/^CleaKjn
WATER POLLUTION CONTROL RESEARCH SERIES  I3030ELY4/7I-T
REC-R2-TI-7
OWR NO. l7V-IO
BIO-ENGINEERING ASPECTS OF AGRICULTURAL DRAINAGE
SAN JOAQUIN VALLEY, CALIFORNIA
REMOVAL OF NITRATE BY AN ALGAL SYSTEM
APRIL 1971
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I

CALIFORNIA DtPARTMENT OF WATER RESOURCES

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BIO-ENGINEERING ASPECTS OF AGRICULTURAL DRAINAGE
SAN JOAQUIN VALLEY, CALIFORNIA
The Bio-Engineering Aspects of Agricultural Drainage
reports describe the results of a unique interagency study
of the occurrence of nitrogen and nitrogen removal treat-
ment of subsurface agricultural wastewaters of the San
Joaquin Valley, California.
The three principal agencies involved in the study are
the Water Quality Office of the Environmental Protection
Agency, the United States Bureau of Reclamation, and the
California Department of Water Resources.
Inquiries pertaining to the Bio-Engineering Aspects of
Agricultural Drainage reports should be directed to the
author agency, but may be directed to any one of the three
principal agencies.
THE REPORTS
It is planned that a series of twelve reports will be
issued describing the results of the interagency study.
There will be a summary report covering all phases of
the study.
A group of four reports will be prepared on the phase of
the study related to predictions of subsurface agricul-
tural wastewater quality  one report by each of the
three agencies, and a summary of the three reports.
Another group of four reports will be prepared on the
treatment methods studied and on the biostimulatory
testing of the treatment plant effluent. There will be
three basic reports and a summary of the three reports.
This report, "REMOVAL OF NITRATE BY AN ALGAL SYSTEM",
is one of the three basic reports of this group.
The other three planned reports will cover (l) techniques
to reduce nitrogen during transport or storage, (2) possi-
bilities for reducing nitrogen on the farm, and
(3) desalination of subsurface agricultural wastewaters.

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BIO-ENGINEERING ASPECTS OF AGRICULTURAL DRAINAGE
SAN JOAQUIN VALLEY, CALIFORNIA
REMOVAL OF NITRATE
BY AN
ALGAL SYSTEM
Prepared by the
California Department of Water Resources
William R. Gianelli, Director
The agricultural drainage study was conducted under the direction of:
Robert J. Pafford, Jr., Regional Director, Region 2
UNITED STATES BUREAU OF RECLAMATION
2800 Cottage Way, Sacramento, California 95825
Paul DeFalco, Jr., Regional Director, Pacific Southwest Region
WATER QUALITY OFFICE, ENVIRONMENTAL PROTECTION AGENCY
760 Market Street, San Francisco, California 94102
John R. Teerink, Deputy Director
CALIFORNIA DEPARTMENT OF WATER RESOURCES
1416 Ninth Street, Sacramento, California 95814
DWR-WQO Grant #13030 ELY
DWR-USBR Contract #14-06-200-3389A
April 1971

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REVIEW NOTICE
This report has been reviewed by
the Water Quality Office, Environ-
mental Protection Agency and the
U. S. Bureau of Reclamation, and
has been approved for publication.
Approval does not signify that the
contents necessarily reflect the
views and policies of the Water
Quality Office, Environmental
Protection Agency, or the U. S.
Bureau of Reclamation.
The mention of trade names or
commercial products does not
constitute endorsement or recom-
mendation for use by either of the
two federal agencies or the Cali-
fornia Department of Water Resources.

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ABSTRACT
An algal system consisting of algae growth, har-
vesting and disposal was evaluated as a possible means of
removing nitrate-nitrogen from subsurface agricultural drain-
age in the San Joaquin Valley of California. The study of
this assimilatory nitrogen removal process was initiated to
determine optimum conditions for growth of the algal biomass,
seasonal variations in assimilation rates, and methods of
harvesting and disposal of the algal product. A secondary
objective of the study was to obtain preliminary cost esti-
mates and process design.
The growth studies showed that about 75 to 90 per-
cent of the 20 mg/1 influent nitrogen was assimilated by
shallow (12-inch culture depth) algal cultures receiving 2
to 3 mg/l additional iron and phosphorus and a mixture of 5
percent C02. Theoretical hydraulic detention times required
for these assimilation rates varied from 5 to 16 days, de-
pending on the time of the year. The total nitrogen removal
by the algal system, assuming 95 percent removal of the
algal cells, ranged from 70 to 85 percent of the influent
nitrogen.
The most economical and effective algal harvesting
system tested was flocculation and sedimentation followed by
filtration of the sediment. The algal cake from the vacuum
filter, containing about 20 percent solids, was then air- or
flash-dried to about 90 percent solids. The market value for
this product as a protein supplement was estimated to be
about $80 to $100 per ton.
Preliminary estimates indicate that the removal of
nitrate from tile drainage by an algal system will cost about
$135 per million gallons of treated water. This figure
includes engineering and contingency costs and recovery of
some cost by the sale of an algal product. The estimate will
be refined at the end of operational studies to be completed
in 1970.
Key words: Algae stripping, nutrients, tile drainage,
nitrogen removal, treatment costs.
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BACKGROUND
This report is one of a series which presents the
findings of intensive interagency investigations of practical
means to control the nitrate concentration in subsurface
agricultural wastewater prior to its discharge into other
water. The primary participants in the program are the Water
Quality Office of the Environmental Protection Agency, the
United States Bureau of Reclamation, and the California
Department of Water Resources, but several other agencies
also are cooperating in the program* These three agencies
initiated the program because they are responsible for pro-
viding a system for disposing of subsurface agricultural
wastewater from the San Joaquin Valley of California and
protecting water quality in California's water bodies. Other
agencies cooperated in the program by providing particular
knowledge pertaining to specific parts of the overall task.
The ultimate need to provide subsurface drainage
for large areas of agricultural land in the western and
southern San Joaquin Valley has been recognized for some time.
In 195^, the Bureau of Reclamation included a drain in its
feasibility report of the San Luis Unit. In 1957, the
California Department of Water Resources initiated an investi-
gation to assess the extent of salinity and high ground water
problems and to develop plans for drainage and export facili-
ties. The Burns-Porter Act, in i960, authorized San Joaquin
Valley drainage facilities as part of the State Water
Facilities.
The authorizing legislation for the San Luis Unit
of the Bureau of Reclamation's Central Valley Project, Public
Law 86-488, passed in June i960, included drainage facilities
to serve project lands. This Act required that the Secretary
of Interior either provide for constructing the San Luis Drain
to the Delta or receive satisfactory assurance that the State
of California would provide a master drain for the San Joaquin
Valley that would adequately serve the San Luis Unit.
Investigations by the Bureau of Reclamation and the
Department of Water Resources revealed that serious drainage
problems already exist and that areas requiring subsurface
drainage would probably exceed 1,000,000 acres by the year
2020. Disposal of the drainage into the Sacramento-San Joaquin
Delta near Antioch, California, was found to be the least
costly alternative plan.
Preliminary data indicated the drainage water would
be relatively high in nitrogen. The then Federal Water Quality
Administration conducted a study to determine the effect of
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discharging such drainage water on the quality of water in the
San Francisco Bay and Delta, Upon completion of this study in
1967# the Administration's report concluded that the nitrogen
content of untreated drainage waters could have significant
adverse effects upon the fish and recreation values of the
receiving waters. The report recommended a three-year re-
search program to establish the economic feasibility of
nitrate-nitrogen removal.
As a consequence, the three agencies formed the
Interagency Agricultural Wastewater Study Group and developed
a three-year cooperative research program which assigned
specific areas of responsibility to each of the agencies. The
scope of the investigation included an inventory of nitrogen
conditions in the potential drainage areas, possible control
of nitrates at the source, prediction of drainage quality,
changes in nitrogen in transit, and methods of nitrogen re-
moval from drain waters including biological-chemical processes
and desalination.
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TABLE OF CONTENTS
Page
ABSTRACT 		i
BACKGROUND		ii
CHAPTER I - CONCLUSIONS AND SUMMARY		1
Conclusions ..... 	 . 	 1
Summary ........ 	 .	1
CHAPTER II - INTRODUCTION		5
Algal Growth 				. .	7
Growth Characteristics 		8
Light ........ 		9
Temperature .......... 		10
Carbon Source 	 .........	11
Inorganic Macronutrients 		12
Micronutrients		 .	13
Algal Harvesting 	 .....	13
CHAPTER III - MATERIALS AND METHODS		15
Laboratory - Chemical and Biological .......	15
Algae				 .	17
Water					18
Algal Growth		22
Lightbox		22
Miniponds 				2k
Rapid Growth Pond (RGP)			26
Algal Harvesting		28
Laboratory		28
Pilot-Scale Separation Studies 		29
Sedimentation Tank		30
Shallow Depth Sedimentation Tajik		30
iv

