 1.
The extractable anthocyanin of coleus leaves exposed to continuous UV radiation
(1 FS-40 + 3 cool white fluorescent lamps at 16 cm) for 1-24 hours decreased with
increasing time of UV exposure (Table 2). The trend in pigment reduction was
most striking between 16 and 24 hours. However, due to considerable variability
in initial pigment intensity, the differences were not statistically significant
at the 5% level by the F test. When improved sampling techniques were used,
-------
the anthocyanin concentration was found to be significantly decreased
by exposure to unfiltered UV radiation for 12 hours (Table 3).
EXPERIMENT 2: EVALUATION OF UV DOSE - RESPONSE STUDIES
The leaf injury in coleus, produced by exposure to UV radiation, can be
easily observed (7,8). In previous experiments with unfiltered UV radiation
(a 700-900% increase in BUV) exposure for 12 hours produced a rapid disap-
pearance of the anthocyanin pigment without concomitant changes in leaf
shape. However, exposure to FS-AO lamps filtered with 5 mil CA (42 hours
under a 300-500% increase in BUV) caused distortion of leaf margins (Figs.
2, 3). The expanding apical leaves assumed a "sickle shape," resulting
from decreased development of the.leaf half nearest to the UV source.
Although this symptom did not preclude further leaf expansion, the curva-
ture remained until senescence, even with a subsequent reduction in UV
exposure. Both symptoms were used in developing a leaf injury index (Table
4). Using these parameters, injury produced by the UV treatments over an 8-
day period ranged from slight injury, with the 10 mil CA filter, to intense
injury with the 5 mil CA filter. The range of UV injury from filtered
treatments is shown in Fig. 4.
The spectral transmission curves (Fig. 7) indicated that 5 mil Mylar
had an effective cutoff at approximately 310^-315 nm; the spectral cutoff
for the CA filters ranged from approximately 286-292, depending upon the
thickness of the filter.
The contrast in UV injury between the plants grown under Mylar (Fig. 5)
and those grown under 5 mil CA (Fig. 6) was striking. Symptoms of UV injury
under 5 mil CA included marginal distortion, glazing of the surfaces and
pigment changes in the apical leaves; under 5 mil Mylar, no visible changes
were observed.
-------
Statistical analysis (Table 5) indicated that the anthocyanin content
of plants irradiated under 5 mil CA decreased significantly with increases
in the UV dose.
EXPERIMENT 3: DETERMINATION OF SPECTRAL CHANGES OF METHANOL-EXTRACTABLE
SUBSTANCES IN COLEUS LEAVES
The visible- spectra of methanol-extractable constituents of coleus
leaves were modified by exposure to UV radiation. The extracts from unirradiated
control plants and those filtered with Mylar, produced strong absorbance peaks in
the 412-434, 520-535, and the 640-660 nm regions. UV treatment was characterized
by a reduction in the absorbance at the 520-535 peak and increases at 412-434
and 640-660 nm. The contrast between the unirradiated control plants and those
irradiated for 24 hours under unfiltered UV is shown in Fig. 7.
Similar results occurred for a 6-hour unfiltered treatment when the
plants were more highly pigmented. The visible spectrum for greenhouse
grown coleus is shown in Fig. 8. Strong absorbance bands occurred at 420
and 525 nm, with a minor peak at 657 nm. The spectrum for the 6 hour
unfiltered UV treatment was characterized by reduced absorbance at 525 nm
and substantial increases at 425 and 657 nm (Fig. 9). This pattern can
be used to demonstrate the effect of filtered treatments over time, shown
in Experiment 3. Exposures of plants to 3, 6, and 8 days of UV under
FS-40 lamps + 5 mil CA were associated with progressive increases in the
absorbance at 425 and 657 nm and relative decreases at 525 nm (Fig. 10).
The UV spectrum of the extracts from control plants shows absorbance
peaks at 275-295 and 320-340 nm. Twenty-four hour exposure to unfiltered
UV radiation reduced the absorbance in both regions (Fig. 11). The effect
of UV radiation can be visualized by calculation of absorbance ratios.
Table 6 contains these data for a 6-hour unfiltered treatment and the
corresponding unirradiated control.
7
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Paper chromatography and subsequent spectroscopy were used to determine
the methanol-extractable components of coleus leaves responsible for the
observed spectral characteristics. A summary of the spectral characteristics
of these bands is given in Table 7.
Band 7 produced a large, but poorly defined peak in the region of a 210-
230 run and sharp'maxima at 415 and 657 run. UV treatments increased the absor-
bance at 415 and 657 nm (Figs. 12 and 13). Band 6 contributed a small peak
at 290 nm and a large peak at 330 nm, which decreased with intensity of UV
treatment (Figs. 14 and 15). Band 4 (Figs. 16 and 17) absorbed at 270 and 330
and appeared as a large UV-absorbing area on the chromatograph. The spectrum of
band 2 showed strong peaks at 280 and 520-535 (Figs. 18 and 19). UV treatment
was particularly effective in eliminating this band.
With the exception of band 2, which appeared to be a derivative of cyanidin
(11), identification of the compounds involved in this separation has not been made,
The destructive effect of UV radiation on coleus leaf constituents, which
absorb in this region was expected. However phenols, such as cinnamic or ferulic
acids, did not increase measurably due to UV treatment. .
SUMMARY
Short-term exposure (12 to 16 hours) of 3-week-old coleus plants to high
\
levels of UV radiation (an increase in BUV of 100% or more) produced by
unfiltered FS-40 fluorescent sunlamps resulted in a significant decrease in
the concentration of anthocyanin that could be extracted from the leaves.
Although the concentration of the pigment in unexposed plants was variable
and depended on other environmental conditions, such as temperature, light
intensity, and nutrition of the plants, the specific decrease due to UV
treatment was observed by evaluation of the changes which occur in portions
-------
of the same leaf and in the opposite leaves of the same plant. Significant
degradation of the anthocyanin pigment occurred after 12 hours of exposure
and was intensified with an increase in exposure up to 24 hours. Complete
degradation of the pigment occurred within 36 to 48 hours, depending on
the experimental material. The quantitative relationship between the
concentration of the pigment and the UV dose, observed at a high level of
exposure (700 to 900% increase in BUV), did not apply proportionally to a
lower level of UV radiation (less than a 100% increase in BUV) produced with
CA filters. Exposure to a 100 percent increase in BUV caused distortion of
the leaf margins, and an inhibition of leaf expansion, as well as some
degradation of anthocyanin.
The visual leaf injury index appears to be a more useful indicator of
injury than does the measured anthocyanin content of coleus leaves at
levels of exposure constituting less than a 100 percent increase in BUV.
However, the anthocyanin content determined after a 24-hour UV exposure
to a 300 to 500 percent increase in BUV was more reliable as an indicator
of UV dosage than was the index. The effect of UV radiation on the
methanol-extractable constituents of coleus leaves was particularly dramatic.
The UV-absorbing components of the leaves were substantially decreased by
UV exposure. In the visible range, the strong absorbance at 525 (anthocyanin)
was decreased by maximuraal UV exposure, whereas the peaks at 410-420 and
657 were increased. Semeniuk (7) reported large increases in absorption
of extracts from coleus leaves examined at 268, 286, and 328 nm. This
relationship was not resolved by this study. The reason for increased
absorbance at 410-420 and 657 nm following UV exposure was not determined.
-------
Although the underlying mechanisms of UV injury in coleus remain to
be explained, the plant responses to enhanced UV radiation observed
(glazing of leaf surfaces, deformation of expanding leaves and specific
decreases in anthocyanin pigmentation) may serve as useful predictors of
UV damage in higher plant.
10
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LITERATURE CITED
1. Ambler, J. E., D. T. Krizek and P. Semeniuk. 1975. Influence of UV-B
radiation on the early seedling growth and translocation of Zn from
cotyledons in cotton. Physiol. Plant 34:177-181.
2. Cams, H. R., R. Thimijan, and J. M. Clark. 1977. Outline of irradiance
distribution of UV fluorescent lamps and combinations. Symposium on
Ultraviolet Radiation Measurements for Environmental Protection and Public
Safety. June 8-9, 1977. Nat'l Bureau of Standards Gaithersburg, MD (Abstract)
3. Krizek, D. T. 1977. Current UV measurement methodology and future needs
in photobiological research. Symposium on Ultraviolet Radiation Measurements
for Environmental Protection and Public Safety. June 8-9, 1977.
Nat'l Bureau of Standards Gaithersburg, MD (Abstract).
4. Norris, K. H. 1977. Development of a portable, automated UV-B spectroradio-
meter. Symposium on Ultraviolet Radiation Measurements for Environmental
Protection and Public Safety. June 8-9, 1977. Nat'l Bureau of
Standards, Gaithersburg, MD (Abstract).
5. Robinson, Trevor, 1975. Flavonoids and Related Compounds. Chap. 9, p. 190-
223. In_ The Organic Constituents of Higher Plants. Third Ed. p. 347.
Cordus Press, New Amherst, Mass.
6. Rowan, J. D. and K. H. Norris. 1978. Instrumentation for measuring
irradiance in the UV-B region. Final BACER Report submitted to the
Environmental Protection Agency.
7. Semeniuk, P. 1977. Response of Coleus blumei Benth. to enhanced UV-B
radiation. HortScience 12 (4) Section 2:395. Abstract.
8. Semeniuk, P. 1978. Biological effects of ultraviolet radiation on plant
growth and development in florist and nursery crops. Final BACER
Report submitted to the Environmental Protection Agency.
11
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9. Skelly, J. M., M. F. George, H. E. Heggestad, and D. T. Krizek. 1978.
Air pollution and radiation stresses. Chapter 2.5. I_n ASAE Monograph.
Modification of the Aerial Environment of Plants.
10. Swain, T. 1976. Flavonoids. p. 166-205. In. T. W. Goodwin, (ed.), Chemistry
and Biochemistry of Plant Pigments. Vol 2, Academic Press, NY.
11. Thimijan, R. W., H. R. Cams, and L. E. Campbell. 1978. Radiation sources
and related environmental control for biological and climatic effects
UV research. Final BACER Report submitted to Environmental Protection
Agency. .
12. Wellmann, E. 1974. Regulation of flavonoid synthesis by ultraviolet light
and phytochrome in cell cultures of parsley. Ber. Deutsh. Bot. Ges. 87:267-273.
13. Wellmann, E. 1975. UV dose-dependent induction of enzymes related to
; flavonoid biosynthesis in cell suspension cultures of parsley. FEES
Letters. 51:105-107. '
12
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Table 1. Weighted and unweighted UV measurements of UV irradiance
under FS-40 fluorescent sunlaraps covered with various
filters.
Filter
5 mil Mylar
20 mil CA
10 mil CA
5 mil CA
UV-B
Irradiance
mWm
129.7
982.0
1494.5
2161.4
BUV2
,^,-2
mWm
0.23
4.52
9.44
18.15
"Tieasurements were taken with a spectroradiometer 12.7 cm above
the canopy
2
A base line level of biologically effective UV (BUV) radiation was
equivalent to 3.06 mWrc for a standard Beltsville sun.
13
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Table 2. Concentration of anthocyanin in Coleus blumei Benth. leaves as a
function of UV exposure and node position. Basal node = 1.
(Absorbance/g fresh wt. of leaf tissue). One unfiltered FS-40
lamp placed 16 cm above the top of the plants.
Time of
Exposure
(Hours)
1
2
4
8
16
24
Average anthocyanin concentration
(Abs/g fresh wt . ) 525 nm
Node
4
.499
.352
.298
.600
.385
.451
-UV
Node
5
.501
.470
.330
.386
.445
.518
+UV
Average
.500
.411
.314
.493
.415
.485
Node
4
.560
.384
.370
.575
.394
.083
Node
5
.509
.403
.405
.696
.292
.007
Average
.535
.394
.388
.636
.343
.045
14
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Table 3. Change in anthocyanin concentration of Coleus blumei Benth. leaves
with duration of UV exposure under one unfiltered FS-40 lamp at
30 cm. (Absorbance per 3-leaf disks, each 11 mm diameter.)
Time of
Exposure
0
12
24
Anthocyanin
Node 5
.506a*
.264 b
.117 c
Concentration
Abs/3 leaf disks
Node 4
.492a
.269b
.215 b
Average
.499
.267
.166
*Mean followed by a common letter are not significantly different at
the 5% level according to Duncan's multiple range test.
15
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Table 4. Effect of UV exposure on visual leaf injury index.
Duration Injury index
of Exposure*
Days*
3
6
8
Weighted mWm
% Increase in
Mylar 5 mil CA
0 +
0 ++
0 -H-f
0.23 18.15
BUV - 493
10 mil CA 20 mil
0
o
+ 0
9.44 4.
209 48
CA
52
* 6 hours/day
Key: 0 = no injury
+ = moderate (slight leaf distortion with negligible pigment loss)
H- = severe (leaf distortion with 50% pigment loss)
-H-f = intense (leaf distortion with 50% pigment loss)
16
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Table 5. Anthocyanin concentration in Coleus blumei Benth. leaves as a
function of UV exposure.
Duration
of Exposure**
Days
3
6
8
-2
Weighted mWm
%
Increase in BUY
Anthocyanin
Mylar
1.66 ab**
1.28 bcde
1.37 abed
Concentration (Abs/g fresh
5
1.
1.
0.
0.23 18.
- 493
mil CA
50 ab
04 cde
84 e
15
10 mil CA
1.
1.
1.
9.
209
27 bcde
51 ab
49 abc
44
weight)
20 mil
1
1
1
4
48
.74 a
CA
.43 abc
.47 ab
.52
* 6 hours/day
**Means followed by a common letter are not significantly different at
the 5% level according to Duncan's multiple range test.
17
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Table 6. Absorbance/g fresh weight for leaf extracts from unfiltered
UV treatments (FS-40 at 30 cm).
Absorbance/gm at
Treatment Wavelength (nm)
330 420 525
Absorbance Ratio
320/420 420/525 330/525
Control 0.472 0.052 0.068
Unfiltered 0.351 0.100 0.14
UV
9.07 0.77 6.94
3.51 7.14 25.0
18
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Table 7. Chromatographic separation of methanol-extractable components
from the leaves of Coleus blumei Benth.
Band
1
2
3
4
5
6
7
Rf
(x 100)
10.5-19.2
24.5-33.3
35.1
36.0-57.9
59.6
61.4-94.7
94.7 .
Color
Visible
Pink
Red
x
X
X
X
Yellow-green
UV
Rose
Rose
x
dark brown
(absorbing)
x
bright blue
yellow-brown
Absorption maxima
Visible UV
* *
520-535 280
* *
* 270,
* *
* 280,
415, 657 210,
(run)
330
330
230
* No observed maximum
Colorless
19
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LEGENDS TO FIGURES
Figure 1. Influence of shading Coleus. blumei Benth. leaf from UY radiation.
Figure 2. Abnormal growth of upper Coleus blumei Benth. leaves exposed to ;
UV-B (2 FS-40 lamps filtered vdLth 5 rail cellulose acetate and
mounted at 30 cm). Left to right: Plants irradiated 6 hour/day
ป
for 6, 9, and 12 days, respectively.
Figure 3. Response of expanded leaves of Coleus blumei Benth. to UV-B.
Left to right: Plants irradiated for 6, 9, and 12 days respectively
under 2 FS-40 fluorescent sunlamps filtered with 5 mil cellulose
acetate lamps mounted 30 cm above the plants.
Figure 4. Effect of UV radiation on 4-week-old Coleus blumei Benth cuttings
irradiated 6 hour/day for 8 days. Filter treatments (1-r) 5 mil
Mylar, 5 rail CA, 10 mil CA, and 20 mil CA.
Figure 5. Four-week-old Coleus blumei Benth. cutting irradiated 6 hour/day
for 8 days under 2 FS-40 lamps filtered with 5 mil Mylar.
Figure 6. Four-week-old Coleus blumei Benth. cutting irradiated 6 hour/day
for 8 days under 2 FS-40 lamps filtered with 5 mil CA at a 493%
increase in BUV.
Figure 7. UV spectral irradiance obtained under 2 FS-40 fluorescent sunlamps
filtered with 5 mil CA (setup 1), 10 mil CA (setup 2), 20 mil CA
(setup 3) and 5 mil Mylar (setup 4) at a distance of cm above the
canopy. '
Figure 8. Visible absorption spectrum of methanol-extractable constituents
of Coleus blumei Benth. leaves. Plants were grown in a greenhouse
without supplemental UV treatment.
Figure 9. Visible absorption spectrum of methanol-extractable constituents
of Coleus blumei Benth. leaves after 6 hours of exposure to
unfiltered FS-40 fluorescent sunlamps.
-------
Figure 10. Visible absorption spectra of methanol-extractable constituents
of Coleus blumei Benth. leaves irradiated 6 hour/day for 3, 6, and
8 days under FS-40 lamps filtered with 5 mil CA.
Figure 11. Comparative UV absorption spectra of methanol-extractable components
of Coleus blumei Benth. leaves left unirradiated (bottom curve)
.or exposed to unfiltered FS-40 fluorescent sunlamps for 6 hours
(top curve).
Figure 12. Visible absorption spectrum of band 7 obtained from extract of
Coleus blumei Benth. leaves following 6 hours of UV irradiation
under unfiltered FS-40 lamps.
Figure 13. Visible.absorption spectrum of band 7 obtained from extract of
Coleus blumei Benth. leaves taken from an unirradiated control plant.
Figure 14. UV absorption spectrum of band 6 obtained from extract of Coleus
blumei Benth. leaves taken from unirradiated control plant.
Figure 15. UV absorption spectrum of band 6 obtained from extract of Coleus
blumei Benth. leaves exposed for 6 days to 2 FS-40 lamps + 5 mil
CA at 30 cm above the canopy.
Figure 16. UV absorption spectrum of band 4 obtained from extract of Coleus
blumei Benth. leaves taken from unirradiated control plant.
Figure 17. UV absorption spectrum of band 4 obtained from extract of Coleus
*
blumei Benth. leaves following 6 hour of UV irradiation under
unfiltered FS-40 lamps.