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TABLE OF CONTENTS (Continued)
Upflow Clarifier .
Upflow Sand Filter
Microscreen . . .
Vacuum Filter . .
Centrifugation . .
CHAPTER IV - RESULTS AND DISCUSSION 	
Algal Growth 	
Lightbox Studies 	
Nutrient Requirements 	
Temperature 	
Algal Species 	
Water Source		 . . . .
Miscellaneous Lightbox Studies 	
Minipond Studies 	
Nutrient Additions 	
Phosphorus 	
Iron	
Carbon Dioxide 	
Mixing 	
Detention Time 	
Culture Depth 	 	
Soil Ponds 	
Biomass Control 	
Addition of Fish 	
Rapid Growth Pond 	
Biological and Chemical Observations 	
Biomass Production 	
Algal Genera Noted		 . . . .
Predatory Organisms 	
Dissolved Effluent Nitrogen 	
Dissolved Oxygen 	
pH and Alkalinity ... 	
RGP Evaporation and Temperature Data . . . .
Algal Harvesting ..... 	
Laboratory		 .
31
32
34
34
35
37
37
38
38
42
44
44
46
47
47
47
50
5}
i8
60
62
66
67
69
70
70
7il
74
75
U
80
80
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TABLE OF CONTENTS (Continued)
Pilot Scale Separation Studies 		87
Sedimentation Tank		87
Shallow Depth Sedimentation Tank
(Water Boy)		88
Upflow Clarifier 		92
Upflow Sand Filter (Sanborn Filter) ....	92
Vacuum Filter 		95
Micros creen		95
Centrifuges		96
Drying				97
Regrowth Studies 		97
Botulism Studies 		100
CHAPTER V - DISPOSAL	103
Animal Food Supplement	103
Use of Algae as a Soil Conditioner	108
Miscellaneous Possible Markets for an Algal
Product	110
Use of Algae to Produce Methane Gas .	Ill
CHAPTER VI - PROCESS EVALUATION	113
Removal Efficiencies 	 ....	113
Process Configuration 		115
Cost Estimates	115
CHAPTER VII - AREAS FOR FUTURE INVESTIGATION	123
ACKNOWLEDGMENTS 		125
LIST OF REFERENCES	126
PUBLICATIONS	131
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FIGURES
Number	Page
1	Growth of Algal Population in Batch Culture
(Hypothetical)			 8
2	Photomicrograph of Scenedesmus 	 18
3 Seasonal Variation in Total Dissolved Solids and
Nitrate-Nitrogen in Agricultural Drainage Water
Available at Wastewater Treatment Center ... 19
4	Algal Growth Facilities at Agricultural Waste-
water Treatment Center 	 21
5	Minipond Designs Used at the IAWTC ...... 25
6	Rapid Growth Pond Used for Large-Scale Algal
Growth Studies 	 27
7	Typical Dilution Curve Used to Determine Total
Solids from Percent Transmittance 	 29
8	Schematic and Flow Diagram of Water Boy
Shallow Depth Sedimentation Unit		  30
9	Effect of Tube Angle on Settling Path of
Suspended Particle ..... 	 31
10	Schematic Diagram of Upflow Clarifier 	 32
11	Schematic Diagram and Principle of Operation
of Sanborn Filter				 33
12	Principle of the Belt Filter	35
13	Effect of Phosphorous Addition on Nitrogen
Assimilation 	 39
14	Effect of Iron Addition on Nitrogen Assimila-
tion by Scenedesmus Cultures 	 39
15	Effect of Type of CO2 Addition on Nitrogen
Assimilation by Scenedesmus Cultures 	 40
16	Nitrate-Nitrogen Remaining at Day 11 in
Scenedesmus Cultures with Various Combinations
of Fe, P, and Carbon Dioxide	4l
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FIGURES (Cont'd)
Number	Page
17	Comparison of Nitrogen Assimilation by
Scenedesmus Cultures Using Water from Three
Different Tile Drainage Systems ....... 45
18	Comparison of Nitrogen Assimilation in Various
Tile Drainage Systems - October Sample ... 46
19	The Effect of Phosphate Addition on Nitrate
Assimilation in Outdoor Growth Units .... 49
20	Comparison of Nitrogen Assimilation in Mini-
ponds With and Without the Addition of 2 mg/1
P04-P . 		 49
21	Effect of Fe Additions on Algal Nitrate
Assimilation in Outdoor Growth Units in
Late Spring	 50
22	Effect of Fe Addition (3 mg/l) on Algal
Nitrogen Assimilation in Outdoor Growth
Units in Late Summer	... 50
23	Effect of CO2 Addition on Effluent Quality
in Outdoor Growth Units in Late Spring ... 52
24	Effect of Different Levels of COg Addition on
Nitrogen Assimilation in Ponds at 11.4 Days
Detention Time. 				5^
25	Effect of Continuous Mixing on Nitrogen
Assimilation, MP Run 3 				55
26	Effect of Mixing Duration on Average Total
Nitrogen Assimilation in Minipond Run 4A . . 56
27	Effect of Time of Mixing on Total Nitrogen
Assimilation 	 57
28	Effect of Detention Time on Nitrogen Assimila-
tion, 5 Percent CO2 Added		 . . . . 59
29	Effect of Detention Time on Nitrogen Removal -
Summary of Data from 12-Inch Ponds from July-
December 1969 	 ........... 60
30	Maximum and Minimum Minipond Temperatures -
July to December 1969 . . . 			 6l
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FIGURES (Cont'd)
Number	Page
31	Plot of Detention Time required to Assimi-
late Approximately 80 Percent of the Total
Nitrogen vs Average Run Temperature .... 6l
32	Effect of Three Different Culture Depths on
Total Nitrogen Assimilation 		 62
33	Average Total Nitrogen Assimilation for
Various Culture Depths, Minipond Runs 6-9B 63
34	Slope of Depth vs Percent Removal Curves
Compared with Average Solar Radiation During
Corresponding Run . . . .	 64
35	Comparison of Average Total Nitrogen Assimi-
lation at Two Culture Depths, 8 and 12 Inches
Inches, and Various Detention Times .... 64
36	Comparison of Total Nitrogen Assimilation in
One Soil Pond with a Comparably Operated
Minipond and Standard Minipond with Highest
Nitrogen Assimilation 	 ... 65
37	Comparison of Volatile Solids in Miniponds
With and Without Biomass Control      67
38	Percent Total Influent Nitrogen Assimilated
in Miniponds, With and Without Biomass
Control 		 67
39	Proportionate Increased Nitrogen Assimi-
lation Attributed to Pish, By Season ... 68
40	Operation of Rapid Growth Pond During 1969	69
41	Average and Range - Percent Volatile Solids
of Total Suspended Solids at Different
Levels of CO2 Addition	 71
42	Diurnal Fluctuations in Effluent Volatile
Solids	 71
43	Effluent Nitrogen and Volatile Solids Used
in Biomass Production Figures 	 72
44	Change in Nitrogen Forms During Minipond Run 75
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FIGURES (Cont'd)
Number	Page
45	Diurnal Fluctuations in Dissolved Oxygen . . 77
46	Diurnal Changes in Bicarbonate Alkalinity.  78
47	Average Daily Evaporation in Rapid Growth
Pond During 1969	 78
48	Daily Maximum and Minimum Temperatures in
Rapid Growth Pond During 1969		 79
49	Concentrations of Mineral Coagulants
Required for at Least 90 Percent Algae
Removal, Jar Tests			80
50	Concentrations of Lime, Alum and Ferric
Sulphate Required for 90 Percent Algae
Removal - Before and After Addition of
Ferric Chloride to Rapid Growth Pond ... 82
51	Percent Transmissions of Water Boy Influent
and Effluent Samples During First Three
Months of 1970 			89
52	Relation Between Solids Loading and Run
Length, Sanborn Filter 	 94
53	Algal Growth Responses of Various Combin-
ations of Untreated Agricultural Tile
Drainage and "Algal Stripped" Water Mixed
with San Joaquin River Water (Delta) ... 99
54	Nitrogen Assimilation in Various Minipond
Runs	 114
55	Flow Diagram of Algal Stripping Plant ... 115
56	Algae Growth Pond	 116
57	One of Three 12-Pond Groups Constructed
per Phase	 117
58	Predicted Seasonal Variation of Tile Drainage
Flow and Nitrogen Concentrations from San
Joaquin Valley, California 	 118
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TABLES
Number	Page
1	Relation Between Incident Light Intensity,
Depth and Algal Concentration at I
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TABLES (Cont'd)
Number	Page
15	Water Boy Operational Criteria and Results
for May 1970 	 90
16	Cost Comparisons of Pour Coagulants	 91
17	Composition of Slurry from Water Boy ... 91
18	Summary of Operational Data for Sanborn
Filter, 11/17-21/69 	 93
19	Number of Hours Required to Reach 80 Percent
Solids, from Original Sample Containing 6.^- 97
Percent Solids  ..
20	Composition of Algae Grown on Sewage .... 104
21	Protein, Carbohydrate, and Lipid Contents of
Some Freshwater Algae ..... 	 106
22	Amino Acid Composition of Oven-Dried RGP
Product Algae 	 107
23	Estimated Algal Production by an Algal
Stripping Plant, 1975-2000 	 109
2k	Rail Freight Rates for 100-Lb. Algae Packages
from San Francisco, California	110
25	Algae Stripping Design Criteria 	 , 119
26	Treatment Costs for Algae Stripping 	 121
27	Capital Cost for Algae Stripping ...... 122
xii

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CHAPTER I - CONCLUSIONS AND SUMMARY
Conclusions
1.	Algal growth and harvesting is a technically feasible
method of removing nitrate-nitrogen from subsurface
agricultural tile drainage in the San Joaquin Valley.
This method can be used to reduce 20 mg/l of influent
nitrogen to 3-5 mg/l which includes organic nitrogen
remaining in unharvested cells.
2.	Laboratory algal growth assays comparing algae-treated
and untreated tile drainage mixed with potential
receiving waters showed that treatment lowered the bio-
stimulatory nature of the waste.
3.	Preliminary cost estimates for the system described in
this report are about $135 per million gallons. This
figure is based on results of feasibility studies and
may be revised after 1970 operational studies.
Summary
Tile drainage will support extensive algal growth,
providing environmental conditions are optimum for such growth.
The effect of several chemical and physical factors on algal
growth were studied in laboratory and outdoor cultures. The
following summary of the factors includes the best estimates
of optimum levels for outdoor cultures.
1.	Three nutrient additives appeared necessary to
support sustained massive growth of Scenedesmus quadricauda
in tile drainage. About 2 milligrams of phosphorus per liter
of waste were needed regardless of season. The addition of
5 percent carbon dioxide was required during part of the
year, as was 2 to 3 mg/l of iron.
2.	Mixing data obtained before carbon dioxide was
added routinely to the outdoor cultures consistently showed
that rapid mixing was required for maximum nitrogen removal.
During the time when additional carbon dioxide was a growth
requirement, ponds with carbon dioxide addition only had
nitrogen assimilation rates comparable to those with both
carbon dioxide and mixing. The exact mixing requirements of
an algal system have not been defined but it appears that the
maximum requirement will be about four hours of mixing at an
average velocity of about 0.25 feet per second.
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3.	When phosphorus was available to the algae,
detention time was normally the most important variable
studied in individual outdoor culture experiments. The theo-
retical hydraulic detention times required for maximum nitro-
gen assimilation varied from 5 to 16 days and appeared to be
directly related to pond temperature within the range of 12C
and 25 C, and independent of temperature within the range of
25C to 33Co As shown in items h and 5 depth and biomass
control also significantly affected detention time.
4.	The optimum culture depth in these studies was
eight inches, but by a three to four day increase in deten-
tion time, comparable nitrogen assimilation rates could be
obtained at a 12-inch depth. Comparison of the difference
between nitrogen assimilation at two depths, 8 and 16 inches,
showed that the difference varied seasonally and was directly
related to available light. That is, depth had less effect
during summer light conditions than during the winter months.
5.	Some mechanism may need to be incorporated into
the growth units to control algal biomass. This mechanism
may consist of a settling area in the pond itself. The set-
tling area would remove the heavier, older, and less meta-
bolically active cells from the system. With such a system,
a minimum summer detention time of five days at a 12-inch
depth should be feasible. Without it, about eight days
detention time may be needed.
The two series of laboratory studies which compared
rates of nitrogen removal in effluents from various tile
systems in the Valley showed that the Alamitos sump water
used in these studies provided results comparable to the re-
maining systems studied. These studies also showed that the
addition of iron, phosphorus, and carbon dioxide was required
for maximum nitrogen assimilation by algae grown in water
from any of the systems studied.
During the investigation relatively little definitive
work was accomplished on soil-lined ponds. This process
probably involves a combination of algal and bacterial meta-
bolic pathways. Data from the two soil ponds at the IAWTC
showed that removal efficiencies often were comparable to
those of the best algal stripping pond and required only the
addition of phosphorus to achieve these results.
Harvesting of algal biomass is divided into three
steps or stages, namely, concentration, dewatering, and drying.
These steps differ in the amount of moisture remaining in the
algal product. Studies at the Interagency Wastewater Treatment
Center demonstrated that algae can be readily separated from
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agricultural tile drainage and concentrated to 1 to 2 percent
solids (by weight) either by coagulation-flocculation and
sedimentation with any of several chemical coagulants or by
use of a rapid sand filter (Sanborn filter) with backwashing.
The slurry resulting from the concentration process can then
be dewatered to about 10 to 20 percent solids by vacuum
filtration or by self-cleaning centrifugation. Using these
processes, the effluent algae concentration will be only 5
to 10 percent of the influent, at least within the influent
range of 100-600 mg/1 of algae.
Laboratory jar tests were conducted to determine
the effectiveness of various mineral coagulants (lime, alum,
and ferric sulfate) to determine their effectiveness in
achieving coagulation and flocculation of the alga Scenedesmus
in growth pond samples. The studies showed that the addi-
tions of these minerals could effect 90 to 95 percent removal
of the algae (influent suspended solids concentration of 100
to 600 mg/1) during all seasons of the year; however, the
required concentrations varied with changes in operation of
the growth unit. When iron (PeCl3) was added to the rapid
growth pond as an algal nutrient, the concentration of the
reagents required to remove 90 to 95 percent of the algae
was about 5 mg/l for ferric sulfate, 20 mg/1 for alum, and
40 mg/1 for lime. These are compared to about 80 mg/1, 100
to 140 mg/1, and 180 to 200 mg/1 for the same compounds when
iron was not present in the growth pond.
Approximately 60 polyelectrolytes were tested alone
and with the mineral coagulants to evaluate their effective-
ness in algal separation. Of the compounds tested, both
anionic and cationic, IT polyelectrolytes were found to aid
coagulation and to be economically comparable to mineral
coagulants. With iron added to the growth unit and no carbon
dioxide addition, almost complete (99 percent) algal separa-
tion was obtained with less than 0.2 mg/1 of the cationic
polyelectrolyte, Cat-Ploc.
The algal product was usually air dried to about
90 percent solids, although one sample was dried by a De Laval
spray drier at the company's test facilities. Two to three
days air drying was normally required to reduce the moisture
content to the desired level.
Literature review and market predictions indicate
that a market can probably be developed for an algal product
that will retail at about $80 to $100 per ton. The question
of marketability can be answered more completely when the
evaluations of the algal product are obtained from the com-
panies that received representative samples. The high ash
content of dried algae (30 to 50 percent) from a nitrogen
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removal plant operated on tile drain effluent may preclude
its use as a food for livestock but not as a protein supple
ment for fowl, or as a soil conditioner. Modification of
the harvesting process to include in-pond settling, as well
as chemical coagulation-flocculation may result in the pro-
duction of two different by-products, one with 10 percent
ash, and the other with about 50 percent ash.
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CHAPTER II. INTRODUCTION
This is the final report of field studies con-
ducted by members of the Interagency Nitrogen Removal
Group on the feasibility of using algal growth and har-
vesting (algae stripping) as a method of removing nitrate-
nitrogen from subsurface agricultural drainage in the
San Joaquin Valley. These studies were conducted at the
Interagency Wastewater Treatment Center (IAWTC) located
near Firebaugh, California. Field work began in January
1968 and continued through December 19693 however, opera-
tional studies will be conducted through December 1970.
They will be discussed in a later (June 1971) report.
The original impetus for considering the algal
process came from a formal feasibility report submitted to
Department of Water Resources (DWR) (Oswald, ej; al 1964).
Based on laboratory culture studies and a review of per-
tinent literature, this panel of consultants concluded
that algae stripping was a technically feasible means of
removing nutrients from tile drainage and that removal of
these nutrients would reduce the potential of the drainage
for causing deleterious algal blooms in the receiving waters.
The consultants proposed that a two-stage field study be
initiated to determine the exact levels of nitrogen re-
moval which could be realized by the process. The first
stage involved construction and operation of a small
(50 x 200 feet) prepilot plant. If the results of the
prepilot studies show that algal stripping is a promising
method, two pilot plants, each having a capacity of one
million gallons per day (mgd), would be built. The pre-
pilot plant was designed by engineers of the California
Department of Water Resources following suggestions by
the consultants and construction was started in July 1967.
Algae stripping is an assimilatory removal pro-
cess in which the nutrient in question is first incorpor-
ated into cellular tissue and the cells then removed from
the medium. The process includes three distinct areas of
activity: growing the algae, separation of the algae with
the incorporated nutrient from the liquid phase (including
drying of the algae), and disposal of the algal product.
The effluent nitrogen from an algae stripping plant will
consist of two fractions  the influent dissolved nitrogen
not assimilated by the algae and the particulate cellular
nitrogen not removed by the separation process. At the
IAWTC the effluent total nitrogen limit was 2 milligrams
per liter (mg/l) as recommended by the 1967 Environmental
Protection Agency (formerly the Federal Water Quality
Administration) report entitled "San Joaquin Master Drain,
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Effects on Water Quality of San Francisco Bay and Delta".
This amounts to a 90 percent reduction of the 20 mg/1 aver-
age nitrogen concentration predicted for a combined
San Joaquin Valley tile drain effluent. To remove 90 per-
cent of 20 mg/l by the algal process will require 95 percent
assimilation by the algae, with subsequent removal of 95
percent of the algal biomass.
The concept of using algal growth to remove
nutrients from wastewaters is not original with this
project, but its application to agricultural wastewaters
has received little consideration. Oswald and Gotaas
(1957), Fitzgerald (i960), Hemens and Mason (1968) and
North American Aviation (1967) are some of the workers
reporting on the use of the algal process to remove nutri-
ents, mainly nitrogen and phosphorus, from secondary sewage
effluent. These studies all indicated that tertiary treat-
ment by photosynthetic algae effectively removed nutrients
but that some problems were encountered. Because algal
growth is light and temperature dependent, growth and
nutrient assimilation were reduced during periods of cloud
cover and/or cold weather. The practical problem of eco-
nomically removing the suspended algae was also noted.
This investigation was intended to determine if these
limitations were equally Important in an area of moderate
climate and with agricultural wastewater as the culture
medium.
The general objective of the feasibility phase of
this study was to determine whether algae stripping would
effectively remove nitrogen from agricultural tile drain-
age effluent. In working towards this objective, several
questions had to be answered before a true assessment could
be obtained. Among the most important of these questions
were:
1.	Will agricultural tile drainage support
sustained algal growth?
2.	What levels of nitrogen assimilation can
be expected from this growth?
3.	What environmental conditions are needed
for maximum nitrogen assimilation and growth?
4.	Can the algae be readily and economically
separated from the liquid phase?
5.	What levels of total nitrogen removal can
be obtained by the algal stripping process?
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6.	Is there a potential market for an algal
product, or how do you dispose of the algal product?
7.	What is the total cost of an algal treatment
system, including growth, harvesting, and disposal?
8.	Does the process do the intended Job; that
is, does the treated effluent have less potential for
causing noxious algal blooms in the receiving waters?
9.	Are the results obtained from an isolated
tile system applicable to a combined drainage
facility?
The discussion of the results of a study of the
algae stripping process requires a basic understanding of
various concepts of algal growth and separation. A brief
review of some of the more pertinent facets of these two
areas will be included in the following sections.
Algal Growth
Algae cells contain a group of pigments, the
chlorophylls, which enable the organism to produce organic
material through a series of reactions requiring light
energy, water, carbon dioxide, and various inorganic nutri-
ents. The photosynthetic process can be summarized by the
following approximation (Jewell and McCarty, 1968):
aCOg	+ ePCty ^ + (c+3e)H^" + l/2(b-c-*3) HgO +
sunlight 	^CaHbNcodPe + (a+b/4+5c/M-d/2+5e/4)02
as Indicated by this equation, the rate of nitrogen (or
phosphorus) assimilation by algal cells is a function of the
rate at which organic material is synthesized. In a nu-
trient removal system, such as that studied at the IAWTC,
the ultimate goal of systems design is to have the undesir-
able nutrient  nitrogen for example  to limit formation
of cellular material. Using this concept, all other re-
quired elements of photosynthesis should be available in
optimum or excess amounts. To accomplish this goal, the
effect of various factors on photosynthesis must be
understood.
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Growth Characteristics
Although this category does not fit conveniently
into the classification of factors affecting photosynthetic
activity, some explanation of algal growth in two general
types of algal systems may be helpful. A generalized algal
growth curve for algae grown in a culture without nutrient
replenishment (batch culture) is shown in Figure 1. The
initial period of
adjustment to the
medium (lag phase)
is followed by a
period of rapid
cell division (log
or exponential
phase). Cell growth
is eventually lim-
ited by nutrient
availability or by
light limitation
caused by mutual
cell shading. The
algal biomass, or
cell numbers, may
then decline, al-
though in multi-
algal cultures the
original species
may be replaced by
another alga with different nutritional requirements. The
specific growth rate of cells during the exponential (log)
growth phase in this type of system is a function of cell
concentration and can be described by the equation:
dN - KN
where K is the specific growth rate (day *), N is the cell
concentration (in any applicable unit), and t is the time
in days.
In most waste treatment systems utilizing algae,
algal nutrients flow continuously through the system and
thus are constantly renewed. The theoretical analysis of
this type of cellular growth, continuous culture, has been
comprehensively treated by Monad (19^9) and Its application
to algal systems reviewed by Retovsky (in Malek: and Fencl,
1966) and Shelef, et_ al (1968). The importance of this
type of process to algal cultures is that by manipulation
of environmental characteristics (nutrient concentration,
z
UJ
N
V)
z
o

_l
a
a.
2
TIME-DAYS
FIGURE I - GROWTH OF ALGAL POPULATION
IN BATCH CULTURE (HYPOTHETICAL)
-8-

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light, and so forth) the algal population can be maintained
at steady-state, or constant, population density. Con-
versely, manipulation of population density can cause some
factor (nutrient) to be rate-limiting. As will be shown in
the Materials and Methods chapter, the growth units used in
this study were designed to be approximations of a continu-
ous-flow type of system, with nitrogen as the rate-limiting
factor.
Light
Absorption of light energy by dense algal cultures
follows the Beer-Lambert law:
Id - loe"E0d
where I0 is the incident light intensity, Id is the inten-
sity of light at any depth, d is depth in centimeters, c is
the algal biomass in mg/1, E is the extinction coefficient
in cm2/mg, and e is the base of natural logarithms. As
shown by this equation, light penetration to X* is directly
affected by incident light and inversely affected by depth
and culture density. Optimum light intensities for maxi-
mum algal growth range from 200 to 400 foot-candles (ft-c),
and the lower limit may be 100 ft-c. Using an extinction
coefficient of 2 x 10"3 cm2/mg (determined experimentally),
Bogan, et al (i960) calculated the depth at which various
lncidenlPlight levels would penetrate several different
concentrations of algae and still leave 100 ft-c. These
values are shown in Table l.
TABLE 1
RELATION BETWEEN INCIDENT LIGHT INTENSITY,
DEPTH AND ALGAL CONCENTRATION AT Id - 100 ft-c
(Prom Bogan, i960)
centration	" cm for corresponding I0
(mg/l) 1,666 tt-0 2,660 ft-c ,000 ft-c 10,000 ft-c
50
23
30
39
46
100
11.5
15
19.5
23
200
5.8
7.5
9.8
11.5
400
2.9
3.8
4.9
5.8
-9-