Figure 18. Visible absorption spectrum of band 2 obtained from extract of
Coleus blumei Benth. leaves taken from unirradiated control plant.
Figure 19. UV absorption spectrum of band 2 obtained from extract of Coleus
blumei Benth leaves taken from unirradiated control plants.
21
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FINAL REPORT
EFFECTS OF UV-B RADIATION ON PHOTOSYNTHESIS AND GROWTH
OF SELECTED AGRICULTURAL CROPS
J. H. Bennett
Plant Stress Laboratory
Plant Physiology Institute
Beltsville Agricultural Research Center
Beltsville, Maryland 20705
EPA-IAG-D6-0168
Project Officer:
R. J. McCracken
Federal Research, Science and Education Administration
U.S. Department of Agriculture
Washington, D.C. 20250
Prepared for
Environmental Protection Agency
BACER Program
Washington, D.C. 20460
-------
ABSTRACT
Selected snap bean, soybean, clover, cotton, cucumber and wheat
varieties were exposed to UV-B radiation over 2 to 6 week periods (6
or 24 hr/day) under greenhouse and growth chamber conditions. Biologically
effective UV-B irradiances based on an experimentally determined action
spectrum ranged from 1-8 Sun Equivalents (SE). Carbon dioxide exchange
rates (CER), plant biomass production, stomatal diffusion and transpiration
were determined. UV-B effects on CER and foliar diffusivities were correlated
with the amount of visible injury induced (i.e., chlorosis, leaf and petiole
pigmentation, leaf stipple). In the absence of visible injury, CER, leaf
conductances, and biomass production were not measurably depressed in
experimental plants given the extended UV-B exposures.
The plant species differed markedly in their susceptibilities to
UV-B radiation. Greenhouse-grown snap beans and soybeans sustained high
levels before they were injured (in excess of 125 hr exposures3 weeks,
6 hr/dayto more than 4 SE). Wheat and clover were not injured by the
maximum UV-B exposures tested (3-4 week treatments at 2 SE). Poinsett
cucumber developed marginal chlorosis when irradiated for 1-3 weeks at
1-2 Sun Equivalents. Cotton petioles and midveins at the leaf bases
*
became (red) pigmented in the 4 SE trials. Snap bean plants grown under
low-light conditions in the growth chamber were more sensitive to UV-B
injury than when grown in the greenhouse.
-------
INTRODUCTION
Ultraviolet B (280-320 nra) irradiation corresponding to enhanced
levels reaching the earth's surface due to projected stratospheric ozone
(03) destruction by halocarbon emissions has been reported to suppress
photosynthesis in certain agricultural plant species (1-4). Some data
indicate that vegetation grown and exposed to UV-B under low photo-
synthetically active radiation (400-700 nra) regimes may be more sensitive
than when irradiated in bright sunlight (4). Photorepair mechanisms may
be important in mitigating the plant damage.
Experimental plants showing UV-B depressed carbon dioxide exchange
rates (CER) produced less plant biomass when irradiated over an extended
period of time. Brandle et al. (1) observed chloroplast structural damage
in UV-B injured leaves and correlated it with reduced Photosystem II activity.
This was proposed as a causal factor in depressing photosynthesis and
growth. They further concluded from their studies that CER suppression was
not caused by stomatal closure induced by UV-B damage to the leaf epidermis.
Stomatal resistance to gas exchange might increase without specific injury
to the epidermal cells, however, as mesophyll and chloroplast disruption can
result in stomata closure as a consequence of higher C02 concentrations
\ .
within the leaves (as well as other factors).
Little information has been published concerning visible injury
on UV-B irradiated leaves showing reduced photosynthesis and growth.
It was not possible to effectively assess the extent to which growth
reduction correlated with tissue damage and to integrate this with CER
suppression.
Considering the potential importance of the reports referred to above
on impending governmental regulatory actions dealing with the environmental
-------
impact of chlorofluorocarbon use In the United States, this research was
conducted as part of a Federal interagency cooperative project to corroborate
earlier findings and to expand the available information on UV-B radiation
effects on agricultural plants. Research reported here present data on
UV-B exposures and the exposure ranges required to measurably depress
C(>2 assimilation in selected crop plants. Special attention was given to
integrating the results with incipient visible injury. UV-B effects on gas
diffusion through the upper and lower leaf surfaces under typical growth
chamber and greenhouse conditions were also investigated.
Soybean, cotton, wheat, clover, cucumber, and two snap bean varieties
were tested.
METHODS AND MATERIALS
Experimental plants, with the exception of cucumber, were grown
and irradiated with UV-B under (i) common growth chamber conditions
or (ii) in a glass greenhouse. The plants were cultured in 15-cm dia.
clay pots containing a sand-silt (1:3) soil mix. They were fertilized
weekly with Peter'si' fertilizer containing micronutrients. The (heated
or wet-pad, fan cooled) greenhouse was equipped with a high-volume
charcoal air filtration system to prevent plant injury from oxidant air
pollution (5). Cucumbers were grown and irradiated in fiberglass green-
houses in 12.5-cm dia. pots filled with a peat-vermiculite mix (1:1).
They were watered daily with 1/4 strength Hoagland's solution during the
first 3 weeks and with 1/2 strength Hoagland's solution thereafter.
' Mention of a trademark or a proprietary product does not constitute a
guarantee or warranty of the product by the USDA and does not imply
its approval to the exclusion of other products that may also be
. suitable.
-------
The cucumber plants tested were part of another U.S. Department of
Agriculture (USDA) study (6) concerned with UV-B injury and growth
reduction in the sensitive Poinsett variety and more tolerant Ashley
variety.
Growth chamber (Controlled Environments, Inc., Model PCW 36)
conditions are shown in Figure 1. The plants were given 12-hr photo-
periods (0800-2000) utilizing cool white fluorescent + incandescent
lighting. Greenhouse plants received solar radiation. Photosynthetically
Active Radiation (PAR) was monitored with a LI-COR 190-S Quantum Sensor.
UV-B exposures were carried out using standardized procedures (7)
developed for USDA UV-B studies employing FS-40 sunlamps with 5 rail
cellulose acetate (CA) filters. Mylar filters were used for the controls.
Spectral irradiances (250-370 nm range) were measured with an Optronic
Laboratories, Inc., Model 725 spectroradiometer developed by the
Instruments Research Laboratory (IRL), SEA, USDA (8). Spectroradiometer
data were obtained for every nanometer over this range. Routine broadband
UV-B measurements were made with an IRL UV-B Radiometer. Plants were
irradiated for 6 hours per day (between 1000-1600) or, in some companion
experiments, for 24 hours per day.
\
Figure 2 shows a typical UV-B spectral irradiance curve for the CA-
filtered FS-40 sunlamp system obtained at a distance providing a total
_2
irradiance for the 280-320 nm wavelength range of 1079 mWm . The
plant injury Action Spectrum, empirically determined for cucumber and
certain other plants (9), and the weighted biologically effective
ultraviolet irradiance (BUV) are also plotted. For the FS-40 sunlamp
system the summed BUV over the 280-320 nm region, called the Action
Integral ZAป represented approximately 1% of the total UV-B irradiance
-------
f\
(cf. 10 mWm in Figure 2). This varies somewhat as the CA transmission
changes with exposure time (solarizes). UV-B spectral transmission of
the CA filters was routinely monitored with a Beckman DB UV-visible
recording spectrophotometer. Spectroradiometer data were also obtained
for different exposure times. The CA filters were changed every 4 days.
The unweighted and BUV-weighted irradiance curves in Figure 2
represent UV-B exposures for Figure 1 experiments.
Paired (control vs UV-irradiated) intact leaves or whole plants
stratified according to age, position, stage of development, and condition,
were used in each CER trial. The paired foliar subjects were examined
simultaneously under identical conditions in matched Physiological
Activity and Diagnostic Chambers (PhAcDC) which permitted C02 and water
vapor exchange rates and leaf and air temperatures to be continuously
monitored during the experimental runs (10,11). The PhAcDC tests
regulate and standardize physical parameters that enter into the leaf
energy balance equation [ie., Radiant Energy (input) = Reradiation +_
Convection Hh Evapotranspiration ฑ Metabolic Energy (net photosynthesis
or respiration )]. PhAcDC cuvettes were equipped with internal mini-
systems for humidity and wind control. PAR was derived from Quartzline
\
lamps filtered through a 10-cm 1^0 heat filter. Dual PhAcDC experiments
9 1
were conducted at PAR intensities (900 H^ 100 yE nT^sec ) that gave maximum,
but light-unstressed, steady-state apparent photosynthesis (Pmax) rates.
Chamber temperature, relative humidity, and wind conditions were: 27 Hh
2ฐC, 60 + 5% and 0.5 + 0.1 m sec"1, respectively. Soil temperatures were
25 + 2ฐC.
4
Leaf, air, and soil temperatures were monitored with an 11-channel
YSI Telethermometer (Probe types: T2600, T2631). Leaf temperature
-------
calibrations and spot checking were made with a Mikron 15 IR noncontact
Thermometer. Wind speeds were determined with Hastings RF-1 and AB-27
Air Meters (Probes: N-7B and S-27). Soil moisture and pH x^ere determined
with a Bouyoucos Soil Moisture Meter and the Kelway Soil pH/Moisture
Tester, Model HB-2.
Each paired run required approximately 4 hours in the dual-PhAcDC
systems for complete examination and diagnosis including steady-state
Pmax rates, dark respiration rates, and leaf diffusive resistances.
Relative CER data given in Tables 1 and 2 were calculated from steady-
state Pmax rates." Foliar diffusive resistances were computed from
evapotranspiration rates and data taken with a Lambda Diffusion Porometer
(10,11). Plant biomass data were determined from leaf area measurements
and fresh and dry weights. The experiments were conducted during
January-September 1977.
RESULTS AND DISCUSSION
Leaf diffusive resistance; Effects of UV-B and experimental conditions
Results of experiments to investigate the influence of exposure
conditions on gas diffusion through the stomata of UV-B irradiated plants
are shown in Figure 1. Two snap bean varieties (Phaseolus vulgaris cvs.
\
Bush Blue Lake 290 and Astro) grown and exposed to UV-B radiation in
the growth chamber and greenhouse are compared. An inset gives their
relative plant biomasses at harvest. :
Ratios for the upper ru vs lower r-^ leaf surface diffusive resistances
to transpired water vapor are plotted as functions of the total leaf
resistance R. Foliar diffusive resistances reflect the leaf health and
stress physiologyresponding to the moisture balance, phytotoxic agents,
aging, mesophyll C0ฃ levels, and a number of other factors. The adaxial
and abaxial surfaces were compared since it was postulated that UV-B
-------
irradiation could cause more epidermal injury to the upper exposed
surface leading to an early effect on gas permeation through this surface.
Leaf diffusion was more restricted in growth chamber plants than greenhouse-
grown plants. [Growth chamber plants: R = 3-10 sec cnT^; greenhouse plants;
R = 1-3 sec cm""*.] Stomatal opening and development, light dependent
_2
processes, were undoubtedly suppressed under the lower PAR levels (270 pE m
sec~l) of the growth chamber. Consequently, relative photosynthetic rates
would be restricted by diffusion limitations as well as by the lower PAR
available for light-harvesting chloroplast reactions. PhAcDC studies
_2
indicated that snap bean leaves required PAR intensities of about 800 yE m
sec" for maximum photosynthesis.
Diffusive resistances given in Fig. 1 are mean values for all
(<3/4 expanded) first, second and third trifoliates sampled biweekly
over the 4-week UV-B irradiation period. The data were taken during mid-
morning to noonon sunny days in the greenhouseon well-watered plants.
Lower surfaces of greenhouse and chamber-grown leaves exhibiting moderate
diffusive resistances (i.e., 2-5 sec cm"*) were about twice as permeable
as the upper surfaces for both UV-B exposed and control plants. The ru/ri
ratios increased rapidly as higher or lower total diffusive resistances
*
were measured. The arrayed data do not indicate that diffusion through
the upper surfaces of UV-B irradiated plants was significantly altered
relative to that of the lower surface; though, there may be a slight
tendency for increased ru/ri ratios in UV-B exposed plants.
After several weeks in the chamber some UV-B injury was observed on
Bush Blue Lake 290 (BBL 290) bean leaves. [UV-B injury symptoms: Red
pigmentation of the petioles and leaves with slight leaf stipple, i.e.,
scattered small "flecks" of necrotic cells.] Two to three weeks after
-------
losing their cotyledons BBL 290, which is more sensitive to a number of
known environmental stresses than Astro (10,11), showed a gradual loss
of vigor in both UV-B exposed and control plants. As the foliage became
stressed with time in the chamber (perhaps by inadequate photosynthesis),
leaf diffusive resistances increased accordingly. The Astro variety
withstood the growth chamber conditions better than BBL 290 and was also
less sensitive to UV-B injury. The plants were removed after 4 weeks in
the growth chamber and returned to the greenhouse to check for recovery.
At harvest (2 weeks later), UV irradiated BBL 290 plants had less biomass
than the controls (Fig. 1 inset). Astro plants exposed to UV-B radiation
in the chamber did not differ statistically from the controls.
The healthy, vigorous greenhouse plants grew larger than the chamber
plants (cf: C/G, Fig. 1 inset). Foliar resistances remained low throughout
the experimental period. At harvest neither BBL 290 nor Astro plants grown
and irradiated with UV-B in the greenhouse differed in biomass from their
Mylar controls. No visible UV-B injury occurred on the greenhouse plants.
The results tend to corroborate previous reports that plants under
low PAR regimes may be injured more by UV-B irradiation than plants in
bright light. The more sensitive BBL 290 snap bean variety, furthermore,
grew less vigorously in the low PAR chamber environment than the Astro
cultivar. Preliminary trials with Pennseott clover showed this clover
variety to grow well in the growth chamber. The chamber-tolerant clover
_2
was not injured by comparable BUV exposures (E^lOmWin , 6 hrs/day) during
a 4-week trial.
UV-B effects on CER, plant injury, and growth
Table I summarizes the results of CER experiments conducted during
the spring and summer of 1977 on six crop plants exposed to increasing
-------
UV-B doses. Table 2 gives an abridged array showing: visible injury
index ratings for the PhAcDC investigated leaves; comparative leaf
conductances for evapotranspired water vapor (See footnote, Table 2); and
relative plant biomasses of the irradiated plants given as percent of
control. The experimental plants were cultured and irradiated with specified
UV-B doses in the greenhouse and transferred with their paired controls to
leaf or whole plant dual-PhAcDC systems for examination and characterization.
The CER trials were restricted to UV-B exposures that produced no more
than 20% leaf injury (Injury Index II). Data for snap bean, soybean,
cotton, cucumber, and clover were taken on intact leaves or trifoliates.
Wheat and additional clover data were obtained from whole plant studies.
Equivalent numbers of leaves on the controls and UV-B exposed plants were
used. The data were normalized on a weight basis.
UV-B exposed leaves showing no visible injury at the time of testing
did not statistically differ from their paired controls in CER or foliar
conductances, nor was plant biomass reduced by the extended exposures.
CER values were lowered roughly in proportion to the amount of visibly
damaged leaf tissue. Snap bean leaves with an Injury Index I rating
(1-10% injury) showed mean CER values 8% below those of the controls.
Soybean and cucumber leaves assigned to the Injury Index II class (10-
20% injury) gave CER values averaging 14% and 16% below the controls.
High UV-B radiation levels were required to injure greenhouse grown
snap bean and soybean plantsin excess of 125 hrs exposure (3 weeks,
fy
6 hrs/day) to more than 12.5 mWru of biologically effective UV-B radiation.
Cotton was marked by this dosage. Poinsett cucumber was the most sensitive
plant tested. Wheat and clover were not injured by the highest doses given
in those particular experiments. Visible symptoms of injury for cucumber
8
-------
and cotton were well-defined chlorosis along the leaf margins (cucumber)
and red pigmentation of the petioles and juncture with the leaf base (cotton),
Snap bean injury symptoms were described previously. UV-B injured soybean
leaves showed increased pigmentation or bronzing of the leaves with scattered
necrotic stipple.
-? -2
Dividing the Z^(mWm ) in Table 1 and Figure 1 by 3.06 mWm gives
plant BUV exposures relative to one control sunshine equivalent SE. One
SE is the weighted BUV integrated over the UV-B region for the control
sunshine used at the Beltsville Agricultural Research Center (6,8,9).
_ 9
Five mWm , the minimum Z^ included in Table 1, represents 1.6 SE, or a
60% increase above 1 SE. One hundred hours' daytime exposure to 1.6 and
2.1 SE was insufficient to cause injury to any of the soil-grown greenhouse
plants tested. Gregg cotton developed basal leaf and petiolar pigmentation
in the 4 SE exposure tests, but no cellular necrosis or statistically
significant reduction in CER. The very high UV-B levels required to
visibly injure greenhouse snap bean and soybean plants, 8 SE, combined with
nighttime irradiation (the plants were irradiated 300 hrs, 24 hrs/day) are
exceptionally adverse treatments and will not be discussed further. The
data are included to show the range of exposures given.
%
Poinsett cucumber leaves showed some chlorosis when subjected to
ca. 1.2 SE. [See Krizek (6) for experimental data.] The sensitivity of
this cucumber variety varied slightly with season, increasing in the fall
t-
and winter and decreasing in the spring and summer. The seasonal effect
was attributed largely to photorepair in plants grown under the higher PAR.
PhAcDC examined Poinsett leaves exposed to 1.6 and 2.1 SE showed mean
Injury Index ratings of I and II based on the amount of marginal chlorosis
-------
observed (cf. Table 2). Nonchlorotic portions of the leaves appeared to
be healthy and functional. Predicted biomass (dry weight) loss, based on
data generated from the larger population from which the plants were drawn
(48 plants), indicated that 1.6 and 2.1 SE exposures would be expected to
cause 3% and 6% reduction in plant dry weights, respectively (6). This was
less than the mean leaf chlorosis and reduction in CER observed.
Increased BUV corresponding to 1.6 - 2.1 control sun equivalents
represent values greater than the 1.4 SE predicted for the maximum
stratospheric ozone depletion due to chlorofluorocarbon emissions.