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Using similar calculations Golueke and Gotaas (1958) deter-
mined that 4.5 inches was the theoretical optimum pond
depth for domestic wastes, but practical experience indi-
cated that optimum culture depth below which light no longer
limited algal growth was about 8 inches.
The practical implications of the light-absorption
equation are that if light-limiting conditions are to be
avoided, high-rate algal ponds must be shallow and that depth
may have to be adjusted seasonally. Even in the relatively
shallow cultures used in chemostat (laboratory continuous-
flow) studies, only a small percentage of the incident light
energy is converted to cellular energy. Oswald (1963) re-
ported that in laboratory studies with settled sewage, an
average of 4 percent of the incident energy was fixed by
the algal cultures. Conversion efficiency varied inversely
with intensity, duration of light, and detention time, and
directly with temperature and carbon dioxide concentration.
Another possible method of increasing the avail-
ability of incident light to individual cells is to move the
algae into the light path by induced turbulence (mixing).
Kok (1953) found that increased algal yield could be ob-
tained in Chiore11a cultures by using intermittent or
flashing light, and that the optimum proportion of light-
to-dark period appeared to be about 1:9. In these studies
the flash time varied from 3 to about 200 milliseconds. In
outdoor cultures the flashing light effect can be achieved
by mixing a sufficiently dense algal culture; however, the
mixing would have to be such that the duration of exposure
to light and dark were of the correct time intervals.
Mixing of this type would not be economically feasible in
conventional wastewater treatment systems.
Temperature
As with all organisms, temperature affects the
growth rate of algae, normally following the Van't Hoff rule
according to which the growth rate doubles for each 10C
increase in temperature, within the range of temperature
tolerance. Oswald (1963)# working with Chlorella
pyrenoldosa. noted that optimum light conversion efficiency
occurred at 20C. Using mixed cultures of Chlorella and
Scenedesmus, Witt and Borchardt (i960) found that the light
saturation intensity level (lowest light level at which
maximum growth rate was attained) was directly affected by
temperatures between 20 and 30C. Because it is normally
impractical to heat an outdoor algal culture, seasonal tem-
perature variations cause changes in the required detention
times of the system. In periods of cold weather, the algae
-10-

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are allowed a longer contact time with the nutrient supply,
which compensates for slower growth rates.
Carbon Source
Algae normally use free carbon dioxide as an in-
organic carbon source, although some algae have been reported
to use the bicarbonate ion. Osterlind (1950) stated that
Scenedesmus quadricauda could freely use the bicarbonate ion
but that Chlorella pyrenoldosa would only grow if provided
with carbon dioxide (CO?) In poorly buffered systems, use
of CO2 and bicarbonate THCO3)causes the equilibria in the
following equations to shift to the right, accompanied by
a rise in pH.
COg + 1^0 ^ HgCOg	H+ + HCOg H* + CO^
The concentration of any of the components of the
carbon dioxide-bicarbonate-carbonate buffer system is a
function of temperature, pH, and total dissolved solids as
well as the concentrations of the remaining components. The
equilibrium equation for the formation of hydrogen and bi-
carbonate ions from carbonic acid is:
[HCOo + H^j - Kj,
where the dissociation constant, Ki, has been reported to be
3.5 x 10"7 at 18C (Chemical Rubber Publishing Co., 1951;
At a pH of 8 the ratio of carbonic acid to bicarbonate ion
Is 0.0286, at a pH of 7 It is 0.286, and at a pH of 6 it is
2.86 (McKee and Wolf, 1963). a similar equilibrium reaction
between HCO3 and COo* has a reported K2 of 4.4 x 10""11 at
25C (Chemical Rubber Publishing Co., 1951); thus at pH 7
the ratio of bicarbonate to carbonate ions would be 2,270
to 1, whereas at pH 11 the ratio would be 1 to 4.4 (McKee
and Wolf, 1963).
At pH values above 9, carbonate precipitates as
calcium and magnesium salts, thus decreasing total alka-
linity. These precipitates also remove many algal nutrients,
especially phosphorus and heavy-metal trace elements, by
forming complex salts. In outdoor algal cultures exposed to
the atmosphere, afternoon pH values may be as high as 10.5
to 11.
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Inorganic Macronutrients
Inorganic macronutrient requirements include nitro-
gen and phosphorus. Most algae containing chlorophyll are
able to use either ammonia or nitrate as a nitrogen source,
and many species can use nitrite providing the concentration
is low, about 0.001 molar (Fogg and Wolfe, 1954). Ammonium-N
has been reported to be used in preference to nitrate, when
both sources are provided to the same culture (Schuler,
et al, 1953). As shown by the following equations, the use
oT either of these nitrogen sources can cause undesirable
changes in culture pH (Cramer and Meyers, 1948).
1.0(N03)+5.7(C02)+5.4(H20)->'C5#7H9^802^3N1#o+8.25 02+1.0(0H")
i.o(nh4+)^.7(co2)+3.4(^0)^05^^0^31^0+6.25 Og+i.oCff1-)
Nitrate assimilation results in the production of OH" ions
which causes a rise in pHj whereas ammonium assimilation
lowers the pH by formation of hydrogen ions.
In addition to dissolved nitrogen sources, many
blue-green algae can fix atmospheric nitrogen (Fogg, 1947).
A requisite for nitrogen fixation appears to be an extremely
low concentration of available dissolved nitrogen in the
growth medium. Dominance of nitrogen-fixing algae in a
treatment plant would preclude the use of algae stripping
as a means of removing nitrogen.
Algae usually use phosphorus as orthophosphate
(P0i|~3). The ratio of nitrogen to phosphorous concentrations
in a typical algal cell is about 10:1. Phosphorus is es-
sential to algal growth. Without It, no growth will occur,
regardless of the algal species. Sawyer (1952) found that
the N:P ratios in natural waters that were seemingly optimal
for algal blooms varied from 30:1 to 15*1* depending on the
algal species involved. Some algae, when provided with
quantities of phosphorus in excess of their requirements,
can "store" the element up to a certain concentration
(Ketchum, 1939a). Zabat (1970) found that, regardless of
the availability of phosphorus, the maximum uptake by
Chlorella soroklawa was 2 percent of the cellular dry weight.
Phosphorus uptake can be influenced by light (Gest and Kamen,
1948) and hydrogen ion concentration. Hydrogen ion concen-
tration affects availability and has been mentioned
previously.
-12-

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Mlcronutrlents
Many trace minerals are required for algal growth,
although the actual concentrations required may be beyond
the limit of detection by routine analytical procedures.
The elements most commonly mentioned are iron, molybdenum,
manganese, vanadium, cobalt, zinc, copper, sodium, and
boron. In laboratory cultures, an accepted method of pro-
viding these nutrients is by means of a soil extract (the
supernatant from a sample of soil boiled in distilled water).
The trace element concentration in agricultural tile drain-
age may resemble a soil extract in that the water passes
through several feet of soil before entering the drainage
system.
This brief summary shows that the growth studies
at the IAWTC had to include the investigation of several
variables, some of which (temperature and light, for example)
were fixed by natural conditions.
Algal Harvesting
This phase of the study of the algae stripping
process was designed to determine the most feasible method
of removing the algae from the growth units' effluent while
maintaining an acceptable concentration of algae in the
plant effluent. The harvesting process can be divided into
three distinct areas of activity, based on the amount of
water retained in the algal product. The first step, con-
centration, increases the solids from 0.015 to 0.040 per-
cent by weight (depending on the concentration of algae in
the growth unit) to 1 to 4 percent. The second step is de-
watering which then brings the solids to 8 to 20 percent,
and finally in the third step, the algal mass is dried to
85 to 92 percent solids by weight. At the latter moisture
content, algae can be stored almost indefinitely without
decomposition (Oswald and Golueke, i960).
Relatively little work has been done on algal
separation, especially at prepilot or pilot-scale levels.
Oswald and Golueke (1968) present a comprehensive review of
the results of several years of work on the problem of sepa-
rating microscopic algae grown on secondary sewage effluent.
In general, their results showed that the algae could be
most economically concentrated by coagulation, flocculation,
and sedimentation. (Coagulation is defined as the destabi-
lization of colloidal particles, and flocculation as the
mixing of the destabilized particles to encourage the for-
mation of larger "floe masses.) Dewatering was accomplished
-13-

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by centrlfugation, with final drying in the open. An alter-
native to separate dewatering and drying steps was to spread
the concentrated algal slurry on sandbeds, which brought
about the desired moisture content without the intermediate
step of dewatering. North American Aviation (1967) also
found sandbed dewatering and drying feasible in the harvest-
ing of sewage-grown algae. In their studies, the algae were
concentrated by sedimentation after coagulation and floc-
culation and then spread on sandbeds. These studies were
conducted in Central and Southern California, which are areas
of moderate climate.
A primary consideration in the algal harvesting
process is the concentration of algae remaining after
routine separation of the algal biomass from the liquid
phase. Based on 2 mg/1 total nitrogen in the effluent, the
maximum allowable concentration of algae in the effluent
would be about 15 mg/l. The 2 mg/1 effluent nitrogen will
probably contain about 0.5 mg/1 dissolved nitrogen with the
rest as particulate nitrogen. Algae contain about 10 percent
N; therefore, 15 mg/1 of algae can provide up to 1.5 mg/1 N.
Based on these figures, about 90 to 95 percent of the algae
would have to be removed from the effluent of an operating
algal treatment plant. Without this level of removal,
organically-bound nitrogen would be regenerated by bacterial
decomposition of the algae and would be available for algal
growth downstream from the treatment site. Jewell and
McCarty (1968) found that the amount of nitrogen and phos-
phorus regenerated by the aerobic decomposition of algae
varied with culture age, with almost no regeneration taking
place in older cultures. The average amount of N regenerated
after about 300 days of aerobic decomposition was about 49
percent of the original amount present ih the cells. Poree
and McCarty (1968) reported that after 200 days of anaerobic
decomposition, algal cultures retained about 60 percent of
the original particulate nitrogen. These studies indicate
that algae cellular nitrogen must be considered to be part
of the total nitrogen released from a nitrogen stripping
plant.
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CHAPTER III - MATERIALS AND METHODS
The studies described in this report, with the ex
ception of regrowth studies, were conducted at the Inter-
agency Wastewater Treatment Center. In both algal growth
and harvesting, studies were conducted using laboratory
"bench-scale" experiments and expanded prepilot and pilot-
scale studies. In general, the laboratory tests in both
phases of the algal system were designed to screen a series
of compounds or variables before testing in the larger
systems. The following sections describe in detail the
various materials and methods.
Laboratory - Chemical and Biological
A chemical and biological laboratory was located
at the site and contained the equipment needed to perform
routine chemical analyses. This included a Kjeldahl distil-
lation and digestion unit, gas chromatograph, specific ion
electrodes, spectrophotometer and other routine laboratory
equipment. For special analyses, samples were sent to the
Department of Water Resources' laboratory at Bryte,
California. Samples for trace metal analysis were sent to
the U. S. Geological Survey laboratory in Sacramento for
analysis by emission spectrograph. Almost all chemical
analyses used in these studies followed the procedures
lined in Standard Methods for Examination	Wa;3te
Water, American Public Health Association,lyop Table 2
lists the routine chemical analyses used and the normal
frequency of analysis.
As shown in Table 2, the analytical emphasis was
on nitrogen forms, especially nitrate and nitrite.
inary studies on all the listed constituents indicated that
sampling and analysis at a greater frequency than shown was
not necessary to follow changes in the algal system. Most
of the analyses listed were completed within two to five
hours of sample collection and were not preserved in any
way.
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TABLE 2
CHEMICAL ANALYSIS SCHEDULE
Constituent
Frequency
Method
Nitrate
3 times/wk.
Brucine, specific


ion electrode
Nitrite
3 times/wk.
Diazotization
Ammonia
once/wk.
KJeldahl - Distil-


lation
Organic Nitrogen
twice/wk.
Kjeldahl
Orthophosphate
twice/wk.
Stannous chloride
Iron
once/wk.
Phenanthroline
Chemical Oxygen Demand
Occasionally
Dichromate refluxing
Dissolved Oxygen
Occasionally
Winkler-Azide Modi-


fication
PH
Daily
Electrode
Alkalinity
twice/wk.
Titration - pH meter
Electrical Conduc-
Occasionally
Wheatstone Bridge
tivity