Experimental data generated from this study give no evidence that the snap
bean, soybean, wheat, clover, or cotton varieties tested should be
measurably damaged by the proposed UV-B increases. The potential exists
for injury to Poinsett cucumber plants but anticipated growth reductions
of the magnitudes expected would be difficult to statistically detect in .
nature even when some observable chlorotic injury might result.
10
-------
LITERATURE CITED
1. Brandle, J. R., W. F. Campbell, W. B. Sisson, and M. M. Caldwell.
1977. Net photosynthesis, electron transport capacity, and ultra-
structure of Pisum sativum L. exposed to ultraviolet-B radiation.
Plant Physiol. 60:165-169.
2. Sisson, W. B. and M. M. Caldwell. 1976. Photosynthesis, dark
respiration, and growth of Rumex patientia L. exposed to ultraviolet
irradiance (288 to 315 nanometers) simulating a reduced atmospheric
ozone column. Plant Physiol. 58:563-568.
3. Van, T. K. and L. A. Garrard. 1975. Effect of UV-B radiation on net
photosynthesis of some C- and C, crop plants. Proc. Soil and Crop
Sci. Soc. of Florida 35:1-3.
A. Van, T. K., L. A. Garrard, and S. H. West. 1976. Effects of UV-B
radiation on net photosynthesis of some crop plants. Crop Sci.
16:715-718.
5. Heggestad, H. E. 1973. Photochemical air pollution injury to potatoes
in the Atlantic coastal states. Amer. Potato J. 50:315-328.
6. Krizek, Donald T. 1978. Differential sensitivity of two cultivars of
cucumber (Cucumis sativa L.) to increased UV-B irradiance: I. Dose-
*
response studies. BACER Final Report, U.S. Dept. of Agr., 33 pp.
7. Krizek, D. T. 1977. Current UV measurement methodology and future
needs in photobiological research. In Symposium on Ultraviolet
Radiation Measurements for Environmental Protection and Safety.
pp. 49-52. Held at National Bureau of Standards, Gaithersburg, Md.
June 8-9, 1977.
11
-------
8. Norris, K. H. 1977. Development of a portable, automated UV-B
spectroradiometer. In Symposium on Ultraviolet Radiation Measure-
ments for Environmental Protection and Public Safety, pp. 72-72.
Held at National Bureau of Standards, Gaithersburg, Md. June 8-9,
1977.
9. Cams, H. R., R. Thimijan, and J. M. Clark. 1977. Outline of irradiance
distribution of UV fluorescent lamps and combinations. In Symposium
on Ultraviolet Radiation Measurements for Environmental Protection
and Public Safety, pp. 74-76. Held at National Bureau of
Standards, Gaithersburg, Md. June 8-9, 1977.
10. Bennett, J. H. 1977. Ozone and leaf physiology. In Proceedings of the
Fourth Annual PGRWG Meetings, Hot Springs, Ark. pp. 323-330.
11. Bennett, J. H. 1978. Foliar exchange of air pollutants and
physiological gases. In S. Krupa, W. W. Heck and S. N. Linson (eds.)
"Handbook on methodology for the assessment of air pollutant
effects on vegetation", (Chapter 10), Proc. Specialty Conference
of the Air Pollut. Contr. Assoc. Held minneapolis, Minn.,
April 19-21, 1978.
\
12
-------
7 _
6 _
5_
4 _
3 _
2 _
1
O Astro (Control)
Astro (UV-B)
O BBL 290 (Control)
BBL 290 (UV-B)
CHAMBER
X
co CQ--'
*"-*
R range
1it
10
GREENHOUSE
o
R range
Relative Plant Biomass at Harvest
UV /Control:
Chamber
Greenhouse
C(Control).
G(Control)'
(7. Control)
BBL 290 Astro
89* 95
96 102
69*
88*
Significant at
level
-iiiiง
R Csec/cmJ
10
| Experimental Conditions
PAR Light (uE nT2 sec"1)
BUV HA (mW m-2)
Temp (ฐC) [day/night]
R H (7.)
t
Experimental Period: Jan
Growth Chamber
270 ฑ30
10ฑ2.5
[23/16]
35-55
- Mar, 1977.
Greenhouse
1300 (sunny day)
10ฑ2.5
[20-30/15-20]
Usual Range: 30-60
Figure 1. Relative foliar resistances to transpired water vapor
diffusion through the upper r and lower r. leaf surfaces of UV-B
irradiated and control plants plotted as functions of total leaf
resistance R(sec/cm). The data compare plants exposed under growth
chamber and greenhouse conditions. Experimental conditions and
relative plant biomass at harvest (8 weeks of age) are summarized
in block inserts.
13
-------
IU rT
" 1
2
^^ fO
3 'B
09
.u
4J
CO
CC s
d
280
300
320
WAVELENGTH (run)
Figure 2. UV-B irradiance curve for Figure 1 experiments produced
by the FS-40 sunlamp systems filtered through 5 mil cellulose acetate
(exposed 6 hours). Also shown are the Action Spectrum for UV injury
to cucumber (and other) plants (9), and the Biologically-harmful UV
Action IntegralZ. (inset). The Action Integral represents the integ-
rated product of the Action Spectrum Y times the Irradiance I as a
function of wavelength.
14
-------
Table 1. Summary of relative photosynthetlc rates (nee carbon dioxide exchange rates, CER)* for UV-B
irradiated and control plants [n 5-8 replications per treatment).
UV-B Exposure
(Expt'l. Period: Apr.-Sept. 1977J
Relative CKR
(Z of Control)
Exposure Range UV-B Snap beans Soybean Clover Wheat Cotton Cucumber
IA (mWatts m-2) Exposure Exposure CER Test "BBL 290" "York" "Pennscott" "Monon" "Cregg" "Polnsett"
(Cf. Fig. 2) (Hours) Class (Weeks)
5.0*0.5(6 hr/day) 100*20
ซ.5ฑ0.5( " ) "
12.5ซ.5( " ) 125ฑ25
25*5 (24 hr/day) 300150
2-3
3-4
4-6
I
MEANS INSIDE BOUNDARY NOT SIGNIFICANTLY DIFFERENT I
105 99 103 107 [ 84*
97 102 95 |
92* 86*
*CER for Intact leaves calculated as ug C02 dm" mln"1; Wheat and clover normalized on weight basis.
One control sun equivalent (SE) equals 3.06 mWatts or2 based on action spectrum given.
*Meซns Significant at P QJ level
Table 2. Summary Table showing relative net carbon dioxide exchange rates for UV-B exposed and control
plants arrayed to Include leaf Injury indices, leaf conductance ratios for transpired water
vapor, arid relative plant blomass.
Flint/Variety
Snap bean
"Bush Blue
Lake 290"
Soybean
"York-
Clover
"Pennscott"
Wheat
"Monon"
Cotton
"Cregg"
Cucumber
"Polnsett"
UV-B
Exposure
Class
[See Table 1]
C
D
C
D
B
B
C
A
B
Plant
Blomass
(7. Control)
105
94
96
92
101
99
93
97C
94C
Relative
CER
(T. Control)
97
92*
102
86*
% 103
107
95
94
84*
Ratio8
LUV/LC
1.0
0.9
1.1
0.9
1.1
*
1.0
1.0
0.9
Leaf
Injury
Indexb
0
I
0
II
0
0
Tr
I
II
Symptoms of Injury
No visible Injury
Leaf pigmentation; stipple
No visible Injury
Leaf bronzing; stipple
No visible Injury
No visible Injury
!
Pigmentation of petiole and
base of leaf
Marginal chlorosis of
ii . n ii
leaf
II
Ratio of leaf conductance L (reciprocal of leaf resistance) for UV-B exposed and control plants.
Conductance roefflcients linearly relate gas exchange rates with concentration gradients.
blnjury Index Scale: 0 (no visible injury); Tr ซ17. injury); I (1-107.); II (10-207.)
cPlant btomass (dry weight) based on regression data generated from 48 cucumber plants
15
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ffi.au
. ซ??
BIOLOGICAL AND CLIMATIC. EFFECTS RESEARCH
TERRESTRIAL NON-HUMAN ORGANISMS
EXECUTIVE SUMMARY
EPA-IAG-D6-016S
Project Officer:
R. J. ^IcCracken
Agricultural Research, Science and Education Administration
U.S. Department of Agriculture
Washington, B.C. 20250'
Prepared lor
Environiiiental Protection Agency
BACER Program
Vfashington, D.C. 20460
-------
EXECUTIVE SUMMARY
Tlits final report describes the research undertaken by the U.S. Department
of A'.'r tculture during the period of October 1, 1976, to February 28, .1978, as
;i parr of the Biological and Climatic Effects Research (BACER) Program
conducted under interagency agreement with the Environmental Protection Agency.
Tin- objective of this research was to assess the biological impact of in-
c>.v.risecl radiation in the UV-B region (280-320 nui) reaching the earth's surface
on ;i)\r {.culturally important and native terrestrial plants and animals that
ini>;ht; result from stratospheric ozone reduction caused by inadvertent release
of ciilorofluoromcthanes (CFM's). . - -
The organizations reporting and participating in this research arc
li:;ted below:
U.S. Department of Agriculture
U.S.. Forest Service, Fort Collins, Colorado
Science and Education Administration
Arizona-New Mexico Area, Las Cruces, New Mexico
Beltsville Agricultural Research Center, Beltsville, Maryland
Agricultural Equipment Laboratory
Chemical and Biophysical Control Laboratory
Florist and Nursery Crops Laboratory
Instrumentation Research' Laboratory-
Organic Chemical Synthesis Laboratory
Plant Stress Laboratory
Florida-Antilles Area, Gainesville, Florida
National Animal Disease Center, Ames, Iowa
Northern Regional Research Center, Peoria, Illinois
-------
Federal Research Contracts
University of Florida, Fruit Crops Department, Gainesville,
Florida
Colorado State University, Department of Horticulture, Fort
Collins, Colorado
Major accomplishments resulting from the U.S. Department of Agriculture
liACER Program are as follows:
1. An1 automatic UV spectroradiometer was designed, constructed and
successfully tested capable of measuring UV radiation every.
.
nanometer (ma) from 250 to 400 nra, with a wavelength precision
of,0.1 run (1A). Broad- and narrow-band radiometers to monitor the
output of artificial UV sources for laboratory, growth chamber, and
greenhouse' experiments were also developed.
2. Substantially all major crops, including many horticultural species
and varieties, and some native species have been screened for
sensitivity to increased.levels of UV-B. ...
3. Injury threshold levels were found to vary widely among species
ancl within cultivars and varieties of the same species.
4. It was almost universally observed that field grown plants were
more tolerant to enhanced UV-B levels than we're the identical
selections grown in greenhouses or controlled environment chambers.
A seasonal difference in sensitivity to enhaiiced levels of UV-B
was also observed, with plants showing greater tolerance during
the summer months. Substantial evidence for the existence of a
high-light intensity photorcpair or photoprotection mechanism in
higher plants was obtained.
-------
5. Some economic plant species, such as cantaloupe, soybeans, and cotton
have been shown to be sensitive to present levels of UV-Ji reaching
the earth's surface.
6. Highly pigmentcd insects, especially those with a high melanin
content, were found to be highly resistant to UV-B. Honeybees,
for example, were unaffected by very high levels of UV-B. Some
leaf disease organisms display similar responses.
7. We have established that "cancer eye" may have been induced in
Hereford cattle by high levels of UV-B.
These accomplishments provide a significant increase in knowledge
of the potential responses of terrestrial plants and animals to increased
UV-B irradiance. Because of continued uncertainties involved in this
research, the results do not permit the development of conclusive
statements concerning the possible harmful effects to be expected from
increased UV-B reaching the earth's surface as a result of the projected
reduction in stratospheric ozone as a result of release of the chloro-
fluoromethanes in. the biosphere.
The uncertainties are addressed below:
Artil'icJat Enhancement Sj^rc
-------
Figure 1
10BO00
JLS_1O..5SC!{ _
JES.40 .ZJ3CHi5J.IIl.JX
J81i.4MLG.Mo-.LM . ..Z
100.000-
18,009-
k l.B0a
1.J.1
C.J
eg
0,109-
0.010-
0.001-
WAVELENGTH nm IRL AHRI FR USDA
3 a
-------
The first problem seen is that the region of; major interest 280-320 nra
is one of rapidly decreasing irradiance; decreasing over four orders of
magnitude. Unfiltered lamps do not parallel this decrease in intensity,
displaying higher intensities as the. wavelength decreases. As can be .seen,
this discrepancy can be partially overcome by use of suitable filters; 5 mil
cellulose acetate for example (+ 6 hrs. 5 m CA).
The second, and perhaps as serious a difference is the greatly decreased
irradiance from artificial sources from approximately 310 nin and above. The
possible effect of this reduced irradiance level in the UV-A region can only
be conjectured at our present state of knowledge.
A third uncertainty, which we believe to be at least partially resolved
is the. fact that energy emitted by a fluorescent lamp and falling on a
horizontal surface parallel to the long axis of the lamp is unequal; being
highest directly under the center of the lamp and falling off rapidly in
all direction;; from the center. By using improved instrumentation and
through development of equations defining this energy variable, careful
energy-defined biological research has been accomplished.
However, continued use of these sources for UV enhancement experiments
will require improvements in UV filters and in the sensitivity, accuracy
and utility of UV radiometer and spectroradiometer equipment. Development
of new sources more nearly simulating*the sun will prove expensive and
probably require the development of new or improved technology.
Ae_ticm Spectra. Since the artificial enhancement sources do not
simulate the sun and because biological organisms and systems do not respond
equally to all wavelengths of UV radiance, the interpretation of data derived
from UV enhancement experiments requires the use of a suitable weighting
-------
function derived from an action spectrum to describe the comparative
biological effectiveness of different wavelengths in the. UV region to which
the organism is subjected. At the beginning oC the short-term BACliR
program there was no universally accepted action spectrum for higher plants .
although several had been proposed (Figure 2). The Uwo action spectra (AS)
which have been evaluated for data interpretation in the current program are
graphically shown as 9AS (AE9) and 21AS (Aฃ21).
Weighting functions derived from these two spectra have been used by
participating.principal investigators. The 9AS weighting function has
generally been applied by most researchers. However, the University of
Florida and the Florida-Antilles Area report their results using the 21AS
weighting function. .
The Agricultural Equipment Laboratory report contains a presentation
of results obtained by Beltsville Agricultural Research Center investigators
which suggest that the 9AS weighting function is more valid than the 21AS
weighting function. It should be emphasized, however, that data are still
lacking as to how accurate or how universally applicable this weighting
function will prove to be. Use of the 21AS weighting function in place of
the 9AS tends to overestimate the effectiveness of the longer wavelengths
thereby underestimating the amount of biologically effective UV radiation
(BUV) received. Thus, tesฃ organisms are subjected to higher total UV
irradiances with the 2.1AS weighting function than with the 9AS weighting
function for a supposed equal amount of biologically effective UV.
To provide a. basis for interlaboratory comparison of data, all principal
investigators have provided irradiances used in experimental set-ups in
absolute uirweighted mW;n~ " thus, when a verified action spectrum has been
established, all submitted data may be reinterpreted on the basis of the
accepted action spectrum.
-------
Figure: 2
.
CD
CO
t"
C.J
IIAVELENGTH nm IRL AMRI FR U'S
5a
-------
Contro 1 Sii_nghino. AnoLhpr major problem in interpretation of UV-
irracliance enhancement studies is the variation in the sun's UV-irradiance
as a function of latitude, elevation, zenith angle, and the sun's variability
from clay to day as the result of stratospheric and atmospheric changes. This
is illustrated in Figures 3 and 4 showing linear and log plots of UV
irradianccs obtained every nanometer from 290-320 nm using a single or double
raonochromator spectroradiometer. Starting with the sun spectrum with the
highest overall spectral energy, the data were obtained at: Snowmass,
Colorado, 9,777 ft. (2980 in) on August .10, 1977, 1400 hours; Beltsvillc,
Maryland, 186 ft. (56.7 n), June 30, 1977, 1337 hours; Gainesville, Florida,
180 ft. (54.9 in), April 28, 1977., 1432 hours, (University of Florida
report. Table 10); and Be.Usville, Maryland, March 21, 1977, 1400 hours.
Asterisks on Figure 3 represent the control sunshine used by investigators
at BARC derived from data obtained by the Smithsonian Radiation Research
Laboratory, Rockville, Maryland, and the Instrumentation Research Laboratory,
Beltsville, Maryland. It basically represents the average sunshine for
the months of June and July .1976 in the Washington, D.C., area.
As can be seen on Figure 4, because of stray light response of the
instrument used at Gainesville, Florida, irradiances are overestimated at
the shorter wavelengths (290-300 nm); this presents a serious problem when
'
weighting functions are required since greater weight is applied to shorter
x>7avelengths. .
9
Table 1 presents the total unweighted mWrn from 290-320 nm, the Aฃ9
-2
weighted mWm " and the fraction of one average control sunshine used in
the Beltsville investigations. The biologically effective UV-B measured
at Snowmass, Colorado,, was 2.7 times that of the Washington, D.C., area.
A 20 to 40 percent enhancement of the measured biologically effective UV-B
-------
0. 8-
633. B
Figure 3
J1QRLQA Jil^ J JB._Z7_2^.H JSI
JEITSVUIH SUN.. G 32J7 i, 37 P;
mORAl2Q^OJ3_ZLJ BOOT
.!
., WAVELENGTH nr., IRL AHRI FR USDA
6a
-------
Figure A
WAVELENGTH nm IRL AMRI PR USDA
6b
-------
_o
the. Colorado location would range from 9.95 to 11.61 weighted mWm ,
whereas an identical enhancement at Washington, D.C., would range from
-?
3.67 to 4.28 weighted mWm ~. The need for biological research to cover a
wide range of biologically effective UV-B irradiances is obvious.
To properly evaluate the results presented here United States-wide
will require accurate knowledge of the sun's spectral energy in all geographic
regions. The development of a program for regular monitoring of the UV
spectral irradxan.cc at selected locations is a necessity which cannot be
ignored in future programs.
Research results obtained by each principal investigator are summarized
in the following pages.
-------
Table 1
Location
Snowmass, CO
Beltsville, MD (high)
Gainesville, FL
Beltsville, HD (low)
Control sunshine
Total
ir radiance
290-320 nm
o
mWra
4098
2499
1742
1274
2847
Biologically
effective UV Fraction of
irradiance AE9 control sunshine
290-320 nm
2
v:eighted raVrm
8.29 2.7
3.94 1.3
2,, 68 0.7
1.60 0.5
3.06 1.0
(Beltsville)
-------
RESEARCH PROGRESS . -
lojinienI: of Ins 1:rument ation for MeasurIng UV--V> J
-------
scan, reverses the wavelength drive and returns the monochromator to the
starting wavelength. At tlie completion of the scan, the data can be stored
on a magnetic tape for future analyses or transferred to an automatic plotter.
A miniature low-pressure mercury-arc lamp is used to provide a precise check
of wavelength accuracy. The calculator computes the position of the 233.7-nin
and the 296.. 7-nm mercury lines to a precision of + 0.01 nm.
In order to provide investigators in the program with a simple instrument
for monitoring the output of artificial UV sources in laboratory, green-
house, and growth chamber studies, broad-band and narrow-band UV radiometers
were also developed by the Instrumentation Research Laboratory. These
instruments feature a teflon bubble cosine receptor, a so.lar--b.lind phototube,
a battery-powered photometer circuit and a small rugged housing. In order
to provide a basis for interlaboratory comparisons of UV data, the sensitivity
of the broad-band radiometer was adjusted to give the same full scale
reading under a common UV source CFS40 fluorescent sunlamps filtered with
i
!
5 mil cellulose acetate). Correction factors were developed for use under j
other lamp-filter combinations. . j
i
The specifications for these instruments were made available to industry,
and commercial models have now been developed and obtained by cooperating
locations. v
Spectra.'l. Characteristics of Fluorescent Lamps arid Testing of Weighting
Functions
The Agricultural Equipment Laboratory at Beltsville, Maryland, developed
mathematical equations describing the distribution of normalized UV irradiance
levels in any desired combination of lamps, either parallel or end-to-end,
at any defined distance of the lamps from the illuminated surface. A computer
10
-------
program was written which permitted scientists to accurately design
biological experiments and interpret the results.
Lamp fixtures were fitted with specially designed reflectors to provide
greater uniformity and rcproducibility in UV radiation experiments. Multiple
fixture assemblies were designed and built.
An average or standard spectral distribution of UV energy of the sun
for .the Middle Atlantic area was derived from data collected by the
Smithsonian Radiation Laboratory, Rockville, Maryland, in June-July 1976,
and by the Instrumentation Laboratory, Beltsville, Maryland. This is now
being used by Beltsville scientists as a base, for design of UV enhancement
experiments.
In conjunction with the Florist and Nursery Crops Laboratory and the
Plant Stress Laboratory, experiments with higher plants were undertaken
to more clearly define the weight, ing function that must be used to interpret
results, since the spectral distribution of the fluorescent lamps does not
duplicate that of the sun. Several weighting functions were investigated.
The equation providing the best fit was derived and is being used by the.
majority of investigators. We are continuing to obtain data testing the
applicability of this weighting function.
Biological E ffects of UVB Radiation/von Plant Growth and Function
Greenhouse, growth chamber, laboratory, and -field studies were conducted
by the Plant Stress Laboratory at Beltsville, Maryland, on a wide range' of
vegetable and agronomic crops to determine the relative sensitivity or
resistance to increased L1V-B radiation. Data were collected on various
physiological responses to increased UV-B radiation including: photosynthesis,
respiration, ion uptake, translocation of rad.ioisotopcs, stomatal activity,
changes in chlorophyll and anthoc.yanin content, leaf movement, germination,
1.1
-------
seedling growth and reproductive development. Studies were also conducted
On UV-B interactions with plant disease organisms.
Broad-band UV-B studies were conducted in the greenhouse and growth
chamber on over 20 species and cultivars of vegetable and agronomic crops.
Plants were exposed to a gradient of IJV-B radiation representing a 50 to
500 percent increase in biologically effective UV radiation. Plants studied
included cotton, peanut, wheat, rice, alfalfa, cucumber, pea, beet, tomato,
rutabaga, okra, bean, radish, and turnips Most plants were exposed to UV-B
for 4-5 weeks from time of planting the seed, but a few (wheat:, rice, alfalfa,
cotton) were grown to maturity under elevated UV--B. Visual injury was observed
in over half of the species and cultivars s.tudied. In most cases only slight
or moderate UV damage was noted even when the. plants were exposed to an
increased level of biologically effective UV radiation as high as 300-AOO
percent.
The most dramatic evidence of UV-B injury was chlorosis in pea and
cucumber, necrosis in pea leaves and pods, and reduction in leaf size in
pea and cucumber. ^
Dose-response studies conducted on cucumber varieties demonstrated
significant differences in UV-B sensitivity; Poinsett cucumber was extremely
sensitive and Ashly cucumber was only slightly sensitive. Evidence, was
obtained for UV-B induction of chlorosis of the-leaves, inhibition of leaf
enlargement, and reduction in biomass. These effects were most pronounced
under conditions for low photosynthetically active radiation, and high UV-B
exposure.
High levels of UV-B irradiation in the greenhouse (100-400 percent
increase in biologically effective UV) reduced the total number of kernels
in Pacific Triple Uwarf wheat by 20 percent, but had no appreciable effect
on the average yield.
12
-------
Translocation of radioactive zinc from the cotyledons to other plant
parts of the young cotton was not inQuencod by a lOO-AOO percent increase
in biologically effective UV; however, the transport of radioactive calcium
was depressed 12-30 percent over this range of UV irradiation.
Based on linear regression analysis of plant data .obtained in the
greenhouse of one of the more sensitive plants (Poinsett cucumber) exposed
to-increased UV irradiation (from 50-30CH- percent increase in biologically
effective UV), it was estimated that a maximum decrease in stratospheric
ozone content of 20 percent would cause a 10 percent reduction in dry matter
accumulation and a 15 percent decrease in leaf area. It is not possible
at the present time to determine whether these estimates 'can be applied to
oUier species of higher plants.
Measurements were made on net photosynthesis rates, plant biomass
production, stomatal diffusive resistance, and transpiration rates in
selected plants of snap bean, clover, cotton, cucumber, and wheat irradiated
in the greenhouse and growth chamber. In general UV-B effects on net COp
exchange rates and foliar gas exchange were correlated with the amount of
visible injury induced.
Chromatography and subsequent UV and visible spectroscopy of acidic
methanol extracts of Cole us bljjmejL_ 'leaves taken from UV-B irradiated plants
demonstrated a degradation in UV absorbing compounds. Similar results were
obtained with reflectance measurements. Increasing the UV-B. irradiance
resulted in increased degradation of anthocyanin pigment, reduction in the
rate of leaf expansion, inhibition of apical growth, and abnormal development
of the leaves.
.13
-------
Field studies were conducted on UV effects at Be.ltsville on a range
of agronomic and vegetable crops using a gradient of UV radiation
developed by the Agricultural Equipment Laboratory, Crop plants studied
included Contender bush bean, Early Prolific, straightneck yellow squash,
Amsoy-71 soybean, sugar beet, Golden Cross Bantam corn, R-720 sorghum, and
Waltham 29 broccoli, A fall crop of winter grains was also grown that
included Potomac, Redcoat, and-Abe whea.t, Pennard and Monroe barley, and
Abruzzi rye.,
Increasing the biologically effective UV radiation by 100 percent had
no visible or consistent effect on crop performance under field conditions.
P.lant Disease T.nterac tion _w.iLth UV-B Radiat_icm
The Plant Stress Laboratory has studied the effects of UV-B radiation
on plant diseases. The results of increased levels of UV-B irradian.ce on
spore germination indicate that although plant leaf pathogenic fungal
species vary considerably in sensitivity to UV-B, relatively high irradiance
levels are required to reduce germination percentage. Pigmented spores such
as Cladosporium, Steinphyllium, and Alternar.ia were found to be more, resistant
to increased UV-B irradiance than hyaline spores (Mycosphaerella t
ColletotrichuTTi). -
Disease .severity of Co 11 eto tr i c hum lagenarium on cucumber v;as decreased
with increasing UV-B irradimic.es. A linear decrease in the percentage of
leaf area diseased with increased irradiances was found.
Increased levels of UV-B irradiance did not affect disease severity
ฐ^ Cladosporium cucumerinum. The disease tended to reduce plant growth
equally regardless of UV-B irradiance levels.
. There was no noticeable UV-B effects on either Che Ste-mphyllium
botryosuni pathogen or the host, alfalfa.
14
-------
In summary, recognizing that our results represent only a small
sampling of leaf disease organisms and of plant disease-interaction
experiments, they appear to support the following: (1) considerably higher
levels of UV-B irradiances than those expected from the projected ozone
depiction will be required to adversely affect germination and growth of
pathogenic fungi, and (2) where fungal germination and growth are affected,
disease severity in the host plant can be expected to be reduced as UV-B
irradiances increase.
Response of Florist arid -Nursery Crops to Incre ased UV-B Rad.1 at ion
Greenhouse and growth chamber studies were conducted by the Florist
and Nursery Crops Laboratory at Beltsville, Maryland, on a wide range of
florist and nursery crops to determine their relative sensitivity or
resistance to increased UV-B radiation. Selected plants were also chosen
for reflectance and fluorescence measurements and for microscopic examination
in the laboratory. After 2-8 weeks of exposure, visible injury was observed
in eight of the 58 species irradiated, and then only when applied in excess
of projected levels of UV-B radiation expected to result from CFM-cata.lyzcd
reduction of stratospheric ozone. .
The most typical response to high levels of UV-B irradiation (100
percent or greater increase in biologically effective UV) included break-
down, of chlorophyll and anthocyanin and a glazing and browning of the tissue,
generally attributed to the presence of oxidized, polymerized, phenolic
)
compounds. Other effects observed in some of the test plants included [
!
abnormal leaf growth, characterized by reduced size, twisting and distortion, j
[
and reduced plant height. Plant bioinass was generally unaffected when mature i
I
plants were irradiated; bioinass of young seedlings, however, was frequently !
depressed under high UV-B. . ;
15
-------
There was considerable variation in sensitivity to UV-B exposure,
depending upon species and cultivar, stage of development:, time of. year, and
level of exposure. In general, herbaceous plants were more sensitive to
increased UV-B than were woody plants. Fat si3 japonica was the only woody
species of the ten tested that showed inhibitory effects of high UV-B.
Plants irradiated during the summer months in the greenhouse showed
little or no UV injury, even under the highest levels of UV-B used. This
was in sharp contrast to the spring and winter months when they showed
considerable injury under the same level of UV irradiance.
Poinscttia, Coleus, and Browallia were among the most sensitive examined.
Other species sensitive to increased UV-B irradiation included aster,
hollyhock, vinca, and impatiens.
In order to develop a capability for understanding the basic cellular
and ultrastructural mechanisms of UV-B effects, a UV microspectrophotome.ter
was obtained and. assembled in the Florist and Nursery Crops Laboratory.
This instrument will enable researchers to irradiate single cells or
cellular constituents as small as 0.5 micrometer with narrow band UV radiation,
to make rapid scans of absorbance and reflectance in the region of 250-
1000 ntn, and to make precise measurements of UV fluorescing materials.
A programmable calculator was also obtained to control the instrument and to
provide on-line data acquisition, processing, storage, and display.
Influence pf Solar UV-B Radiatj.cm on Crop Productivity
Greenhouse, growth chamber, and field studies were conducted by the
Fruit Crops Department, University of Florida, Gainesville, Florida,
on a wide range of vegetable and agronomic crops to determine their
relative sensitivity or resistance to increased UV-B radiation.
16
-------
Field studies were conducted at Gainesville under specially constructed
UV-B gradients obtained by mounting the fixtures at an angle over raised
plant beds. Crops grown to marketable size and maturity included corn,
potatoes, tomatoes, field peas, peanuts, rice, squash, mustard, and radish.
Visual effects were observed in corn and rice under high UV-B irradiances
(100 percent or greater increase in biologically effective UV). Both crops
appeared dwarfed and the grain head of the-rice plants were slower to mature
than the unirradiated controls.
Growth chamber studies were conducted by University of Florida researchers
at the Phytotron at Duke University in Durham, North Carolina. Over 100
species and varieties' of agronomic, horticultural and forest plants were
grown from seed for 4-12 weeks under increased UV-B radiation. Under high
levels of biologically effective UV-B radiation (100 percent or greater
increase) plants exhibited a number of abnormal responses. These included:
Marginal and inte.rveinal chlorosis; cupping and epinasty of the leaves;
changes in pigmentation; increased branching; reduced vineness; and reduction
in height, leaf area.> and biomass.
In general., plants within the same family responded similarly to increased
UV-B radiation. By using controlled environment studies it was possible to
identify varieties of soybeans that are sensitive to present levels of
<,
UV-B radiation at Gainesville, Florida.
Preliminary studies conducted in the Phytotron with Jori wheat and
liar dee soybean, at four levels of UV-B radiation and four levels of visible
radiation showed that the extent of UV-B radiation damage was greatly
influenced by the amount of visible radiation present. Other research
accomplishments of the Univcristy of Florida scientists included developing
17
-------
an action spectrum for pigment induction in the avocado leaf having a
maximum effectiveness in the UV-B region at 293 nm. These investigators
also found that increasing the level of biologically effective UV radiation
by up to 100 percent had no significant effect on structural changes or
chemical composition of surface waxes of tomato and pepper plants. Agronomic
plants subjected to high levels of UV-B radiation also produced increased
amounts of ethylene and accumulated larger quantities of abscisic acid than
control plants.
Response of Vegetable Crops to High UV-B Radiation at Ilipji Elevations
UV-B enhancement and exclusion studies were conducted by the Department
of Horticulture, Colorado State University, at a 3000 m site elevation in the
Rocky Mountains.
Supplementing natural solar radiation with additional UV-B radiation had
no significant effect on the growth and biomass of pea, radish, potato, and
wheat grown at this elevation. Shielding wheat plants from natural solar
UV-B, however, resulted in an increase in size of the plants.
Other accomplishments included the development of transmission spectra
for a chlorinated-fluorinated resin film "Aclar" found to be useful in
aquatic studies as a UV transparent film; development of an assay for detecting
loss of electrolytes from UV-irradfated plant tissues; characterization
of the influence of low temperatures on decline1 in lamp output of UV fluorescent
sun lamps; and design of a solar UV-B collector and irradiator.
Response of Arid and S_emi--arid Plants to Increased UV-B Radiation
SEA scientists at Las Cruces, New Mexico, investigated selected native
and economically important species indigenous to the arid southwest United
States. Plants were exposed to increased UV-B irradiation in the greenhouse.
18
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Dose response studies were conducted on alkali sacaton (Spgrobolus avroidcs
Torr.)> mesa dropseed (J3. flexuosus) and Chile pepper (Capsicum frutosceiis) .
Alkali sacaton and Chile pepper plants exposed to high UV-B showed a marked
reduction in leaf growth with increasing UV-B. Mesa dropseed plants, however,
showed no differences in leaf growth between UV-15-irradiated and control
plants. Dock plants (Rumex patientia L.) exposed to high levels of UV-B
showed a reduction in protein synthesis.
Impact of Solar Radiation on Crops and Crop Canopies__
Physiological and ultrastructural studies were conducted by SEA researchers
at Gainesville, Florida, on selected vegetable, agronomic, and citrus crops
exposed to increased UV-B radiation in the field and the greenhouse.
Citrus plants irradiated for 4 weeks under supplemental UV-B radiation
in the field showed no significant reduction in average daily photosynthctic
rate as compared with unirradiated control plants even under a 200 percent
increase in biologically effective radiation. Similar results were obtained
in stomatal diffusion resistance of eight soybean varieties.
Broad-band UV-B enhancement studies were conducted in the greenhouse on
soybeans (Bragg and Altona), peas (Lit;tie Marvel), tomatoes (Rutgers) and
sweet corn (Golden Cross Bantam). Plants were grown for 4-6 weeks under
three levels of UV-B irradiation ranging from approximately a 100-200
percent increase in biologically effective UV. Da'ta were taken on biomass,
C02 uptake rate, chlorophyll content, Hill reaction, RuDP carboxy.1 ase,
PEP carboxylases soluble proteins, absorption spectra of pigment extracts,
and ultrastructural changes in selected cultivars.
19
-------
Ill general, plants exposed to high UV-B irradiation in tlie greenhouse
showed physiological changes. For example, soybean plants showed . ';
a decrease in chlorophyll content, RuDP carboxylnse. activity, soluble protein
content, COo uptake, and fresh and dry weight as compared with control ,
?"
plants receiving only UV-A (320-400 nm) irradiation alone or unirradiated !.
.i.
control plants. In contrast, plants given supplemental UV-B irradiation r
'('
in the. field showed little or no effect. r
l
Differences in species and cultivar response to increased UV-B irradiation j'
i
were also observed. Differences in chemical and structural makeup of the j'
l;
epidermis and palisade parenchjina cells were thought to play a role in j
the response of different plants to enhanced UV-B radiation. " ;
Response of V?pp_dy Plants to Ipcreased UV-B Radiation^ j
' . i;
Various physiological disorders of agronomic and horticultural crops ,
and woody species have been ascribed to high levels of solar irradiation, i
l'.
especially at high elevations. In order to determine the role of UV-B
radiation in solar injury of certain woody plants at high elevations, Forest
Service scientists conducted UV-B enhancement and exclusion studies at the
Rocky Mountain Forest and Range Experiment Station in Fort Collins,
Colorado. Englemann spruce was chosen, as a sensitive species and Lodgepolc
p.ine was chosen as resistant specias. "
Seedlings were irradiated under artificial UV lamps for a total of
AGO hours over a 67-day period or were grown under various filters to exclude
natural UV-B radiation. No evidence of UV injury was observed in any of the
treatments during the first year of the study. Since Engelmann spruce
seedlings transplanted to the natural environment do not show symptoms of
solar radiation injury until after the first winter, seedlings will be
observed for symptoms during the second growing season.