Evaporation
Total Dissolved Solids
Occasionally
Precision and accuracy tests on all nitrogen forms
and orthophosphate indicated that experimental error was
within the limits suggested in Standard Methods. In spite of
this, the routine use of the brucine test for nitrate, with
some procedural modifications, often gave results of ques-
tionable validity. To avoid this problem, use of the nitrate
specific ion electrode was begun early in 1969. The instru-
ment was standardized against known concentrations of nitrate
in denitrified drainage water. A plot of meter readings
versus concentration showed a straight line between 0.5 and
50 mg/l N. Although some problems were encountered, mainly
day-to-day variations in electrode response, the specific
ion electrode proved to be a rapid (up to 150 analyses per
hour), simple, and reliable method of nitrate analysis.
The biology section contained microscopes, an
incubator, bacterial culturing equipment, and other neces-
sary biological supplies and equipment. The primary method
used to determine changes in algal biomass was measurement
of volatile suspended solids. Volatile suspended solids
were normally run two to three times per week, with total
suspended solids once a week. One of the volatile solids
analyses was conducted on the day total organic nitrogen
analyses were made. The procedure for suspended and volatile
-16-

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solids involved filtering 50 to 100 milliliters of algal sus-
pension onto a weighed, preignited Whatman QFA glass filter
disk. The disk was then dried for one hour at 103C, weighed,
and ignited at 56oC for 15 minutes. A final weighing was
then made to determine volatile solids, which were assumed
to represent algal biomass.
Approximate cell counts were made at weekly inter-
vals, using a hemacytometer counting cell. These cell counts
were used mainly to observe the condition of the algae and
changes in species composition.
In some lightbox studies, cell growth was followed
by measuring 3jn vivo chlorophyll fluorescence. A Turner
Model III fluorometer was modified by adding a blue light
source and the proper combination of filters (Corning
CS 5-60 primary and a Corning CS 2-60 secondary) for measure-
ment of chlorophyll a.
The fluorometer, with a different light source and
filter combination, was also used to determine true detention
times in the miniponds. Changes in effluent concentration
of Rhodamine B dye were followed and the detention time and
mixing characteristics of the unit determined.
Adjacent to the laboratory a continuous recording
analyzer monitored rapid growth pond (RGP) water temperature
and pH, and sunlight. A weather station, located on one of
the RQp baffles, was checked daily to determine maximum and
minimum air and water temperatures, evaporation, precipita-
tion, and wind.
Algae
The original inoculum for the rapid growth pond was
obtained in January 1968 from the University of California's
Richmond Field Station, Approximately 100 pounds (20 percent
solids) of a sewage-grown green alga, Scenedesmus quadri-
cauda, or closely related species, were added directly to
the RGP. This amount was added to the RGP on two more oc-
casions, in the summer of 1968 and early in 1969 when other
algal species became dominant in the pond cultures. During
the remainder of 1969* Scenedesmus retained its dominance
and the rgp was used as a source of inoculum for all growth
studies.
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A coenobium of typical Scenedesmus quadrlcauda
cells Is shown In Figure 2. This alga is normally a four-
celled coenobium; however, it is also found in a single-cell
form or with as many
as 32 cells in a col-
ony. The genus has a
cosmopolitan distri-
bution, and one or
more of the 30 known
freshwater species
can be found in most
freshwater environ-
ments in this country.
The choice of this
alga was mainly a
matter of its avail-
ability in large
quantities from the
Field Station. Pre-
liminary culturing
studies using tile
drainage demonstrated that the species would grow in this
type of medium. Additionally, the use of Scenedesmus
quadrlcauda has some inherent advantages over most other
algal species. Because of its wide distribution, planktonic
nature, and ease of culturing, much literature is available
on its physiology and biochemistry. Also, this alga has
been reported to use the bicarbonate ion (Osterlind 1950),
a pathway not available to all algae.

FIGURE 2-PHOTOMICROGRAPH OF SCENEDESMUS
Water
Subsurface agricultural drainage was obtained from
a collecting system called the Alamitos tile system, located
near the site. The Alamitos system was selected for its
easy access from readily available Bureau of Reclamation land
and because extensive sampling had indicated that the water
quality was similar to that predicted for the combined drain.
This particular tile system drained about 400 acres of in-
tensively farmed land on which safflower or barley was grown
in the winter and principally rice and cotton in the summer.
This type of farming resulted In predictable changes in the
quantity and quality of the drainage water, the effects of
which are illustrated in Figure 3. Decreases in nitrate-
nitrogen and total dissolved solids (TDS) concentration in
the late spring were caused by increased tile flows when the
rice crop was flooded. Tile flows varied from more than
1,000 gallons per minute (gpm) in the summer to less than
20 gpm in the winter of 1967-68.
-18-

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10,000
o>

(/)
Q
_J
O
W
a
Ui
o
m
V)
<
h
O
8,000
6,000
4,000
2,000-
JAN
NITRATE-NITROGEN (mq/l)
TOTAL DISSOLVED S0LIDS(mq/l)
30 =
o>
E
z
Ul
o
o
cc.
J-
UJ
<
DC
JUL	JAN	JUL
1966	1967
FIGURE 3-SEASONAL VARIATION IN TOTAL DISSOLVED SOLIDS AND
NITRATE-NITROGEN IN AGRICULTURAL DRAINAGE WATER AVAILABLE
AT WASTEWATER TREATMENT CENTER
TABLE 3
CONCENTRATION OP MINERAL CONSTITUENTS IN TILE DRAIN WATER
(in milligrams per liter)
Constituent
Samples Taken at Alamitos Sump
5/2/67	7/14/6?
(before flooding) (after flooding)
Master Drain,
1969
Calcium
Magnesium
Sodium
Potassium
Carbonate
Bicarbonate
Sulfate
Chloride
Nitrate
Hardness
Total Dissolved
Solids
Boron
373
194
190
106
1,390 ,
875
5.4
4,
0
0
247
373
3,600
2,070
559
304
17.6
9
1,710
921
6,590
3,950
14
9
220
160
1,900
20
0
220
3,500
1,000
20
1,200
6,800
11
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In Table 3 results of analyses made before and
after flooding are compared with quantities predicted
for the Master Drain. From these and other data,
the Alamitos sump water appears to be quite similar to
that of the predicted combined drain effluent during most
of the year. However, flooding of the rice fields had a
marked effect on water quality.
Analysis of the trace mineral results in Table 4
indicates that these constituents remained relatively constant
throughout the year. Because of the semiquantitative nature
of the emission spectrograph, probably no major differences
(with the possible exception of molybdenum) are shown.
These elements are probably of limited solubility; therefore,
their concentrations may be independent of flow.
Because of the fluctuations of water quality illus-
trated in Figure 3, a covered storage pond was used to
provide a constant supply of tile drainage water for the
lightbox and all outdoor units. This pond, which could
store about 820,000 gallons, was lined with polyethylene
plastic to prevent loss of water or contamination by the
ground water. The cover and lining minimized biological
activity, thus providing a reasonably constant 30- to 90-
day water supply depending on depth and detention time of
the rapid growth pond.
-20-

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I
ro
M
i

LEGEN
o - TILE DRAINAGE SUMP d - RAPID GROWTH POND
b - LIGHTBOX TRAILER	e-LABORATORY BUILDING
c-COVERED STORAGE POND f - SEPARATION STUDIES AREA
g-MINIPOND GROWTH UNITS
FIGURE 4 - ALGAL GROWTH FACILITIES AT AGRICULTURAL WASTEWATER TREATMENT CENTER
Et

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TABLE 4
TRACE METAL ANALYSES OF ALAMITOS SUMP WATER DURING
1968-69
Concentration - parts per billion
Element	Sample Date
12/10/68 2/12/69 3/20/69 4/8/69 9/6/69
Aluminum
31
1.4
2.5
1.7
14
Cadmium
23
20
2.5
12
11
Iron
8.6
4.3
3.2
10
6.3
Manganese
12
18
4.4
15
43
Molybdenum
40
17
32
77
57
Nickel
8.0
8.0
7.2
7.7
6.6
Vanadium
1.9
3.1
2.7
2.0
2.3
Zinc
5.7
5.7
10
6.7
5.7
Copper
2.9
1.4
2.5
1.7
1.7
Cobalt
1.4
1.4
2.5
1.7
1.4
Algal Growth
The effect of different variables was tested at
three levels of culture size: lightbox batch assays, small
1,000-gallon outdoor growth units, and a large 1/4-acre
demonstration pond. Because of lack of replicability of the
large pond, most of the actual experimental work was con-
ducted using the lightbox and small outdoor culture units.
An aerial photograph of the Treatment Center (Figure 4) shows
the location of the various facilities used in this
investigation.
Lightbox
The lightbox was located in an air conditioned, con-
verted office trailer provided with forced-air heating. The
trailer was partially Insulated and temperatures were
-22-

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maintained at 25 4C. The lightbox consisted of two s{flves,
12 x 3 feet, which held 200 one-liter culture flasks. No
means of automated, mechanical agitation was provided, but
air (compressed only or enriched with CO2) could be intro-
duced to individual flasks via a central manifold. Air
volume was regulated by short sections of capillary tubing
in each line and resulted in approximately equal amounts of
air being delivered to each flask.
Four 12-foot cool-white fluorescent fixtures, com-
prising a total of eight lamps, were placed about 15 ^hes
above each shelf. Ballasts were separated from the light
fixtures to minimize local heating. Light intensi
the shelves ranged from 350 to 400 foot-candles, depending
on location. The lighting was controlled by a timer to
regulate the light-dark cycle.
Laboratory growth studies were customarily conducted
with 1.000 milliliter (ml) erlenmeyer flasks containing 500
ml of culture.Each variable was tested in triplicate.
Drainage water from the storage pond was used most of the
time. On occasion, water was taken directly from the sump
to determine whether storage affected the water's potential
to support algal growth. A culture containing predominantly
Scenedesmus from the outdoor growth units, usually the rapid
growth pond, was used to inoculate individual flasks. The
initial inoculum was normally on the order of 2,000 to 3,000
cells per milliliter. No attempt was made to obtain uni-
algal or bacteria-free cultures in these studies; however,
the algae in the inoculum were normally 90 to 95 percent
Scenedesmus.
Almost all lightbox studies conducted in 1969 used
continuous lighting. No detrimental effect appeared, and the
experimental time was shortened, allowing more assays to
be run. In the laboratory studies, various levels of CO2
enrichment were tried. The addition of C02 was accomplished
by methods ranging from hand swirling, accelerated
of atmospheric CO2 Into the culture medium, to	0
100 percent CO2. Temperature studies	conducted 1Lt-h*
immersing culture flasks to a one-inch depth in water baths
at controlled temperatures.
Certain observations were made dally during
course of the studies. Because of the prlmary l^erest in
nitrogen uptake, emphasis was placed on the collection ol
daily nitrate data obtained by means of the specific ion
electrode. Other parameters that were often
were pH, maximum and minimum temperatures,	ce*
At the end of a run, the triplicate flasks of each set of
-23-