20
-------
Rejjj>on_s c o f N :L t r o g en - f :i. x ing 0 r v; a n isms to Increased UV-B Radiation
Nitrogen fixation of Anabe.na floss-aquae and other blue-green algae,
free-living and in symbiosis with the water fern Azolla, .is important in
rice culture and in worldwide soil fertility. Laboratory studies were,
therefore, conducted at the SEA Northern Regional Research Center in Peoria,
Illinois, on- the influence of increased UV--B radiation on the nitrogen fixing
abilities of Anabena alone and in association with A/.olla.
Results indicated that while viability and photosynthesis of Anabena
cells were unaffected by UV-B irradiation, nitrogen fixation (as measured by
nitrogenase activity) was markedly reduced by high levels of UV-B irradiation.
*^
When cultures of blue-green algae were exposed to 10 watts/in of UV-B
for 3-5 hours, the algal nitrogenase was inhibited to about one-half of the
activity of the control cultures. In spite of this observed reduction in
nitrogenase activity there was no reduction in the treated cells'
reproductive capacity as demonstrated by plate count studies.
When blue-green algae and its symbiont Axo_lla_ were exposed for 4-6
*) 1 /
days with 10 watts/m of UV-B, photosynthesis (C CU fixation) was not
affected, and nitrogenase activity was reduced to 30-40 percent of that
observed in the control.
Response of Farm Animals to Increased UV-B Radiation
Studies conducted by SEA researchers at the National Animal Disease
Center in Ames, Iowa., investigated the carcinogenic, effect of high levels of
UV-B irradiation on the eyes of four Hereford cattle. Exposing .eyes of
these cattle to high levels of UV-B irradiation for 2 hours per day induced
ocular changes that were consistent with chronic irritation. After 7
months of exposure, one animal developed ocular changes that were considered
ncoplastic after biopsy and pathological examination. Cancer eye, a squaiiious
21
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cell carcinoma, normally takes 5-6 years Co develop in cattle under natural
conditions.
Inspection of; slaughterhouse condemnation .records was made to determine
the extent and incidence, of cancer eye in cattle, using USDA Meat and. Poultry
Inspection data. Based on the total cattle slaughtered since 1950, the
increase in cancer eye was about two-fold. Since other diseases of cattle
have also increased during this time period, it is difficult to interpret
these data.
Response ^of Insects to Increased UV-B Radiation
Studies were conducted in the Chemical and Biophysical Control
Laboratory at Beltsvillc, Maryland, on the influence of increased UV-B
radiation on the physiology and behavior of selected beneficial and
harmful insects.. These studies demonstrated that lightly pigmented insects,
such as the pink bollworm, codling moth, and the face fly, were much more
sensitive to exposure to UV-B irradiation than heavily pigmented ones such
as the house fly.
Brief exposure of pink bollworr.t. eggs to UV-B radiation levels (10-50
percent above natural levels at Beltsvillc), for 1 to 3 hours greatly
reduced the life span, of larvae hatching from these eggs. High UV-B levels
.. v
(100 percent increase in biologically effective radiation) also had a
highly lethal effect on face fly pupae irradiated for 1 hour per day for
3 days.
Adult honeybee workers, however, were able to tolerate a 10-50 percent
increase in UV-B radiation for 6 hours per clay for 6 days without apparent
injury.
22
-------
High UV-B irradiation increased pigment; formation in the .larvae of
butterflies and moths, and the pupae of face flies, but had little or no
effect on the pigment content of tobacco budworm larvae, house fly pupae,
or honeybees.
Physiological studies on respiration indicated an increase in oxygen
uptake in codling moth larvae irradiated 6 hours per day for 2 days, but no
effect on honeybee workers.
Tobacco budworms allowed to feed on bean leaves exposed to UV-B irradiation
did not show increased mortality. The eggs of pink bollworms, however,
irradiated on cotton leaves showed a redxiction in number that hatched.
Stab i_lity _of_ Ag_r icult ural Chem ic.a Is Under In c r_e_asฃd_ JlYzlLJLl? r' li
The Organic Chemical Synthesis Laboratory at Beltsville, Maryland,
constructed and successfully put into operation a "merry-go-round" type
photolysis apparatus for investigating the stability of pesticides and other
agricultural clie.mi.cals under increased UV-B irradiation. Such studies are
being conducted to determine the efficacy of various agricultural chemicals
under a high UV-B environment.
Photoclegradation of test compounds was obtained by exposing the samples
to 313 nm radiation in the UV-B region and quantum yields measured.
Preliminary studies with aqueous solutions of pesticides confirmed the
.. %
dependence, of quantum yield (the number of pesticide molecules consumed
per quantum of UV-B radiation absorbed) upon concentration.
-------
FINAL REPORT
EFFECTS OF UV-B RADIATION ON SELECTED LEAF PATHOGENIC
FUNGI AND ON DISEASE SEVERITY
H. R. Cams
J* H. Graham
S. J. Ravitz
Plant Stress Laboratory
Plant Physiology Institute
Beltsville Agricultural Research Center
Beltsvilleป Maryland 20705
EPA-IAG-D6-0168
Project Officer:
R. .J. McCracken
Agricultural Research, Science and Education Administration
U.S. Department of Agriculture
Washington, D.C. 20250
Prepared for
Environmental Protection Agency
EAGER Program
Washington, D.C. 20460
-------
ACKNOWLEDGMENT
We acknowledge the dedicated involvement of Ms. Sheryl Olsen who
provided valuable aid in conducting experiments, data collection, and
analyses throughout these investigations.
-------
ABSTRACT
The results of increased levels of UV-B irradiance on spore
germination indicate that although plant leaf pathogenic fungal species
vary considerably in sensitivity to UV-BS relatively high irradiance levels
are required to reduce germination percentage. Pigmented spores such as
Cladosporium, Stemphylium, and Alternaria were found to be more resistant
to increased UV-B irradiance than hyaline spores (Mycosphaerella,
Colletotrichum).
Disease severity of Col1etotrichum on cucumber was decreased with
increasing UV-B irradiance. A linear decrease in the percentage of leaf
area diseased with increased irradiance was found.
Level^of UV-B irradiance did not affect severity of disease incited
by Cladosporium.
There were no noticeable UV-B effects on either the Stemphylium
pathogen or the host, alfalfa. Alternaria, the tomato pathogen, reacted
similarly to Cladosporium on cucumber, disease severity being unaffected by
UV-B irradiance levels.
Recognizing that our results represent only a small sampling of leaf
disease organisms and plant disease-interaction experiments, they appear
\
to support the following: considerably higher levels of UV-B irradiance
than those expected from the projected ozone, depletion will be required to
adversely affect germination and growth of leaf pathogenic fungi; and, where
fungal germination and growth are affected, disease severity in the host
plant can be expected to be reduced as UV-B irradiance increases.
-------
INTRODUCTION
The impact of increased radiation in the 280-320 nra region (hereafter
referred to as UV-3) on overall plant growth, development, and critical
metabolic processes involved is a major concern in the short-term Biological
and Climatic Effects Research Program. Of equal 'concern is the possibility
that crop yield and quality may be affected indirectly by influencing plant
susceptibility to various plant pathogens and/or by affecting the pathogens
directly.
Numerous studies have been made on the influence of visible light on
sporulation of fungi; a limited number of studies have been conducted on
the effects on plant infection and disease severity. A few reports have
dealt with the effects of ultraviolet radiation; however, most researchers
used germicidal lamps (major energy in the 254 nm range) (UV-C). UV-C
from germicidal lamps enhanced sporulation of some fungi and inhibited or
retarded it in others. UV-C effects on pathogen spore germination, mycelial
growth and subsequent infection and disease severity have also been studied
and generally an adverse effect on the pathogen has been reported. In
addition, limited information is available on the effects of other UV
unspecified or poorly defined wavelengths on fungal behavior. Numerous
references are cited in reviews by Marsh et al. (1959) and by Leach (1971).
Our research was undertaken to provide preliminary information on the
effects of UV-B radiation on spore germination, mycelial growth, infectivity
and disease severity of three fungal leaf pathogens of cucumber (Cucumis
sativus L.)s one of tomato (Lycopersicon esculenturn Mill.), and two of
alfalfa (Medicago sativa L.). The pathogens selected were: Colletptrichum
lagenarium (Pass.) Ell. & Halst., causing cucumber anthracnose;
-------
Cladosporium cucumerinum Ell. & Arth., causing cucumber scab;
Mycosphaerella melonis (Pass.) Chiu & J. C. Walker, causing cucumber black
rot; Stemphylium botryosum Wallr., causing an alfalfa leafspot; Uromyces
striatus Schroet. var medicaginis (Pass.) Arth., causing alfalfa rust;
and Alternaria solani (Ell. & Mart.) L. R. Jones & Groot, causing tomato
early blight.
MATERIAL AND METHODS
UV-B Radiation - Measurements, Instrumentation, and Methodology
UV-B enhancement facilities were developed cooperatively with the
Agricultural Equipment Laboratory (AEL), Beltsville Agricultural Research
Center. UV-B enhancement was provided by the required assembly of
Westinghouse FS40 or FS20 fluorescent sunlamps, either filtered with 6-
hour-aged 5 mil cellulose acetate (CA) (plus UV-B) or 5 mil Mylar (minus
UV-B).
Spore germination and mycelial growth experiments were carried out in
temperature-controlled incubators equipped with one FS20 lamp each as the
UV-B source. Three incubators were equipped with one FS20 lamp as the only
radiation source and one incubator was additionally equipped with two
Westinghouse 14-watt cool white fluorescent lamps to provide visible energy.
Greenhouse and growth chamber experiments dealing with pathogen
infactivity and disease severity were carried out in a fiberglass green-
house or in plant growth chambers equipped with FS40 lamp assemblies
provided by AEL.
_!/ Mention of a trademark, proprietary product, or vendor does not
constitute a guarantee or warranty of the product by the U.S. Department
of Agriculture and does not imply its approval to the exclusion of other
products or vendors that may also be suitable.
-------
UV-B irradiance levels were determined for each plant or fungal
location in each experiment with either an Optronics Laboratories, Inc.
Model 725 UV-B Radiometer or an Instrumentation Research Laboratory (IRL)
UV-B Radiometer described in the IRL final report. Radiometer readings
were verified by spectral irradiance determinations (250-369 nm) with an
automated spectroradiometer as described in the IRL report at selected
locations in the experimental irradiation areas.
~2
Weighted irradiance levels are reported as mWra BUV, the biologically
effective UV derived from the AZ9 weighting function, and unweighted
__ f\
irradiance as mWm obtained by summing the measured or calculated values
2 _9
at each nanometer from 280-320 nm. Dividing mWm BUV by 3.06 (the mWm BUV
of control sunshine) provides the fraction of BUV received by each plant or
fungal location relative to that of one control sunshine.
Since all UV irradiation for experiments reported here was filtered
through cellulose acetate, BUV was limited to the UV-B region (280-320 nm).
For details concerning average control sunshine, spectral character-
istics of UV fluorescent lamps and filters, and the weighting function, see
the EAGER final reports of the Agricultural Equipment Laboratory, and
the Instrumentation Research Laboratory, Beltsville Agricultural
Research Center.
Spore ''Production
Sporulating cultures of Cladosporium c me urnerinurn and Mycosphaerella
melonis were grown on potato dextrose agar at room temperature (22-24ฐC).
Colletotrichum lagenarium was maintained under the same conditions on
V-8 juice agar. For Stemphylium botryosum, sporulation was induced by
placing V-8 juice agar cultures in a 22ฐC incubator fitted with four
-------
Westinghouse 14-watt cool white fluorescent lamps on a 12-12 hour
light-dark cycle. Alternaria solan! spores were obtained by growing
mycelia on lima bean extract agar. After 7 days of culture, the mycelia were
scraped with a scalpel in a sterile transfer chamber and the Petri dish lid was
removed. Spores were produced the following day,. Uredospores of Uromyces
striatus var. medicaginis, an obligate parasite, were obtained from infected
alfalfa plants.
Spore Germination
From a distilled water suspension of fungal spores, a drop was pipetted
onto 2 percent water agar in polyethylene plastic Petri dishes and allowed to
.dry. As the uredospores of Uromyces striatus do not readily suspend in
water, spores were dispersed onto the agar surface by dusting with a sterile
camel's hair brush. The lids of the dishes were removed and replaced by
filter squares of 5 mil CA, pre-solarized for 6 hours, or 5 mil Mylar as
required. Dishes were then transferred to incubators at predetermined
positions under the FS20 lamp such that they were subjected to weighted
irradiance levels of 6.25 mWni~2BUV (644 mWm~2), 6.69 mWm~2BUV
(691 mWnT2), 8.06 mWm~2BiJV (8.33 mWm~2) and 10.52 mWm~2BUV (1087 mWm"2).
Samples were irradiated for 6 hours, then left in the dark for 18 hours
after which microscopic counts of ,spore germination (200-500 spores per
dish) x^ere made. Each UV-B irradiance was duplicated within each of three
2 1
incubators. A fourth incubator was additionally supplied with 25 yEm s
of photosynthetically active radiation (PAR) provided by cool-white
fluorescent lamps. These lamps x^ere allowed to remain on during the 6-hour
UV-B irradiation and for an additional 6 hours thereafter.
Myce1ia1 Gr owt h
Fungal cultures were exposed to UV-B in the same incubators used for
spore germination. The 5 mil CA filter was placed as a collar around
-------
Che FS20 lamp, lids were removed and the Petri dish bottoms were enclosed
by UV-B transparent polyethylene bags to keep the agar from drying out
over the course of the experiments.
Samples were inoculated by placing a 7 nun diameter core of mycelia
upside down in the center of the agar plates which were then placed at the
predetermined positions in the chambers. Samples were irradiated daily for
6 hours until growth reached the perimeter of the Petri dish or sufficient
data points had been accumulated. Growth was determined by daily measuring
the diameter of the colonies.
Disease Development
Cucumbers
For epidemiological experiments with cucumbers, the UV-B sensitive
cultivar, Poinsett, was germinated in a synthetic soil mix of peat and
vermiculite (Jiffy Mix) in 12.5 cm pots in a fiberglass greenhouse, five
seedlings per pot. Temperatures ranged from 24-27ฐC during the day and
19-21ฐC at night. Plants were subjected from emergence to UV-B radiation
supplied by eight FS40 lamps, filtered by pre-solarized 5.mil CA. Filters
were changed every fourth day. Plants were irradiated with'UV-B between
the hours of 1000 and 1600 dailv for the duration of the. experiment. Figure
1 shows a typical experimental design used in the plastic greenhouse. '
As the cotyledons became fully expanded (5-7 days after seeding), the
seedlings were selected for uniformity and thinned to one per pot. Plants
were inoculated when the first leaf was fully expanded (11-14 days).
Colletotrichum lagenarium. Figures 2 and 3 diagram the experimental
design and provide the weighted and unweighted UV-B irradiances, respec-
tively, for each pot location.
-------
Figure 1. Typical experimental set-up used in UV-B enhancement
"studies in the greenhouse containing a four-fixture,
txtfo lamps per fixture, array of FS40 fluorescent sunlamps
(filtered with 5 mil CA). 'Poinsett cucumber plants infected
with Colletotrichura are shown. The typical chlorotic lesion
response induced by UV-B irradiation is evident. Disease
symptoms are visible at the bottom of the photograph on
plants receiving the lowest levels of UV-B irradiance.
-------
-
.
-------
4.6 6.1 7.7 9.2 10.1 10.1 10.1 9.5 8.3 7.0
5.8 4.3 3.1 2.1 1.5 1.2 1.0
CO
4'9 6'4 8.0 9.5 10.4 11.0 11.0 .10.1 9.2 7.7 6.1 4.3 3.4 2.1 1.5 1.2 1.0
5.2 6.7 8.6 10.4 11.3 11.9 11.6 11.0 9.8 8.0 6.4 4.6 3.4 2.1 1.9 1.2 1.0
5.5 7.0 9.2 11.0 11.9 12.6 12.2 11.3 10.1 8.3 6.4 4.6 3.4 2.5 1.8 1.2 1.0
5.2 7.0 9.2 10.7 11.6 12.2 11.9 11.3 10.1 8.3 6.4 4.6 3.4 2.5 1.8 1.2 1.0
**
5.2 6.7 8.6 9.8 11.0 11.6 11.6 10.7 9.5 8.0 6.1 4.6 3.4 2.1 1.5 1.2 1.0
4.6 6,1 8.0 9.2 10.1 10.4 10.4 9.8 8.6 7.0 5.8 4.3 3.1 2.1 1.5 1.2 1.0
4.3 5.8 I 7.0 8.3 9.5 9.8 9.8 9.2 8.0 6.7 I 5.2 3.7 2.8 1.8 1.5 1.2 1.0
Figure 2. Colletotrichum greenhouse experimental arrangement. Each number represents the biologically
effective UV-B irradiance in mWnf^jjUV at each plant canopy. Plants were 0.15 m apart.
Brackets indicate positions of fixtures, with the outer fixtures being 0,8 m, and the inner
fixtures 1.0 m from the top of the pot.
-------
474 631
791 948 1043 1043 1043 982 854 724 599 441 316 221 158 125 98
506 665 823 982 1072 1138 1138 1043 948 791 631 ' 441 349 221 158 .125 98
537 696 BbB 1072 1168 1233 1199 1138 1010 823 665 474 349 221 190 125 98
563 727 948 1138 123'J 1296 1263 1168 1043 854 665 474 349 253 190 125 98
4*
537 727 948 1105 1199 1263 1233 1168 1043 854 665 474 349 253 190 125 98
537 696 858 1010 1138 1199 1199 1105 982 823 631 474 349 221 158 125 98
474 631 823 948 1043 1072 1072 1010 858 724 599 441 316 221 158 125 98
441 599
727 854 982 1010 1010 948 823 696 537 380 284 190 158 125 98
Figure 3. Colie totriehum greenhouse experimental arrangement. Each number represents the total
unweighted UV-B irradiance in mWm~2 at each plant canopy. Plants were 0.15 m apart.
Brackets indicate positions of fixtures, with the outer fixtures being 0.8 m, and the
inner fixtures 1.0 m from the top of the pot.
-------
An inoculum, consisting of 30,000 spores/rni suspended in distilled
water, was prepared from 8-day old Colleto tr ichum cultures. Using an
electric sprayer, the leaves were covered with fine, droplets of inoculum.
Control plants were sprayed with distilled water. To attain the high
relative humidity necessary for infection, the sprayed plants were enclosed
within UV-B transmittable polyethylene bags for a 48-hour inoculation period.
Photographs, fresh weight, dry weight, area of first leaf, and percent
of first leaf diseased were recorded and used to determine UV-B-disease
interaction.
Cladosporium cycumerinum. The weighted and unweighted UV-B irradiances
for each pot location are shown in Figures 4 and 5, respectively. The
inoculum, containing 70,000 spores/ml, was applied as described for
Colletotrichum and the inoculated plants were sealed in polyethylene bags
for 48 hours. Plants xvere harvested 8 days after inoculation. Fresh weight,
dry weight, and area of first leaf were used to determine UV-B-disease
interaction as disease symptoms did not permit precise scoring.
Mycosphaerella melonis. The weighted and unweighted UV-B irradiances,
respectively,, for each pot location are shown in Figures 6 and 7. The
inoculum, containing 60,000 sporesVml, was applied as for Colleto trichum.
Similar data were also taken; however, as with* Cladosporium, disease
symptoms did not permit precise scoring.
Alfalfa
For the evaluation of the effect of UV-B on leaf rust of alfalfa, the
cultivar Arc was grown in a greenhouse for 5 weeks arid then transferred
to two plant growth chambers. Temperature in the chambers was 25-20ฐC
9 1
day-night with a relative humidity of 90 percent. An average of 200 pEm s
10
-------
4.9 6.7
5.2 7.7
6.1 8.3
6.7' 8.6
6.7 9.2
6.7 8.6
6.1 8.0
5.8 7.0
Figure 4.
9.2 10.
9_._5 11.
u ซ
10.7 12.
11.3 13.
11.6 13.
11.0 12.
7 11.9 12.9 12.9 11.9 10,4 8,9 7,0 5.2 3.7 2.8 1.8 1.2 0.9
9 13.2 14.4 14.4 13.5 11.'6 9.5 7.7 5.8 4.3 2.8. 1.8 1.2 0.9
9 14.4 15.0 15.3 14.4 12.6 10.4 8.0 6.1 4.6 3.1 2.1 0.9
5 15.0 -15.9 15.9 15.0 13.8 10.7 8.3 6.1 4.6 3.4 2.1 0.9
5 15.0 15.9 15.6 14.7 12.9 10.4 8.3 6.1 4.3 3.1 2.1 1.2 1.2
9 14.4 15.0 15.0 13.5 11.9 9.8 7.7 5.5 3.7 2.8 1.8 1.2 0.9
10.1 11.
9.5 11.
9 12.9 13.5 13.5 12.6 11.0 9,2 7.0 5,2 3.7 2.8 1.8 1.2 0.9
3 12.2 12.9 12.6
Cladosporium greenhousc0experiment arrangement. Each number represents the biologically effective
UV-B irradiance in mWm BUV at each plant canopy. Plants were 0.15 m apart. Brackets indicate
position of fixtures, with the outer fixture being 0.8 m, and the inner fixture 1.0 m from the
top of the pot.
-------
505
537
631
696
696
696
631
599
Figure
696
792
854
885
943
885
823
727^
5.
948
981
1105
1168
1199
1138
1043
981
1105
1233
1328
1389
1389
1328
1233
1168
Cladosnorium
1233
1358
1484
1547
1547
1484
1328
1263
1328
1484
1547
1641
1641*"
1547
' 1389
1328
greenhouse.
1328
1484
1579
1641
1611
1547
1389
1235
1233 1072 915
1389 1199 981
1484 1235 1072
1547 1421 1105
1515 1328 1072
1389 1233 1010
1235 1138 948
experiment arrangement.
727 537 380 284
792 599 441 284
823 631 474 316
854 631 474 349
854 631 441 316
792 568 380 284
727 537 380 284
Each number represents
190 125 95
190 125 95
234 95
234 95
234 125 125
190 125 95
190 125 95
the total unweightec
UV-B irradiance in mWm at each plant canopy. Plants were 0.15 m apart. Brackets indicate
position of fixtures, with the outer fixture being 0.8 m, and the inner fixture 1.0 m from
the top of the pot.
-------
4.9 6.4
5.2 7.0
5.8 7.7
6.4 8.7
6.4 8.7
8.7 10.
9.2 11.
i
9.8 11.
10.7 12.
10.4 12.
1
0
6
9
9
11.
12.
13.
14.
14.
3
2
2
.**
4 '
4
11.
12,
14.
15.
15.
9
9
1
0
0
11.
12.
14.
15.
15.
9
9
1
0
0
11.
12.
13.
14.
14.
6
2
5
4
4
10.
11.
11.
12.
12.
4 7.7
0 8.0
9 8.7
6 10.1
6 10.4
6.1 4,
6,4 4,
7.0 6.
8.3 5.
8.3 5.
6
9
1
8
8
3.7
3.7
4.0
4.3
4.3
2.8
2.8
3.1
3.1
3.1
1.
1.
2.
2.
2.
8 1.2
8 1.5
1 1.5
1 1.5
1 1.5
.9
.9
1.2
1.2
.9
J6.1 8.0 10.1 12.6 14.1 14.7 14.7 13.8 11.9 9.8 8.0 5,8 4.0 2.8 1.8 1.2
5.8 7.7 9.5 11.6 12.9 13.5 13.5 12.6 11.3 9.2 7.3 5.2 3,7 2.8 1,8 1.2
5.2 6.7 8.1 10.4 11.6 11.9 11.9 11.3 9.8 8.3 6.7 4.6 3.4 2.1 1.8 1.2 .9
Figure 6. Mycosphaerella greenhouse experiment arrangement. Each number represents the biologically
effective UV-B irradiance in mWm~2BUV at each plant canopy. Plants v;ere 0.15 m apart. Brackets
indicate position of fixtures, with the outer fixture being 0.8 m, and the inner fixture 1.0 m
from the top of the pot.
-------
506 661 900 1043 1166 1227 1227 1197 1072 796 628 A66 381 291 186 124 93
537 722 949 1134 1259 1331 1331 1259 1134 827 661 506 381 291. 186 154 93
598 796 910 1197 1520 1455 1455 1394 1227 900 722 628 414 320 217 154 124
661 900 1104 1331 1485 1547 1547 1485 1301 1043 858 .598 444 320 217 154 124
ซ
661 900 1072 1331 1485 1547 1547 1485 1301 1072 858 598 444 320 217 154 93
628 827 1043 1301 1455 1518 1518 1425 1227 910 827 598 414 291 186 124 93
598 796 981 1197 1331 1394 1394 1301 1166 949 753 537 381 291 186 124 93
537 692
835 1072 1197 1227 1227 1166 910 858 691 466 352 217 186 124 93
Figure 7. Mycosphaerella greenhouse experiment arrangement. Each number represents the total
unweighted UV-B irradiance in mWm~ at each plant canopy. Plants were 0.15 m apart.
Brackets indicate position of fixtures, with the outer fixture being 0.8 m, and the
inner fixture 1.0 m from the top of the pot.
-------
visible radiation was provided by sixteen 165-watt cool white fluorescent .Tamps
and twelve 50-watt incandescent bulbs with a photoperiod of 16-hour day
8-hour night. The plants were allowed to acclimate to chamber conditions
for 5 days before being irradiated with UV-B. UV-B radiation was provided
by two FS40 Westinghouse sunlamps (with no reflector) filtered by 5 mil CA
that had been pre-solarized for 6 hours. Filters were changed every fourth
day. Figures 8 and 9 diagram the plant arrangement within the chambers
and show the weighted and the unweighted UV-B irradiances, respectively, at
the canopy height of each plant. Plants were clipped during the experiment
to maintain a distance of 0.37 m from the fixture.
Uromyces striatus. The plants were inoculated with spores of the rust
fungus after 7 days of UV-B irradiation. Inoculum was prepared by scraping
the spores from 10-day-old _!5. botryosum agar cultures, suspending them in
distilled water and filtering the mixture through cheesecloth to remove
mycelial fragments. The resulting spore suspension contained 10,000
spores/ml and was applied to the alfalfa leaves in fine droplets by means
of a chromatography sprayer. Control plants were sprayed with distilled
water only. All plants were then covered for 24 hours with UV-B transmit-
table polyethylene bags to achieve maximum humidity necessary for good
infection. Five days after the end of the inoculation period, all leaflets
v
in the upper 4 centimeters of the plant were scored for type of lesion as
fellows: 1 = pinhead size brown flecks; 2 = lesion approximately 1 mm in
diameter with a brown margin and tan center; 3 = lesion approximately two
to three times larger than #2 type and often with obvious yellow halo out-
side of the brown margin; and 4 = a large blighted area most often found
on leaflet margin.
15
-------
7.3 8.6 8.6 8.6 8.3 ' 7.0
2,1 3.4 5.8
9.1 10,4 10.7 11.0 10.1 . 8.6
2.1 3.4 ! 6.1
8.6 10.1 10.7 10.4 10.1 8.6
2.1 3.4 5.8
6.7 7.7 8.3 8.0 7.7 7.0
. 0.9
! 0.9
*
ป
. 0.9
. 0.9
Figure 8.
Growth chamber experimental arrangement. Each number represents the biologically
effective UV-B irradiance in rr.Wm BUV at each plant canopy (0.37 m from the
fixture). Plants were 0.15 m apart. Brackets indicate the position of the
fixture. The dotted line represents a 5 mil Mylar barrier.
-------
221
221
221
349 599
349
611
349 599
759 885 885 885 854 727
948 1072 1105 1138 1043 885
885 1043 1105 1072 1043 885
696 792 854 823 792 727
95
95
95
95
Figure 9.- Growth chamber experimental arrangement. Each number represents the total
unweighted UV-B irradiance in mWm at each plant canopy (0.37 m from the
fixture). Plants were 0.15 m apart. Brackets indicate the position of
the fixture. The dotted line represents a 5 mil Mylar barrier.
-------
Tomato
The cultivar Chef was germinated and grown in the same plant growth
chambers used for the alfalfa experiments except that the FS40 lamps were
repositioned to adjust UV-B irradiance levels and temperatures were
maintained at 26ฐC .day - 20ฐC night. Figures 10 and 11 diagram the plant
arrangement within the chamber and list the weighted and unweighted
.irradiances, respectively.
Alternaria solani. Twenty-two days after seeding when the third leaf
was well expanded, buds were pinched out; and four days later the plants
were inoculated as follows: The spore suspension was prepared by placing
two spore mats and 100 ml distilled water in a blender for 15 seconds and. then
h
.filtering the resulting suspension through cheesecloth to remove mycelia
and agar fragments. The suspension was applied in a fine mist to the second
and third.leaves by means of a chromatography sprayer. Relative humidity in
the chamber was maintained at 90-100 percent for 24 hours by a chamber
humidity regulator .and the placement of a polyethylene canopy over the plants.
The second and third leaves were harvested for dry weights 48 hours after
inoculation when blighting became severe.
For analyses of plant-disease response as a function of UV-B, in the
.above experiments, .data from .individual plants xsrere combined into irradiance
-? v
groups of increasing 1.5 mWm BUV and subjected to .analyses of variance.
If P= 0.05 or less, the data were further analyzed using linear regression.
RESULTS AND DISCUSSION
Spore Germination
The results of increased levels of UV-B irradiance on spore germina-
tion .are shown in Table 1. They indicate that even in the more sensitive
species, high UV-B irradiance levels are required to reduce germination
percentage; generally, more than double the UV-B of one control sunshine
18
-------
1.8
1.5
1.5
1.2
1.5
1.5
2.5
2.8
2.8
2,8
2,8
2,5
3.7
4.6
4.9
4.9
4.6
3.7
4.6
5.8
6.7
7.0
6.4
5.2
5.5
7.0
8.3
8.3
7.7
6.1
6.1
7.7
8.9
9.2
8.3
6.7
6.7
8.3
9.5
9.5
8.9
7.0
7.0
8.6
9.8
9.8
8.9
7.3
7.0
8.6
10.1
10.1
9.2
7.0
6.7
8.3
9.5
9.5
8.6
6.7
5.5
7.3
8.3
8.3
7.3
5.8
4.6
5.5
6.4
6.1
5.5
4.3
3.1
3.4
3,7
3.7
3.7
3.1
1.8
1.8
1.8
1.8
1.8
1.8
Figure 10, Alternaria growth chamber experimental arrangement. Each number
represents the biologically effective UV-B irradiance in mWra~^BUV
at each plant canopy.
-------
190 253....38.0 474 568 611 696 727 727 696 568 474 316 190
159 286 474 597 727 792 854 885 885 854 759 568 349 190
159 286 506 696 854 915 981 1010 1043 981 854 665 380 190
O
125 286
506 727 854 956 981 1010 1043 981 854 611
380 190
159 286 474 665 792 854 915 915 956 885 759 568 380 190
159 253 380 536 611 696 727 759 727 696 597 441 316 190
Figure 11. Alternaria growth chamber experimental arrangement. Each number represents
the total unweighted UV-B irradiance in mUm at each plant canopy.
-------
was required before a reduction in spore germination was noted. In the
more resistant species, more than three times the level of control
sunshine UV-B was needed to inhibit spore germination.
For Colletotrichum, the most sensitive species, spore germination was
reduced by less than two times control sunshine as indicated by comparison
of the Mylar-filtered controls and the lowest irradiance level used.
Resistance to UV-B appeared to be correlated with spore pigmentation.
Spores of the resistant species Cladosporium, Stemphyliura, Uromyces, and
Alternaria are all darkly pigmented, whereas, spores of Mycosphaerella and
Colletotrichum are hyaline. Our results suggest that pigmentation provided
protection from damage by UV-B.
In the incubator supplied with PAR in addition to UV-B, germination
values were consistently, but only slightly, higher than those observed in
the incubators irradiated by UV-B only, with the most notable increase in the
sensitive Colletotrichum spores. Furthermore, we observed that germ tube length
ซ>
appeared to be considerably increased in nearly all tests in the presence
of PAR. This observation is consistent with other BACER research which provides
evidence for the existence of a photorepair or photoprotection mechanism.
Linear regression analyses indicate that within the range 'of irradiance
levels tested, there is a significant correlation between reduction in
germination and increased UV-B irradiance in the susceptible species.
Mycelial Growth
A measure of the mycelial growth rate under identical UV-B irradiances
and environment used for the spore germination experiments is presented in
Table 2. Growth rate is expressed as increase in colony diameter with time.
In contrast to the spore germination, increase in colony diameter, (as
expressed in terms of percentage of the Mylar control) at the end of the
21
-------
Table 1. Influence of UV-R irradiation on spore germination of six pathogens.
Spores were irradiated in Petri dis-hes in an incubator at 22ฐC under
an FS20 fluorescent sunl'amp filtered with either 5 mil cellulose
acetate or 5 mil Mylar.
Mylar-5 mil
Disease organism <
Colletotrichum lagenarium
Test A
B
Mycosphaerella melonis
Test A
E
C
D
E
Alternaria solani
Test A
B
Stemphylium botryosum
Test A
B
Cladosporium cucumerinum
' '': -' Test A '" "' '- ""'"
B
Uromyces striatus
Test A
B
0.25
Mean
17.82
49.29
52.46
50.21
41.30
33.06
63.08
98.17
98.88
98.03
93.46
93.69
94.73
86.85
90.24
mWm
2BUV
Cellulose acetate-5 mil
6.26
percent
12.70
10.31
57.75
45.48
47.85
29.10
60.04
98.58
99.01
97.44
93.48
90.53 '
90.93
86.70
89.62
6.69
8.06 10
.52
germination
17.79
8.20
57.34
42.34
46.34
28.49
57.41
98.49
99.38
97.35
93.48
89.02
t89.65
85.54
89.90
5.13
2.80
47.85
30.95
37.96
24.59
47.28
98.72
99.14
95.49
93.04
86.37' '
87.80
86.25
89.03
2.03
1.22
35.44
22.94
26.62
19.62
33.90
98.75
99 . 01
94.09
93.29
81.33
83.78
86.10
89.88
r2 '
0.691
0.851
0.945
0.934
0.924
0.885
0.935
0.942 '
0.909
22
-------
Table 2. Influence of UV-B irradiation on mycelial growth of five pathogens.
Mycelia were irradiated in Petri dishes in an incubator at 22 C
under an FS20 fluorescent sunlamp filtered with either 5 mil
cellulose acetate or 5 mil Mylar.
-2
rnWm BUV
Mylar- 5 mil Cellulose acetate-5
Disease organism
No.
days
< 0.25
irradiated
Colletotrichum lagenarium
Percent of Mylar Control
Mycosphaerella melonis
Percent of Mylar Control
Alternaria solani
Percent of Mylar Control
Stemphylium botryosum
Percent of Mylar Control
CladosDorium cucumerinum
Percent of Mylar Control
4
6
8
2
3
4
2
3
4
5
6
3
5
7
11 V
3
5
7
11
22.8
34.0
40.5
45.0
64.0
82.3
29.8
40.0
50.3
60.5
71.2
28.7
41.5
55.7
. 78.0
*
18.2
32.9
47.5
76.0
6.25
Colony
17.5
27.0
35.3
87
44.0
62.0
79.7
97
29.3
39.0
49.7
60.7
70.7
99
14.8
23.2
41.0
72.5
93
12.7
25.0
37.2
59.5
78
6.69
diameter
15.5
26.0
35.0
86
44.5
62.7
80.8
98
29.3
38.7
49.3
59.5
70.2
99
14.0
21.7
39.3
. .68.3
88
14.0
24.8
36.0
58.5
77
8.06
- mm
14.0
26.3
35.8
88
43.2
60.7
78.3
95
28.8
38.8
48. ,8
59.0
69.7
98
13.5
22.3
37.7
.68.3
88
13.8
23.7
34.2
57.7
76
mil
10.52
12.8
24.0
29.3
72
42.7
60.2
76.2
93
28.7
37.5
48.5
59.3
69.7
98
13.3
18.3
33.3
65.0.