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variables were often pooled for volatile solids, total
Kjeldahl nitrogen, nitrite, and cell count determinations.
When the effect of various nutrient additions was tested,
the stock solutions used were prepared from analytical or
reagent grade chemicals.
Miniponds
A total of 22 small, resin-coated plywood ponds,
termed "miniponds", were operated at Firebaugh. Eighteen
of these ponds were 8 feet wide x 16 feet long with a
designed operating depth of 12 inches (3 inches of freeboard).
Approximate capacity of these ponds was 1,000 gallons. Two
other ponds, also 8 x 16 feet, were used to study operating
depths of 8 and 16 inches. The remaining two ponds (12
inches operating depth) were modified in the spring of 19&9
to improve the hydraulic characteristics of the ponds,
specifically to eliminate stagnant zones during mixing.
A comparison of the two designs is shown in Figure 5.
All but two of the miniponds had mixing pumps whose
capacity was approximately 80 gallons per minute (gpm).
This provided average in-pond velocities of 0.25 to 0.5 feet
per second (fps). Timers were placed in the electrical
circuits of the pumps to vary mixing during a 2^-hour period.
The flow of water from the storage pond was metered
into ponds by means of individual flowmeters. All water
lines were made of opaque material to prevent clogging by
algal growth. Effluent was drawn from near the bottom of
the storage pond and discharged through a "broken" siphon
arrangement. This device was also used to maintain a con-
stant water depth.
Normal operation of a minipond run began with the
removal of all algae from a previous run. The ponds were
then started with a common source of algae, usually from the
RGP. For the first few days of a run, the ponds were oper-
ated either on a batch basis or at long detention times.
This acclimatization period was designed to allow algal
biomass to accumulate before being influenced by the experi-
mental variables. Pond samples were collected dally from
near the effluent at about 0830 hours and taken to the
laboratory for analysis. In-pond pH values were determined
once or twice daily by a portable pH meter.
All water delivered to the miniponds was taken
from the storage pond and metered into the ponds by flow-
meters (rotameters) that were adjusted daily. Periodically,
the rotameters were calibrated by timing the flow of a known
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INFLUENT TO
HIXING PUMP
CENTER
BAFFLE
MIXING
PIPING
"^EFFLUENT
INFLUENT
INFLUENT TO
/INFLUENT
LULI* I  v	I	,
MIXING PUMP
MIXING
PIPING
fr1
CENTER
ISLAND
\W f \ll/
p n fT
EFFLUENT
8-0"
STANDARD POND	MODIFIED POND
FIGURE 5- MINI POND DESIGNS USED AT THE I.A.W.T.C.
volume and, if the meters were inaccurate, the glass tubes
and stainless steel floats were cleaned In a potassium di-
chromate-sulfuric acid cleaning solution and then recali-
brated. Nutrients were added to individual ponds or
storage pond. Technical grade sodium nitrate was added to
the storage pond to increase low summer nitrate c?ncentra
tions to near 20 mgN/1. Often fertilizer grade
acid was added to the storage pond, but when phosphorus was
being studied, it was added directly to the miniponds. The
phosphorous concentration for most of the runs wa	*
as P. Iron (PeCl3) was added daily to the growth unlts
when tested as a growth factor.
Except during special mixing studies, the miniponds
were mixed twice daily# from 0800 to 0830	e gtudv^the
and from 1200 to 1530 hours. During most of the istudy, the
miniponds were also swept	to	parhon di-
trated algal deposits caused by eddy current;s. ^niected
oxide (5 or 100 percent) and atmospheric air were injected
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into some ponds through the intake side of the mixing pumps
during the afternoon mixing cycle only. Bottled CO2 was
mixed with compressed air to obtain the desired concentration
of carbon dioxide. The true amount of CO2 injected was esti-
mated by recording changes in cylinder weight.
During some minipond runs in 1969, a settling tank;
consisting of a 50-gallon drum with an influent line entering
near the center of the drum was attached to one of the mini-
ponds, Influent water came from one of the nozzles of the
mixing system and a surface overflow returned the water to
the pond. The algal sludge was periodically removed by
draining from an opening near the bottom of the tank.
During 1968, Sacramento blackfish (Orthodon mlcro-
lepidotus) were added to some miniponds to test the possible
effect of fish on algal growth. The fish ponds with mixing
had a screen placed over the intake side of the mixing pump
to prevent the fish from being brought in the pump suction.
The California Department of Fish and Game used
two miniponds to determine whether Clostridium botullnum
might grow in this type of aquatic environment. One pond
received a one-inch layer of soil from an area with a known
botulism outbreak (Tulare Lake), while the remaining pond
received one inch of soil from the Treatment Center site.
Personnel from the Department of Pish and Game made periodic
visits to the Center to sample the two ponds for bacterial
analyses. These two ponds were also routinely sampled with
the other miniponds; however, the only controlled variables
were water depth (12 inches), P addition (2 mg/l), and de-
tention time. The ponds were neither emptied, mixed, nor
swept during the study period.
Rapid Growth Pond (RGP)
The rapid growth pond (Figure 6) is an asphalt-
lined pond with a 12.5-foot wide folded raceway approximately
800 feet long. The center baffles were originally constructed
of sheets of aluminum attached to both sides of a wooden,
upright frame; however, algae accumulated between the two
sheets and one sheet was removed. The RGP could be operated
at depths from 0.5 to 3 feet, with the effluent taken from
either the top or bottom of the pond. With its four avail-
able mixing pumps, velocities of 1 fps were theoretically
possible at any pond depth. This mixing velocity was con-
sidered necessary to achieve complete resuspension of settled
algae and periodic mixing and aeration of the bottom sludge
layer. Each pump had an individual timer which allowed an
almost infinite variety of mixing schedules.
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FIGURE 6-RAPID GROWTH POND USED FOR LARGE SCALE ALGAL GROWTH STUDIES
Influent from the storage pond was pumped into the
RGP semlcontlnuously throughout a 24-hour period. Detention
time was regulated by varying the length of time the influen
pump functioned during each specified timing interval an y
the flow of the pump. The effluent pump, also on a timer,
could be programmed to pump at a specific time or as a
function of pond depth. Recording flowmeters were attached
to both influent and effluent pumps to record	,
amount of water entering and leaving the pond. A recircuia
tion pump was available to introduce the pond culture into
the influent line.
Oneration of this growth unit resembled that of the
minlponds. The optimum levels of experimental variables
noted in minlpond runs were maintained as nearly ^Possible
in the RGP. Because of its uniqueness, the Pond as
mainly as a demonstration unit and as a source of algae lor
separation studies. During the 1969 operatlonoftheRCP,
daily changes often were necessary in mixing, sweeping, and
so forth, to provide algae for testing variou p
separation equipment.
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Algal Harvesting
Laboratory
The laboratory used in the harvesting studies was
separate from the chemical and biological laboratories des-
cribed previously. Standarized coagulation-flocculation
tests (jar tests") were conducted in this laboratory to
screen potential coagulants in the concentration step of
algal harvesting. The coagulants tested included such com-
pounds as lime (Ca(0H)2)j alum (Al2(S04)3) and ferric sulfate
(Pe2(SOi|)3)> as well as various polyelectrolytes. These
tests were used to estimate the effectiveness of these
different compounds for coagulation-flocculation before test-
ing in the larger, pilot-scale units. Routine tests of the
most promising test compounds, or combinations of compounds,
were also conducted over extended periods of time to deter-
mine possible seasonal variations in their effectiveness.
The experimental procedures used for these tests are out-
lined in the following paragraphs.
Suspensions of algae for the jar studies were ob-
tained directly from outdoor growth units, usually the rapid
growth pond. The suspensions had volatile solids concentra-
tions of from 50 to 800 mg/1. In making the daily compar-
isons of various chemical coagulants, a common supply of algae
was used. It was collected on the morning of the testing
period. The procedure for the jar tests was standardized
as follows: 800 milliliters of algae suspension were placed
In 1,000 ml "tall form" beakers and the calculated quantity
of reagent was added. The samples were then mixed by means
of a multlmixer at a specified paddle speed and time, depend-
ing on the type of coagulant used. The speed and duration
were 40 rpm for eight minutes with lime and 70 rpm for three
minutes with alum. These values were found to be optimum
for maximum flocculation. If polyelectrolytes were used in
conjunction with alum or lime, the conditions optimum for the
principal chemical were used. If polyelectrolytes were
tested alone, the mixing speed and duration most effective
for alum were employed, although mixing speed and duration
were optimized for the most promising polyelectrolytes.
After the mixing operation, the beakers were placed in a
darkened area of minimum air circulation. At the end of one
hour, the sample supernatant was decanted and a spectro-
photometer was used to determine the percent transmittance
of the supernatant at 410 m/j.
During the first part of these laboratory studies,
the amount of algal biomass (and other suspended solids)
removed by coagulation-sedimentation was determined by using
a few random samples to establish the relationship between
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weight of biomass in the flocculated material and percent
transmittance (at MlO m/i) of the supernatant. P..
these values was then used to estimate rnloYa3;j	. ..
manning samples. This method proved to be tedius and it
was modified in the following manner. The 3usp<3Jed .olids
and percent transmittance were determined for a
algae-laden water from a growth unit. The sample was acidi
fied with two drops of 6N H2S04 to remove precipitated
calcium carbonate. Several quantitative dilutio .
made of the original pond sample and the percent transmit
tance measured for each of the acidified dilutions.
calculated suspended solids versus the recorded transmittance
for each dilution were then plotted. A typical curve is
shown in Figure 7. The
percent transmittances
of the acidified super-
natant from the floccu-
lated samples were then
determined, and the
removal percentage or
total suspended solids
remaining were then
obtained from the
dilution curve.
TOTAL SUSPENDED SOLIDS - tn/
uj 80-
z 20-
Chemlcal anal-
yses occasionally per-
formed on the supernatant
included nitrate, nitrite,
organic nitrogen, ortho-
phosphorus, pH, hardness,
calcium, and magnesium.
All analyses were per-
formed as outlined in the
section on growth, except
that calcium and total
hardness were determined	. titrametric
by the	(ethylenediaminetetraaceti^ dlf|erenee between
method, with magnesium
the two ions.
40	60	80	100
PER CENT REMOVAL
FIGURE 7 - TYPICAL DILUTION CURVE
USED TO DETERMINE TOTAL SOLIDS FROM PER CENT TRANSMITTANCE
Pilot-Scale Separation Stales
orminment used in these studies, which re-
?eadily available for lease or rental, dic-
5 t	JLnredures that were followed. (Nothing in
5  i testing	jg intended as an endorsement of any
of the units and testing
procedures are^contained in the following paragraphs.
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Sedimentation Tank. A rectangular sedimentation
tank was borrowed from the Los Angeles County Sanitation Dis-
trict and was used in the removal of algae and sediment from
the rapid growth pond. The unit was approximately 4 feet
wide x 6 feet deep x 22 feet long with a capacity of about
4,000 gallons. Water was pumped from the RGP to the bottom
of the tank and effluent was withdrawn from near the top and
returned to the growth unit. The settled algal slurry was
moved to one end of the tank by blades attached to a continu-
ous chain and was periodically discharged onto drying trays.
Detention time was the only controllable variable.
Shallow Depth Sedimentation Tank. This was a self-
contained water treatment plant called the "Water Boy"
obtained from Neptune Microfloc, Corvallis, Oregon. The
Water Boy was the only unit consistently operated on-line
with the RGP. The plant treated a portion of the pond ef-
fluent and discharged its waste from the site. A schematic
flow diagram of the unit is shown in Figure 8. As illus-
trated in the diagram, the unit consisted of a flocculation
chamber, a settling chamber, and a mixed-media filter.
CHEMICAL
ADDITIONS
INFLUENT-

FLOCCULATION
CHAMBER
do
,/SETTLING^
XZCHAMBER/^
MIXED
MEDIA
FILTER

400 GAL
BACKWASH
STORAGE




EFFLUENT
FIGURE 8 - SCHEMATIC AND FLOW DIAGRAM OF WATER BOY SHALLOW DEPTH SEDIMENTATION UNIT
In the flocculation chamber the algae-laden water was mixed
with a coagulant (or coagulants) in concentrations determined
by jar test studies, and from there passed to the settling
chamber. The chamber differed from the one described in the
preceding paragraph in that it was equipped with a module of
settling tubes inclined at an angle of 7 upwards in the
direction of flow. The theory behind this type of sedimenta-
tion unit has been described by Hansen, et al (1969). Essen-
tially, the settling tubes reduce partlcli" settling distances
and thus lower the time required for sedimentation (i.e.,
detention time). Figure 9 illustrates the effect of tube
inclination on settling distances of discrete particles.
(Diagram and following description are taken from Hansen,
et al, 1969).
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"The path traced by
a particle settling in a
tube is the resultant of
two vectors: V, the ve-
locity of flow through
the tube, and vg the
settling velocity of the
particle. It can be
seen in Figure 9 that if
the settling surfaces
the d^ecWonUS?a??oi? on settling path of suspended part,cle
the settling path of the
particle is altered be-
cause the component of
the settling velocity
which is parallel to the
tube wall, v^, is opposite in direction to the ve-
locity vector v. If V is greater than vs, the required
length of the settling surface decreases as the angle
increases from zero up to about 25 to 30 deg. (at V =*
2.5 vs) and then increases, approaching infinity as the
angle of inclination is increased to 90 deg. For V <
vs the tray length continues to decrease with increasing
angle. "
Use of the tubes also reduces turbulence and short-
circuiting, thus promoting laminar flow of the water. With
periodic flow reversal, or drainage, the inclination of the
tubes facilitates removal of the algal floe. The coagulation-
sedimentation step was designed to remove most of the sus-
pended solids; however, a mixed-media filter provided a
final polishing of the effluent. After passing the filter,
most of the clarified water went to discharge with only a
small portion being stored in the baclcwash tank for back-
washing the filter. The settled algal suspension from this
unit normally was discharged into a holding sump to provide
material for study of dewatering equipment.
The Water Boy was used only to remove algae from
growth pond water, that is, the concentration step.
Controllable variables included hydraulic detention time,
type and amount of flocculate, and frequency of sludge re-
moval. The unit, as tested at the IAWTC, required manual
operation of the filter backwash and sludge removal cycles.
Upflow Clarifies- In the upflow clarifier con-
structed at the site "{Figure 10) a coagulating aid, usually
sodium hydroxide, was added to the influent algae-laden
water from the RGP. The mixture then entered the apex of an
FIGURE 9 - EFFECT OF TUBE ANGLE
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inverted cone and
flowed through a
thin diffuser layer
to mix the. coagu-
lant and algae-laden
liquid. The algal
floe flowed upward
and, as the cross-
sectional area in-
creased, the veloc-
ity decreased,
causing the floe to
settle. A mat of
algal floe formed at
the point at which
the upward velocity
of flow equalled
the settling veloc-
ity of the floe.
The algal sludge was
drawn off from the
mat, and the clari-
fied liquid passed
off at an upper
level. Operational
variables were de-
tention time and
concentration of co-
agulant,.
Upflow Sand Filter. A Sanborn filter was construc-
ted for the algae stripping project by Bohna Engineering and
Research, Inc., of San Francisco, California. The principle
of operation for this unit is illustrated in Figure 11. A
description of the principle is quoted from an unpublished
Bohna internal report dated February 6, 1970:
"The fluid to be filtered is pumped into a
feed chamber at the base of the filter from where
it flows upward in a thin channel called the
feed wick which is closed at the top. The fluid
then flows horizontally across a polypropylene
cloth surface followed by a thin layer of sand.
A second layer of cloth backs up the sand and
allows the clarified liquid to enter a second
channel called the drain wick from where the
filtrate flows upward into a backwash-holding
tank and then to final disposition.
3fc-EFFLUENT
SLUDGE
SLUDGE MAT
DIFFUSER(WIRE MESH WITH
PIECES OF PVC}
INFLUENT
CHEMICAL
ADDITION
FIGURE 10-SCHEMATIC DIAGRAM OF UPFLOW CLARIFIER
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CLOSED IN
BACKWASH
CLOSED
DUMP VALVE-
OPEN IN BACKWASH
BACKWASH CYCLE
EFFLUENT
OPEN IN FILTRATION
BACK
WASH
HOLDING
TANK
NOTE:
ARROWS INDICATE
FLOW DIRECTION
FILTER
CLOTH
SAND OR OTHER
GRANULAR MEDIA
FEED
DISTRIBUTION
CHAMBER
INFLUENT
DUMP VALVE-
CLOSED IN FILTRATION
FILTRATION CYCLE
FIGURE 11-SCHEMATIC DIAGRAM AND PRINCIPLE OF OPERATION OF SANBORN FILTER

-------
"In backwashing, the overflow effluent valve
and feed valves are closed and a dump valve at the
base of the feed chamber is opened instantaneously,
allowing the hydraulic head to force filtered water
down the drain wick, through the sand and out the
dump valve, thereby removing the accumulated solids
from the filter cloth and sand bed."
This filter was operated on water directly from
the rapid growth pond, without any chemical pretreatment,
and the filtered effluent was discharged from the site.
Most of the data for this unit were obtained during two
intensive studies with supervisory help from Bohna, Con-
trollable variables included influent flow rate, backwash
volume, and the pressure differential across the filter
media at which the unit was backwashed.
Mlcroscreen. An automatic rotating drum filter
leased from the Zurn Company consisted of a 2-foot wide,
rotating drum having a diameter of 4 feet and mounted in a
self-contained unit. Algal-laden water entered the interior
of the partly submerged drum and was filtered as the water
flowed outwards through a screen attached to the revolving
drum. Screens of 25 and 35 micron mesh size were supplied
with the unit. Algae caught on the screen were washed off
by a row of water jets located above the drum; and the
algal slurry then dropped into a trough located inside the
drum, above the water level. The microscreen was tested
primarily as a concentrating device, but also as a means to
dewater an algal slurry from the Water Boy. The major vari-
able tested was the amount of solids loading per unit area
of screen.
Vacuum Filter. A continuous belt vacuum filter
was leased from the Eimco Corporation to examine its feasi-
bility both as a concentrating and as a dewatering device.
A schematic diagram of the unit is shown in Figure 12. The
filter media, a continuous porous belt, was immersed in the
filterable material. A vacuum applied on the inside of the
drum caused the suspended particles to form a cake on the
belt. As the belt passed over the discharge roll, a cake
deflector scraped off the cake. The belt was then washed
as it passed back into the feed tank.
The influent algal suspension for this unit con-
tained from 0.3 to 3 percent solids. The number of possible
experimental variables was quite large and included flow,
algal concentration, filter material, drum speed, and amount
of vacuum. With the exception of drum speed, all these
variables were tested at the IAWTC.
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FILTER MEDIUM
FILTER DRUM
RETURN
ROLL-
DISCHARGE
? ROLL
DEFLECTOR
s\ 4 WASH
"SCr HEADER
VALVE
WASH
ROLL
WASH
TROUGH
SEPARATE DISCHARGE TO
DRAIN OR RECYCLE
	_____	"^-J-FEEO TANK
FIGURE 12- PRINCIPLE OF THE BELT FILTER
npntrifucation. Three types of centrifuges were ^
investigated as possible means of primary concentration and/
or dewaterine The first of these units was a Bird Machine
Company continuous flow, solid bowl centrifuge consisting o
two concentric rotating elements inside a stationary housing.
The inner element was a scroll-like unit rotating a a
slightly higher speed than the outer element. Variables
tested, using both straight pond water and a	,n
slurry, included flow, centrifuge speed, and liquid depth in
the bowl.
The second variety of centrifuge was a De Laval
yeast-type separator built by the De kaval
which was designed for continuous feed and removal with
solid/liquid and solid/liquid/liquid extraction. Jentrif
ugal force in this type of unit is constant and
of solids is controlled by varying the ;size
nozzles. The third centrifuge was a De Laval
continuous flow unit. *n.the last-named type o c	g 
suspended solids accumulate on the side of the bowl until
they reach a predetermined level and then are ^Jha^^has
the bowl opens. Once the solids have been ejected, which
occurs in only a matter of seconds, the bowl closes and
normal operation continues. During opening and closing of
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the bowl, bowl speed remains unchanged. Experimental vari-
ables In testing this unit were feed composition (used as
both a concentration and dewatering device), throughput
(flow), length of time the bowl was open, and amount of bowl
discharge.
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CHAPTER IV - RESULTS AND DISCUSSION
Data are presented in this section which were ob-
tained in algal growth and nitrogen assimilation har-
vesting studies at the Treatment Center. Also included are
the data of two other studies which accompanied Jlgf*
growth  regrowth and botulism. The regrowth s^dies were
concerned with the effectiveness of treatment methods for
removal of those substances from agricultural
which promote or "stimulate algal growth. In ^he
work, the research was designed to evaluate the possibility
of botulism outbreaks in algal growth units.
Algal Growth
The investigation of algal growth was divided into
three levels of interest  lightbox, miniponds, and rapid
growth pond. The major emphasis in these studies was to
determine the effect of various factors on the rate at which
nitrogen is assimilated by algae in laboratory and outdoor
cultures. To achieve continuity of results in these studies,
nutritional and physical factors were made most favorable
for the growth of fir.enedesmus. It should be pointed out
that there are other aigae which are probably equally suit-
able for nitrogen removal. The last part of this section on
algal growth will provide some general information on vari-
ous biological, chemical, and physical factors recorded
during the course of the investigation.
In this section many of the results will be re-
ported in terms of nitrogen assimilation, expre:ssed either
as mg/l or percent. In general, "nitrogen assimilation is
expressed or measured as the amount of soluble nitrogen
disappearing (i.e., removed) from a medium as a result oi
algal growth. This method of expressing nitrogen fssimila-
tion is a simplification of the true system found ^ algal
cultures where growth and decomposition are occu^"f 1a p
taneously and a constant flux of nitrogen forms takes place.
For example, in most cultures dissolved organic ""rogen
increased with culture age, probably from ext^acelluiar ex
cretion or cellular decomposition. As defined in tni
report, this would be considered to be	^.1
gen. Nitrogen balances obtained by determinations of both
influent and effluent values indicated that nitroJ*
through anaerobic denitrificatlon and release as nitrogen
gas was not significant.
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Lightbox Studies
Laboratory cultures of Scenedesmus were used to
determine optimum nutritional requirements of the algae, the
effect of temperature on algal growth, possible effects of
seasonal variation in mineral quality of the tile drainage,
and comparisons of algal growth and nitrogen assimilation in
tile systems other than the system used at the Center. In
addition to these studies, the lightbox was used to evaluate
the growth rate and nitrate uptake in tile drainage of algal
species other than Scenedesmus. Because of the large numbers
of flasks involved in the lightbox culture studies, nitrate
was the only nitrogen form determined routinely. Occasional
analyses for other forms indicated that nitrite and ammonia
usually were present in amounts less than 0,10 mg/l and
organic nitrogen in amounts ranging from 0.5 to 0.8 mg/l.
Changes in nitrate were thus valid indicators of the effect
of a variable on total nitrogen assimilation. The results
of the batch culture studies were then used in outdoor
cultures to provide optimum growth conditions for the algae.
The results of these studies are detailed in the following
paragraphs.
Nutrient Requirements. Batch assays were first
used to determine the necessary nutrient additions for opti-
mum algal growth and nitrogen assimilation in the tile drain-
age. Because the ratio of nitrogen to phosphorus (p) in
Alamitos sump water is about 100:1 (as compared to approxi-
mately 10:1 in the algal cell), phosphorus was the first
element tested. Figure 13 illustrates results of a typical
experiment which compares 0 and 2 mg/l P addition to
Scenedesmus cultures. In this particular study, unconverted
nitrate meter readings, which are proportional to nitrate
concentrations, were plotted instead of actual nitrate values.
The meter readings were often used with the assumption that
relative changes were a reliable indication of a variable's
effect. At the end of five days, the culture to which no
phosphorus was added, remained at, or near, the initial
nitrate level; whereas, the culture containing 2 mg/l addi-
tional phosphorus showed a 50 percent reduction in dissolved
nitrate.
In general, all lightbox studies indicated that
little or no growth occurred without supplemental phosphorus.
The 0.2 mg/l originally present in the tile drainage water
was not adequate for attaining the blomass levels of
Scenedesmus cultures required for maximum nitrogen assimila-
tion. Allsubsequent laboratory cultures received 2 mg/l
PO4-P, unless phosphorus was one of the experimental
variables.
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NO P04 ADDED^tn^
uj 150-
2 mg/< PO4-P ADOEO
uj 100-
NQTE-- AERATED, CONTINUOUS LIGHTING
DAYS AFTER INOCULATION
FIGURE 13 - EFFECT OF PHOSPHOROUS ADDITION
ON NITROGEN ASSIMILATION
After the neces-
sity for adding phosphorus
had been established, fur-
ther experiments were con-
ducted on the effect of
additional iron on Scenedes-
mus cultures. A typical
example of this type of
assay is shown in Figure 14
where FeClo was used as the
source of iron and the
lighting cycle was set for
12 hours light and 12 hours
darkness. The combination
of 2 mg/1 P plus 3 mg/l Fe
led to an increase in the
rate and amount of nitrate
assimilation. Experiments
with different concentra-
tions of Fe Indicated that
3 mg/1 (from FeClo) produced
nearly optimum results. Two
additional iron compounds,
ferrous sulfate (FeSOty) and
ferric citrate (FeCgH^Ov)
were tested. The results
of the test showed that
while both were adequate
sources of Fe, optimum con-
centrations differed. With
the use of FeSOi*, the req-
uisite concentration was
similar to that of FeCl3>
or 3 mg/1 Fe. However,
when ferric citrate was
used, about twice as much
iron (as Fe) had to be
added in order to achieve
the same rate of nitrate
assimilation. The use of a
chelating agent, the sodium
salt of EDTA, decreased the
concentration of iron re-
quired. For example, in
one study 3.2 mg/l Fe alone
was as effective as 0.4 mg/l
Fe combined with 5*0 mg/l EDTA addition. EDTA alone also
resulted in an increase in nitrate assimilation, presumably
by making the small amount of iron in the sump water avail-
able for algal growth. Cost analyses of the use of EDTA in
large-scale algal cultures indicated that the use of this
30-
 20-
I
o
z
z
<
5
UJ
oc to
z