83
13.2
23.3
34.3
55.5
73
23
-------
growth period did not differ greatly from colonies receiving no UV-B.
As with spore germination, Colletotrichum appeared to be most sensitive to
relatively high UV-B irradiance levels used. Cladospgrium and Stemphylium
were intermediate in response, while Alternaria and Mycosphaerella showed
little .if any reduction at the highest UV-B Irradiance level used. However,
with all species, mycelial density was visibly reduced when compared to the
Mylar controls. Attempts to obtain dry weights of colonies revealed
. differences between CA-filtered and Mylar-filtered colonies, but the method
used was insufficiently sensitive to distinguish between CA-treatments. We
* conclude that, as with spore germination, relatively high UV-B irradiance
? is required before, the growth of these fungi is impaired.
i Cucumber
{ Growth and disease responses of cucumber to UV-B radiation are shown as
* 'follows: Colletotrichum, Figures 12 through 17; Mycosphaerella, Figures 18
' through .20; and Clad o s porium, Figures 21 through 23. Growth responses of the
f uninoculated control plants, as measured by fresh weight, dry weight, and
I
: area of first leaf, responded similarly in all cucumber disease experiments,
i showing increased repression with each increase in UV-B irradiation level
r. applied.
\
I Disease severity of Colletotrichum and Myco sphaerella on cucumber
j decreased with increasing UV-B irradiances. This is shown graphically for
'' Colletotrichum (Figure 12) for which percentage of diseased leaf area is plotted
| against UV-B irradiance levels. Pictorial representation of the response is
if presented in Figures 16 and 17. Figure 16 depicts the disease response in -the
\ absence of UV-B enhancement (Mylar control)-' In Figure 17, disease
v . - ' .
( response is compared to uninoculated/controls subjected to high and low
i
'. levels of UV-B irradiances. The differences in disease response are obvious.
24
-------
Leaf Area. -Diseased, %
.30
5
Weighted mWm
-2
Figure ..12, ..The effect of .increased UV-B .irradiance on the percent
'-'-. '- -' '.diseased 'area of cucumber leaves infected with Colletotrichum
'.lagenarium. .r _= 0.81, ^standard error := _2._2.
25
-------
Fresh Weight,
19r
Control
5 10
Weighted
*
Figure 13. The effect of increased UV-B irradiance on fresh weight of
Colletotrichum lagenarium infected (r2 = 0.77, standard
'error = 0.63) and rtoninfected (r2 = 0.03, standard error =
0.A3) cucumber plants.
26
-------
Dry Weighting
75ฎ
5OO
Control
Inoculated
5 . 1O
Weighted mWrn"2
.Figure 14. The effect of UV^B irradiance on dry weight of
Colletotrichum Jsagenarium infected (r^ = 0.32,
standard error = A0.6) and noninfected (r^ = 0.79,
standard error = 64.43) cucuciber plants.
27
-------
First Leaf Area,
Control
o
Weighted mWm
-2
Figure 15. The effect of UV-B irracliance on area of first leaf of
V.O
Colletotrichum lagenarium infected (rz = 0.01, standard
error = 2.36) and noninfected (r^ = 0.80, standard error
= 5.74) cucumber plants.
28
-------
Figure 16; Disease response to inoculation of cucumber with Coiletotrichum spores in the
absence of UV-B (Mylar filtered); left to right, uninoculated and inoculated
plants.
-------
.
XA
u
,
'
.
5
30
;
-------
-Figure :17. .'Disease > response ; to 'inoculation' 'of'cucumber'plants' with Colletotric'hurn spores
r sub j ec te'd ; to '-high-arid'low ; level'UV-B'ir radiance. 'Left'to right: high UV-B
juninoeulate'd; I high1' UV-B 'inoculated; 'low'UV-B'uni'riocul'ated; and 'low UV-B inoculated.
-------
r
...
M -ป. -
--
- .
~
.
- -
- - . -
.
-
,' :, ...
v . '-
,
,
**.*ป*>
-------
.Figure 18. The effect of increased UV-B irradiance on fresh weight
of Mycosphaerella melonis, infected (r- = 0,43., .
errorr= 0.19) and noninfected (r'2 = 0.91* standard error
- 0.11) cucumber plants.
Fresh Weight, g
Control
Weighted mWm'
33
-------
Dry Weightjin|
25%-
15ฉ
5
Fieu-r-e 19. The. ef-f-ec-t of- increased- UV-3- irradiance on dry weight of
Mycosphaerella melonis infected (r2 = 0.24, standard
er5oi"' 2_0';i)"and noninfected (r2. =r. 0.89, standard
er-r-qr- =?, 10.95. cuc_umber. plants.._.
34
-------
Leaf Area, cm'
60r
40
C ontrol
Inoculated
5 1O
Weighted mWm~2
15
Figure 20. The effect of increased UV-B irradiance on area of first
leaf of Mycosphaerella melonis infected (r = 0.91, standard
error = 2.61) and noninfected (r = 0.95, standard error =
2.31) cucumber plants.
35
-------
Fresh Weight, g
20 r
16
12
8
O
-------
^;*~^A*^^ A^^A
Dry Weight, g
2.ฉf
1.5-
0 Control
Weighted mWm
Figure 22. The effect of increased UV-B irradiance on dry weight of
Cladosporium cucumerinum infected (r^ = 0.89, standard
error = 0.07) andvnoninfected (r^ = 0.94, standard
error = 0.07) cucumber plants.
37
-------
Leaf Area,
Control
5 T&
Weighted mWm~2
Figure 23. The effect of increased UV-B irradiance on area of first
leaf of Cladosporium cucumerinum infected (r = 0.92,
standard error = 4.0) and noninfected (r^ = 0.92, standard
error = 4.06) cucumber plants'.
38
-------
The decrease in disease infectivity and severity are reflected in
area of first leaf and fresh and dry weight of both Colletotrichum and
Mycosphaerella. With both pathogens, the measured growth responses of
inoculated and noninoculated plants approach unity in the region of 15
_o
mWra BUV irradiance. Colletotrichum and Mycosphaerella both have hyaline
spores. These disease-UV-B interactions are consistent with the spore
germination and mycelial growth data presented earlier. They suggest that
disease response to these organisms is due to the direct effect of UV-B
irradiance on the pathogen, although increased resistance to the pathogens
by the irradiated cucumber plants could also be a factor.
Cladosporium, possessing a pigmented spore, showed no such pathogen-
UV-B interaction. Observed disease symptomology and measurement of first
leaf area, fresh and dry weight, shox^ed a uniformly reduced growth of
inoculated plants regardless of UV-B irradiance levels (Figures 21 through
23); again, this observation is consistent with the effects noted on spore
germination and mycelial growth.
Alfalfa
At the UV-B irradiances used, there were no noticeable UV-B effects
on either the Stemphylium pathogen or the host plant, alfalfa. Stemphylium
spores are also pigmented. Table 3 shows the results of these experiments
undertaken in plant growth chambers and indicates that similar lesion types
occurred at all UV-B irradiance levels. The differences between experiments
were due to harvesting the first experiment 3 days later than the second.
The results indicate that the lesions progressed equally over time under all
UV-B levels. Again, this response is consistent with that of other pathogens
having pigmented spores. Using Uromyces in an experiment with alfalfa grown
in a fiberglass greenhouse, similar results were obtained and supported our
-------
''"'fa'ble'-3. nThe"'reff e'ctrroฃ ''-UV-B^Dn' 'disease''development-"of ifche- alfalfa1:pathogen>:':'Stemphyp.ium..bQtryosum
v Wallr
Ml :Biolo'gically
effective
'UV-B
T;''mUm~2BUV
;-2.i4
"3'.67
'-5l'!20
6.73
8.26
9.76
." : torv A'r cf ' tultivar
:;'No.
1'leaifle'e's/'scoye
'''"48
''';48
/!;'48
108
240
96
:ofi alfalfa.
Experiment
''Growth
' 'chamber. A
:' 2
' 2
/ 2
1
2
2
1
Growth
chamber'.
l 1
: 1
., 2
2
2
1
i 'Lesion score
Growth
B . : chamber
i 1
2
; 1
1
1
1
Experiment. 2
, Growth
A . chamber. B
i 1
; 2
-, 1
1
1
1
-------
belief that the progression of rust on alfalfa is not likely to be affected
by enhanced UV-B, except perhaps at very high irradiance levels.
Tomato
Alternaria, a pigmented spore, reacted similarly to Cladosporium on
cucumber, disease severity being unaffected by UV-B irradiance levels
(Table 4). The differences in dry weight between UV-B irradiances appear
to be more closely correlated with PAR than with UV-B radiation.
SUMMARY
Recognizing that our results represent only a small sampling of leaf
disease organisms and plant disease interactions, they appear to support
the following: ' 1) considerably higher levels of UV-B irradiances than those
expected from projected decrease in ozone that might be caused by chloro-
fluoromethanes will be required to adversely affect germination and growth
of leaf pathogenic fungi, and 2) where fungal germination and growth are
affected, disease severity in the host plant can be expected to be reduced
as UV-B irradiance increases.
41
-------
Table 4. The effect of UV-B on disease development of the tomato pathogen,
Alternaria solani (Ell. & Mart.) L. R. Jones and Groot.
UV-B
weighted
inWnT2BUV
2.5
4.0
5.5
7.0
8.5
10.0
Inoculated
148.5
168.1
190.6
172.6
198.1
131.8
Mean 168.3
r2 < 0.01
Dry weight - mg
yninoculated
177,3
1?2,0
. 211,4
202.9
195,6
162,5
!?0f3
QtP5
-------
REFERENCES
Leech, Charles M. A practical guide, to the effects of visible and
ultraviolet light on fungi. Methods in Microbiology. C. Booth, Ed.,
Chap. XXIII. pp 609-664. Academic Presss London and New York, 1971.
Marsh, Paul B., E. E. Taylor, and L. M. Bassler. A guide to the literature
on certain effects of light on fungi: Reproduction, morphology,
pigmentation, and phototropic phenomena. Plant Disease Reporter.
Supplement 261. U.S. Department of Agriculture, pp 251-312, 1959.
43
-------
FILL-
FINAL REPORT
THE EFFECT OF ULTRAVIOLET (UV-B) RADIATION ON
ENGLEMANN SPRUCE AND LODGEPOLE PINE SEEDLINGS
M. R. Kaufraann
Rocky Mountain Forest and Range Experiment Station
U.S. Forest Service
Fort Collins, Colorado 80521
EPA-IAG-D6-0168
Project Officer:
R. J. McCracken
Agricultural Research, Science and Education Administration
U.S. Department of Agriculture
Washington, D.C. 20250
Prepared for
Environmental Protection Agency
EAGER Program
Washington, D.C. 20460
-------
ABSTRACT
Engelmann spruce and lodgepole pine are conifers found at high elevations
(2,700 to 3,300 m) in the central Rocky Mountains. UV-B enhancement and ex-
clusion studies were performed at about 2,980 ra on transplanted seedlings.
Enhancement studies were performed using standardized light banks consisting
of two FS-40 sunlamps in each of two fixtures 60 cm apart and 110 cm above the
seedlings. Cellulose acetate and Mylar filters were used along with, an un-
treated control to provide appropriate UV-B treatments, with seedlings arrayed
to vary the amount of supplementary radiation. Treatments extended for 67
days, with supplementary radiation for a cumulative period of 400. hours.
Exclusion studies were performed to compare the effects of UV-B in natural
sunlight. Seedlings were placed under filters of cellulose acetate or Mylar,
under lath shade, or in the open to provide different UV-B treatments.
Careful visual observations of the seedlings by several scientists in-
dicated no symptoms (color, necrosis, growth form, etc.) for any of the
treatments, either during the study period or in the subsequent 2-1/2 months
before the seedlings were covered by snow. Analyses of growth QLength. of
terminal leader, number and length of lateral branches) indicated no -major
effect of treatments, either in the enhancement or in the exclusion study.
Importantly, however, in the natural environment transplanted seedlings
of Engelmann spruce do not show symptoms of solar radiation damage until the
summer following exposure. Consequently, seedlings are being kept for con-
tinued observation to determine whether differences among treatments may
appear during the second growing season, perhaps as a result of effects on
food reserves and ability to x^ithstand the harsh winter environment of high.
elevation areas. *
This research was done by the U.S. Forest Service Rocky Mountain Forest
and Range Experiment Station, Fort Collins, CO, under a cooperative agreement
with the Agricultural Research Service, Beltsville, Mt>. and under an agreement
with ARS and the Environmental Protection Agency. This report covers the
period of Oct. 1, 1976 to Sept. 30, 1977. Except for continuing experimental
observations of plants, this work was completed on Dec. 15, 1977.
-------
CONTENTS
: Abstract , , . , . . i
Figures ,,,.ซ,..., ill
; Tables , , , , , , . iv
Introduction ,.,.,,..,,..,,,., 1
Conclusions , i , 2
Recommendations .,.....,..,,.,, 3
i Text 4
General Objectives 4
I Experimental Procedures 4
[ Results 6
' Discussion 14
I References 15
i: Appendix 16
ii
-------
FIGURES
Number Page
1 Distribution of seedlings among 7 radiation levels
beneath the lamp fixtures ..... .... 5
2 Effect of supplemental radiation levels on vegetative
growth of Engelmann spruce 12
3 Effect of supplemental radiation levels on vegetative
growth'of lodgepole pine 13
111
-------
f
[ TABLES
Number Page
| 1 UV-B Radiation Levels at Different Positions Beneath.
I Lamps, Measured with IRL Spec D Spectroradioroeter 7
ป
,l 2 Effect of Natural Sunlight, Shade, and UV-B on Terminal
i Bud Failure and Seedling Mortality in Engelmann
* Spruce and Lodgepole Pine (.Exclusion Study) 8
Effect of UV-B Radiation Levels on Terminal Bud
Failure in Engelmann Spruce and Lodgepole Pine
(Enhancement Study) 9,
Effect of UV-B Radiation Levels on Mortality in
Engelmann Spruce and Lodgepole Pine (Enhancement
Study) 10
Effect of Natural Sunlight, Shade, and UV-B on Vegetative
Growth of Engelmann Spruce and Lodgepole Pine
(Exclusion Study) , 11
Effect of UV-B Radiation Levels on Vegetative Growth of
Engelmann Spruce (Enhancement Study) ,..,,,.,,,., 16
Effect of UV-B Radiation Levels on Vegetative Growth, of
Lodgepole Pine (Enhancement Study) ,...,....,.,, 17
iv
-------
INTRODUCTION
The possibility that the intensity of ultraviolet light (UV-B, 280-320 nm)'
received at the earth's surface will increase is of considerable interest
in the high-elevation forest zone of the central Rocky Mountains, Ronco
(1970 a and b) concluded that mortality of open-grown Engelmann spruce seed-
lings was related to intense solar radiation, whereas seedlings of lodgepole
pine were not adversely affected by full sunlight. The possibility that
solar radiation damage of spruce is caused in part by high levels of UV-B
stimulated interest in evaluating the effects of increased levels of UV-B
on growth and development of these species.
The research reported here was designed to determine the physiological
effects of enhanced UV-B radiation on Engelmann spruce and lodgepole pine and
to establish UV-B tolerance levels for these species in high-elevation forest
ecosystems.
-------
CONCLUSIONS
Observations were made of the effects of UV-B on vegetative growth,
foliage color, and morphological development of new foliage of Engelmann
spruce and lodgepole pine seedlings during and after a treatment period
extending 67 days (400 total hours of supplemental UV-B radiation).
No significant effects of UV-B enhancement or exclusion were observed
during the treatment period or in the subsequent 2-1/2 months before the
seedlings were covered by snow. It must be emphasized, however, that in the
natural environment transplanted seedlings of Engelmann spruce do not show
symptoms of solar radiation damage until after the first winter.
Consequently, it is premature to reach conclusions about the impact on
high elevation conifers of UV-B enhancement or exclusion after a single
growing season.
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RECOMMENDATIONS
No effects of UV-B enhancement or exclusion were observed during a
single growing season.
From an experimental standpoint, it is recommended that observations
on the treated seedlings continue into the second growing season and that
future studies on tree species be long enough to accommodate the perennial,
long-term nature of growth and development.
In the context of environmental impact of UV-B, no assessment or
recommendation can be made at this time regarding tolerance levels for
Engelmann spruce and lodgepole pine.
-------
TEXT
GENERAL OBJECTIVES
Because of thinner atmosphere, solar radiation is attenuated less at
high elevations than at low elevations. In most portions of the solar
spectrum, including UV-B, intensities are higher at high elevations than
near sea level. Thus the study of growth and development of high, elevation
conifers was designed to include exclusion of natural UV-B as well as en-
hancement of UV-B, as might occur through ozone depletion in the stratosphere,
The field research was divided into two parts. The. exclusion study was
conducted to determine if the presence or absence of natural UV-B or a general
reduction across the entire solar spectrum had an effect on physiology and
vegetative growth of conifer seedlings. Simultaneously, an enhancement study
was conducted to determine seedling response to various levels of enhanced
UV-B which might result from ozone depletion,
EXPERIMENTAL PROCEDURES
Experiments were conducted near the Elk Camp Restaurant in the Snowmass
Ski Area, Snowmass Village, Co. at an elevation of 2,980 m. Experiments were
performed using 3-year old Engelmann spruce (Picea engelmannii Parry) and
2-year old lodgepole pine (Pinus contorta Dougl.) seedlings. The seedlings
were grown at the U.S. Forest Service Mt. Sopris Nursery located at Carbondale,
CO, about 20 km northwest from the study site and at an elevation of about
2,800 m. Seedlings were potted June 13-15 and exposed to selected treatments.
\
In the exclusion study, groups of 25 seedlings of each species were
exposed to one of four treatments: (1) natural sunlight control; (2) lath
shade providing 50 percent interception of noon-day sun; (3) natural UV-B,
using a 5 mil cellulose acetate filter; and (4) excluded UV-B, using a 5 mil
Mylar filter. Treatments were replicated, and 200 seedlings of each species
were used in the study.
The enhancement study utilized natural sunlight plus supplemental radiation
from FS-40 lamps. Seedlings were exposed to three treatments: (1) natural
sunlight control; (2) plus UV-B, consisting of natural sunlight augmented by
FS-40 lamps filtered with 5 mil cellulose acetate; and (3) natural UV-B,
consisting of natural sunlight and FS-40 lamps filtered with 5 mil Mylar.
Both cellulose acetate and Mylar remove wavelengths below UV-B; cellulose
acetate passes UV-B'and Mylar removes UV-B. Lamp pairs were located 60 cm
apart and 110 cm above the seedlings. Lamps were operated for 6 hr. per day
for 67 days of treatment. Seedlings were distributed beneath the lamps to
provide seven levels (A through G of UV-B intensity (Fig. 1)).. The positions
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were selected to provide supplemental UV-B radiation in the following approxi-
mate relationship: A:B:C:D:E:F:G = 0.2:0.4:0.6:0.8:1.0;1.3:1.6. Nighttime
measurements with an IRL Spec D spectroradiometer at each, seedling indicated
these relative values to be substantially correct for the seven radiation
levels (Table 1). Thirty-six seedlings of each species were used in each of
the three treatments, and all treatments were replicated. Six seedlings were
subjected to each of the five lower supplemental UV-B levels (A through E) and
three to each of the two highest levels (F and G). A total of 216 seedlings
of each species were used in the enhancement study.
Throughout the treatment period, observations were made of the breaking
of dormancy of terminal and lateral buds. Particular attention was paid to
visual observation of color, deformity, or dwarfing of new foliage. On
Aug. 22-24, 1977, after 67 days of treatment, measurements were made of mor-
tality, length of terminal leader (.if present) , and number and lengths of all
lateral branches. No records were made of color or of needle length., since
variation within treatments obviously greatly exceeded variation among
treatments.
. On Aug. 30, 1977, seedlings were moved to the Fraser Experimental Forest,
CO. (elevation 2,740 m). Visual observations were continued until mid-November
when snow covered the seedlings. Additional observations of mortality and
color and condition of 1977 and 1978 growing season foliage will be made
during 1978.
RESULTS
During the 67-day treatment period, no treatment differences of any kind .
were observed in rate of dormancy break or in appearance of foliage from the
new flush or from the previous growing season. Failure of the terminal bud
of Engelmann spruce to break dormancy ranged from 30 to 52 percent in the
exclusion study (Table 2) and from 25 to 83 percent in the enhancement study
(Table 3). In lodgepole pine, terminal bud failure was 0 to 8 percent in the
exclusion study and 0 to 33 percent in the enhancement study. No significant
effects were found among treatments or supplemental radiation levels in either
study. The high bud failure in spruce is not important, since a lateral bud
on the terminal leader quickly assumes dominance. Seedling mortality ranged
up to 8 percent for spruce and to 17 percent for pine in the two studies
(Tables 2 and 4), but again, no treatment or radiation level effects were
significant.
Vegetative growth data for the exclusion study are summarized in Table
5. No significant treatment effects were observed in terminal leader
length (measured on plants whose terminal bud broke dormancy), number of
branches, and total and mean branch lengths. Vegetative growth data for
the enhancement study are given in Figures 2 and 3 (means with, standard
deviations and statistical evaluations appear in Tables 6 and 7 in the.
Appendix). Significant treatment effects were observed on total branch
length in Engelmann spruce (Fig. 2C) and on number of branches in lodgepole
pine (Fig. 3B). Supplemental radiation effects were significant only for
mean branch length of Engelmann spruce (.Fig. 2D) ; however this effect was
also observed in the untreated control plants and is probably due to random
experimental error. Treatment and radiation level effects in the enhancement
6
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TABLE 1. UV~B RADIATION LEVELS AT DIFFERENT POSITIONS BENEATH LAMPS,
MEASURED WITH IRL SPEC D SPECTRORADIOMETER. UNWEIGHTED POWER CONVERTED
TO WEIGHTED POWER EQUIVALENTS WITH THE AI9 WEIGHTING FUNCTION. VALUES
MEASURED AT NIGHT WITH NO NATURAL UV, AT SEEDLING HEIGHT BENEATH LAMPS
WITH 5 MIL CELLULOSE ACETATE FILTERS.
Location
beneath
lamps
A
B
C
D
E
F
G
Unweighted
power
(mW.m )
47..
9.4
17Q
2Q4
273
.385
478
Weighted
power*
(mW.m )
0.44
0.88
1.59
1.89
2.52
3.57
4.34
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TABLE 2. EFFECT OF NATURAL SUNLIGHT, SHADE, AND UV-B ON TERMINAL
BUD FAILURE AND SEEDLING MORTALITY IN ENGELMANN SPRUCE AND
LODGEPOLE PINE (EXCLUSION STUDY)
Terminal bud failure (%)*Mortality (%)
Treatment Spruce Pine Spruce Pine
Natural sunlight 32 10 0 8
(control)
Lath shade 52 10 0 8
Natural UV~B 40 4 24
(cell, acetate)
Excluded UV-B 30 020
(Mylar)
Significance N.S.** N.S, N.S. N.S,
* Includes trees which died.
** N.S. = not significant at P = 0.05.
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t
I
| TABLE 3. 'EFFECT OF UV-B RADIATION LEVELS ON TERMINAL BUD FAILURE
I IN ENGELMANN SPRUCE AND LODGEPOLE TINE (ENHANCEMENT STUDY)
I Terminal bud failure (%)*
Supplemental radiation level
Treatment A B C D E F G_
Engelmann spruce
Natural sunlight 42** 33 67 33 50 67 67
(control)
Plus UV-B (lamp 42 58 58 33 58 33 50
with cell, acetate)
Natural UV-B (lamp 25 50 42 67 83 50 50
with Mylar)
** Radiation level and treatment effects not significant (P = 0.05)
Lodgepole pine
Natural sunlight 8** 0 0 8 8 17 17
(control)
Plus UV-B (lamp
with cell, acetate) 0 0 8 0 17 17 33
Natural UV-B (lamp 0888800
with Mylar)
* Includes trees which died
** Radiation level and treatment effects not significant (P = 0.05)
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TABLE 4.' EFFECT OF UV-B RADIATION LEVELS ON MORTALITY IN ENGELMANN
SPRUCE AND LODGEPOLE PINE (ENHANCEMENT STUDY)
Mortality ~(%T"
Supplemental Radiation Level
Treatment A B C D E F_
Engelmann spruce
Natural sunlight 0* 0 0 8 8 0
(control)
Plus UV-B (lamp
with cell, acetate)
Natural UV-B (lamp
with Mylar)
* Radiation level and
Lodgepole pine
Natural sunlight
(control)
Plus UV-B (lamp
080
080
treatment effects
0* 0 0
000
0
0
8
0
not significant
8
0
0
17
0 0
0 0
(P = 0.05)
0 17
17 17
with cell, acetate)
Natural UV-B (lamp 0888800
with Mylar)
* Radiation level and treatment effects not significant (P = 0.05)
10
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TABLE 5. EFFECT OF NATURAL SUNLIGHT,. SHADE, AND UV-B ON VEGETATIVE
GROWTH OF ENGELMANN SPRUCE AND LODGEPOLE PINE (.EXCLUSION STUDY) .
VALUES ARE MEANS AND STANDARD DEVIATIONS
Treatment
Terminal
leader
length
(cm)
Number
of
branches
Total
branch
length
(cm)
Mean
branch
length
(cm)
Engelmann spruce
Natural sunlight
(control)
Lath shade
Natural UV-B -
(cell, acetate)
Excluded UV-B
(Mylar)
Significance
4.7+3.0 19.8+10.9 67.4+36.0 3.5+0.8
5.1+2.0 17.4+11.2 60.5+39.1 3.5+0.7
4.6+2.4 17.3+10.3 59.2+33.4 3.3+0.7
5.0 + 2.4 17.1 + 8.0 57.4 + 25.7 3.5 + 0.5
N.S.*
N.S.
N.S.
N.S.
Lodgepole pine
Natural sunlight
(control)
Lath shade
Natural UV-B
(cell, acetate)
Excluded UV-B
(Mylar)
Significance
6.4+2.1 7.0+ 3.3 15.1+ 8.0 2.2+0.8
6.2+2.4 7.8+3.8 15.2+ 8.0 2.1+1.0
6.4+2.5 9.3+ 4.1 18.5+ 8.0 2.1+0.8
t
6.0 + 2.4 9.5 + 4.9 16.2 + 8.2 1.8 + 0.7
N.S.*
N.S.
N.S.
N.S.
* N.S. = not significant at P - 0.05
11
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study are judged to be minor and inconsequential for the 67-day treatment
period.
I
I DISCUSSION
i Most effects of UV-B radiation on higher plants have been observed
I on herbaceous species. Studies on woody species have been very limited,
| although Biggs and Murphy (1977) reported damage of several conifers
Jl exposed to high UV-B levels shortly after germination. The effects of UV-B
,.; on tomato were cumulative (Caldwell et alf, 1974), Hart et al. (.1974) observed
j increased branching of chrysanthemums during exposure to UV-B.
| From these and other studies, it was anticipated that UV-B treatment
I effects might appear on new foliage produced during a treatment period exceed-
ing 2 months. Based on other research, particular attention was paid to i
stunting of needles and branches, color of foliage, and frequency of branching
(e.g. dormancy break of buds and subsequent growth).
The nearly complete lack of response of Engelmann spruce and lodgepole pine i'
to any of the treatments or radiation levels suggests two possibilities. First,
neither species may be sensitive to the UV-B levels used during this study. !
Secondly, because of the perennial nature of these species, a treatment and i-
observation period confined to a single growing season may be too short for r
effects to appear. |
Ronco's (1970 a and b) observations that solar radiation damage of
Engelmann spruce does not appear until the second growing season suggests
that the second possibility is more realistic. Clearly, it is premature to
conclude that UV-B has no effect on these two species,
It seems advisable to continue visual observations of the seedlings into
the second growing season, and for this reason the plants were moved to the
Fraser Experimental Forest where studies can be continued more conveniently.
It has not yet been decided, however, if the plants should be subjected to
the UV-B treatments again during the second growing season. It can be argued
that continual treatment during successive growing seasons is most realistic,
yet to do so would confound the second year's observations of carry-over
first-year effects. High variability and limited numbers of seedlings per
treatment prevent dividing the plants into two groups, one for observation
and the second for retreatment.
14
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i
A
i
j.
}
| REFERENCES
i
.' Biggs, R. H. and S. K. Murphy. 1971. Impact of solar UV-B radiation on
crop productivity. Third Quarterly Report} EPA-BACER, 7 pp.
Caldwell, M. M., W. B. Sisson, F. M. Fox, and J. R. Brandle. 1974.
j Plant growth response to elevated UV irradiation under field and
3 greenhouse conditions. Impacts of Climatic Change on the Biosphere.
1 CIAP Mono. 5, pp. 4-253 to 4-259.
j Hart, R. H., G. E. Carlson, H. H. Klueter, and H, R, Cams, 1974,
1 Responses to economically valuable species to ultraviolet radiation,
j Impacts of Climatic Change on the Biosphere, CIAP Mono, 5, pp.
4-263 to 4-273.
Ronco, F. 1970a. Chlorosis of planted Engelmann spruce seedlings unrelated
to nitrogen content. Can. J. Bot. 48:851-853.
Ronco, F. I970b. Influence of high light intensity on survival of planted
| Engelmann spruce. For. Sci. 16:331-339.
15
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APPENDIX
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TABLE 6. EFFECT OF UV-B RADIATION LEVELS ON VEGETATIVE GROWTH OF ENGELMANN SPRUCE (ENHANCEMENT STUDY)
VALUES ARE MEANS AND STANDARD DEVIATIONS
Radiation Level
1 Treatment
Terminal Leader Length (cm)
Natural sun (control)
Plus UV-B (cell, acetate)
Natural UV-B (Mylar)
A
3.9 +
2.8 +
4.1 +
No significant differences (P
Number of Branches
Natural sun (control)
Plus UV-B (cell, acetate)
Natural UV-B (Mylar)
24.2 +
19.3 +
16.1 +
No significant differences (P
Total Branch Length (cm)
Natural sun (control)
Plus UV-B (cell, acetate)
Natural UV-B (Mylar)
71.9 +
57.7 +
50.5 +
2.1
1.7
2.6
= 0.05)
8.8
8.8
8.9
= 0.05)
30.0
23.6
28.4
B
3.8 +
3.1 +
3.3 +
22.6 +
14.7 +
21.3 +
72.4 +
43.9 +
60.6 +
2.4
2.6
1.8
9.1
8.9
7.9
33.9
25.3
24.0
. Natural sun significantly different from plus UV-B
Mean Branch Length (cm)
Natural sun (control)
Plus UV-B (cell, acetate)
Natural UV-B (Mylar)
Significant radiation
3.0 +
3.1 +
3.3 +
effect
0.6
0.9
1.0
3.3 +
3.0 +
2.8 +
(P = 0.05) ; no
0.7
0.6
0.3
C
2.0 + 1.5
3.9 + 2.9
4.9 + 2.1
17.0 + 9.3
19.0 + 6.6
14.9 + 9.8
49.6 + 27.1
58.2 + 18.4
47.1 + 32.6
and natural
3.0 + 0.6
3.2 + 0.6
3.1 + 0.5
D
2.4 + 1
4.2 + 2
3.0 + 2
18.3 + 5
18.0 + 8
16.2 + 8
59.5 + 21
55.5 + 29
49.7 + 27
UV-B (P = 0
3.2 + 0
3.1 + 0
3.1 + 0
.8
.2
.2
.7
.1
.5
.7
.0
.2
.05)
.8
.5
.8
E
3.0 + 2.0
2.1 + 2.1
5.3+ 0.7
18.4 + 8.9
12.6+ 5.8
20.3+ 6.8
63.8 + 29.3
41.5 + 22.4
64.7 + 17.0
F
6.2 + 1.9
4.1 + 1.2
2.4 + 2.3
24.5 + 13.4
15.5 + 7.7
10.8 + 5.0
84.1 + 41.4
45.5 + 25.6
37.1 + 18.4
; no significant radiation
3.5 + 0.5
3.2 + 0.7
3.3 + 0.4
3.6 + 0.6
2.9 + 0.6
3.4 + 0.5
G
4.3 +
6.3 +
6.2 +
17.3 +
13.7 +
19.3 +
60.4 +
44.3 +
68.0 +
effect
3.7 +
3.8 +
3.6 +
0.6
1.0
2.0
12.3
12.5
11.1
39.5
35.5
41.4
0.9
1.0
0.7
significant treatment differences.
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TABLE 7. EFFECT OF UV-B RADIATION LEVELS ON VEGETATIVE GROWTH OF LODGEPOLE PINE (ENHANCEMENT STUDY) {
VALUES ARE MEANS AND STANDARD DEVIATIONS _ ; i
Radiation Level [
Treatment A B C D E F G |
Terminal Leader Length (cm) |
Natural sun (control) 6.6 + 2.1 5.7 + 2.9 5.0 + 2.3 5.1 + 1.6 6.3 + 1.7 4.7 + 1.3 5.5 + 2.0 j
Plus UV-B (cell, acetate) 5.7 + 2.8 5.5 + 3.0 4.9 + 1.7 6.1 + 2.0 5.6 + 2.9 4.8 + 2.7 5.4 + 3.0 !
Natural UV-B (Mylar) 5.1+ 2.5 4.5+1.7 5.3+2.1 5.4+2.4 5.0+ 2.2 5.8+ 3.7 5.1+ 2.0 f
No significant differences (P = 0.05) t
l
Number of Branches I
Natural sun (control) 6.9+ 3.8 ^6.2+3.2 7.3+3.7 9.4+4.7 8.7+ 3.9 9.8+ 5.6 9.2+ 3.6
Plus UV-B (cell, acetate) 8.7 + 3.1 9.2 + 3.3 7.5 + 4.3 8.8 + 3.0 7.5 + 3.4 8.6 + 3.1 8.0 + 4.7
Natural UV-B (Mylar) 6.5+ 3.3 7.2 + 2.7 6.2 + 2.9 5.5 + 2.3 8.4+ 4.8 6.2+ 2.6 6.2+ 2.8
Natural UV-B significantly different from natural sun and plus UV-B (P = 0.05); no significant radiation effect
Total Branch Length (cm)
Natural sun (control) 16.0+ 6.8 13.3+6.9 15.3+8.9 15.5+6.3 15.9+ 5.8 16.7+14.7 17.1+ 6.8
Plus UV-B (cell, acetate) 15.6 + 6.5 16.4 + 7.3 13.5 + 6.1 16.3 + 6.7 16.5 + 8.5 14.2 + 8.2 13.9 + 11.0
Natural UV-B (Mylar) 14.0+10.0 12.3+4.6 13.0+6.6 12.4+7.0 15.6+11.0 13.2+ 7.1 10,7+ 3.7
No significant differences (P = 0.05)
Mean Branch Length (cm)
Natural sun (control) 2.5+ 1.0 2.2 + 0.5 2.0 + 0.5 1.8 + 0.5 2.1+ 0.8 1-5+ 0.5 1.9 + 0.5
Plus UV-B (cell, acetate) 1.8+ 0.6 2.0+1.3 2.5+2.2 1.9+0.6 2.3+ 0.8 1-7+ 0.9 1-9 ฑ 1-6
Natural UV-B (Mylar) 2.3+ 1.1 1.8 + 0.7 2.1 + 0.8 2.2 + 0.8 1.8+ 0.5 2.1+ 0.8 2.0+ 1.0
i
No significant differences ( P = 0.05) 1^
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