K>
o
z
NOTE : NOW AERATED
LIGHT 12 ON-12 OFF
-2mg PQ4-P
N2mgPO4Fe (FeCI3)
30
10	20
DAYS AFTER INOCULATION
FIGURE 14- EFFECT OF IRON ADDITION ON NITROGEN
ASSIMILATION BY SCENEDESMUS CULTURES
-39-

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compound was not economically practical. After several rep-
lications of the iron studies, 3 mg/1 of Fe (from FeCl3) was
routinely added to lightbox algal cultures.
Based on theoretical considerations, Alamitos sump
water did not contain enough carbon to support the growth
of biomass required to assimilate 20 mg/1 nitrate-nitrogen.
This deficiency indicated that CO2 addition to laboratory
cultures could enhance nitrogen assimilation by Scenedesmus.
Studies were then initiated using a mixture of 4 percent
CO2 in compressed air, compresed air only, and surface re-
aeration (flasks hand-swirled twice daily). The results of
one of the experiments are illustrated in Figure 15, which
shows that the rate of nitrate uptake was significantly
increased by the addition of
of 4 percent 002* an(3 that
compressed air also was an
effective means of provid-
ing some additional carbon.
The pH values were about
7.2 to 7.3 for the 4 per-
cent C02* 9.6 to 9.8 for
the compressed air, and
10.0 to 10.5 for the swirled
cultures. On two occasions,
various amounts of sodium
bicarbonate were added to
inoculated sump water to
explore the possibility of
using this compound as a
carbon source of nitrogen
assimilation. These cul-
tures were then compared to
other cultures receiving
4 percent C02. In both of
these studies, the addition
of bicarbonate did not en-
hance nitrogen assimilation.
conclusively shown to benefit algal growth in the cultures,
in almost all of the subsequent experiments at least two
levels of aeration were applied, namely injection of 4 per-
cent CO2 and diffusion of atmospheric CO2 by swirling the
flasks twice daily. The two levels of CO2 addition were
used to make the data more applicable to outdoor cultures,
some of which received additional inorganic carbon.
The three levels of nutrients  Fe, P, and C0p -
were then combined in a factorially designed experiment to
determine their effect on nitrogen uptake by Scenedesmus
cultures. Five levels of Fe at three levels of P were
compared in flasks that received either 4 percent C02 or


30-]
20-
<
2
UJ
cr

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compressed air. In addition, five levels of Fe were tested
in cultures to which 2.0 mg/l p had been added and which
were swirled twice daily. The nitrate remaining in the var-
ious cultures is shown in Figure 16. These data indicate
that interaction occurred between the three variables tested,
and that in general, the addition of CO2 and Fe increased
the rate of nitrate assimilation. The level of iron addition
which provided the maximum effect was about 3 mg/1 of Fe.
The minimum requirement for complete nitrate assimilation at
11 days was 4 percent C02> 2 mg/l P, and 3 mg/l Fe. The
addition of higher levels of Fe and P probably resulted in
	SWIRLED TWICE DAILY
		COMPRESSED AIR
	4% C02 IN COMPRESSED AIR
I	2 mg/l PO4-P
i	4 mg/l PO4-P
1	8 mg/I PQ4-P
NOTE: LIGHT 12 ON-12 OFF
20-)
	O
-A
Fe ADDITION mg/l
figiibf IR-NtTRATF-NITROGEN REMAINING AT DAY II IN SCENEDESMUS CULTURES
FIGURE 16 N^tRhAT^rN,G|0NMBINATI0NS OF F., P. AND CARBON DIOXIDE
the formation nf an unavailable iron-phosphate complex. By
day leTlll'ae?ateS cultures 
-------
100 percent CO2 was never found to be more effective than
4 percent C02 and often was detrimental to algal growth,
perhaps because pH values were less than 6. This value
falls below the optimum range of 7 to 8.5 for Scenedesmus.
Oswald (1963) found that about 0.5 percent CO2 addition to
mixed cultures of Chlorella and Scenedesmus was most favora-
ble for maximum light conversion efficiency.
Possible seasonal variations in iron and phos-
phorous requirements resulting from changes in the influent
water quality were checked by conducting periodic laboratory
studies in which cultures received various concentrations
of these elements. These tests were normally run when the
storage pond was being refilled. In all but one of these
studies, 2 to 4 mg/l of both elements were necessary for
maximum nitrogen assimilation. In the one exception iron
was not required. Also included in many of these checks
were comparisons of nitrate assimilation rates in sump water
and storage pond water. Little or no difference was shown
between the two waters, provided that both had comparable
amounts of N, P, and Fe. In conducting this type of study
throughout the year, seasonal differences were noted in
nitrogen assimilation rates in those cultures receiving
the optimum nutrient additions. Although these differences
may be attributable to variations in inocula, they could
have been the result of changes In water quality. This
possibility will be discussed more fully in the section on
comparison of water from various tile systems.
On several occasions trace minerals were added to
the cultures to determine their effect on algal growth.
These elements seldom had any significant effect on nitrogen
uptake, either individually or in commonly used trace element
solutions. The few instances in which a beneficial effect
was noted, the results could not be duplicated in further
experiments.
Temperature. A preliminary study on the effect of
temperature on nitrogen uptake indicated that, of the three
temperatures first tested (10C, 20C, and 306C), 20C was
optimum for nitrate assimilation, and that Fe and P were
necessary at all temperature levels. At 30 C, the initial
Scenedesmus culture was replaced by a blue-green alga,
Osclllatoria. The 10C cultures had an extremely long lag
phase and did not enter the exponential growth phase until
near the end of the study. Therefore, another experiment
was designed in which temperatures of about 15C, 20 C, and
-42-

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30C were used. The apparatus used to maintain the various
temperatures did not permit testing the effect of 10C at
the time of the test. This second study differed from the
first in two important respects  (l) three levels of CO2
addition (none, 4 and 100 percent) were tested at all
temperatures and (2) all of the cultures were allowed to
grow at 20C for four days before being subjected to the
various experimental temperatures. (In the preliminary
study, all culture flasks had reached the experimental
temperature when they were inoculated.) When the cultures
had assimilated all the nitrate in the flask, an additional
20 mg/l of nitrate-nitrogen was added along with 2 mg/l P
and 3 mg/l Fe. Growth of Scenedesmus at the high tempera-
tures followed the same pattern as that observed in the
previous study -- rapid initial growth, then dying away,
with eventual replacement by Qscillatorla. This species
change was noted when either no CO2, or 4 percent C02 was
added but not with 100 percent C02. The use of pure C02 at
all temperatures gave the same result: namely, a pale
culture of apparently viable Scenedesmus which did not grow.
The lack of growth may have been due to.the low pH
of these cultures (5.7 to 5.9). The 15 c cultures exhibited
a. surprisingly high rate of nitrogen uptake, especially with
4 percent CO2 addition. Table 5 summarizes the total
nitrate assimilated by the different cultures. With the
exception of the 100 percent COg cultures, the only combina-
tion of variables that produced a notable effect was
4 percent CO? concentration at a temperature of 15 o. ine
inoculum used in this study was obtained from outdoor growth
units which had been at a temperature of about 15 ^ r
several days. Perhaps the period of acclimatization in the
laboratory was not long enough for complete adjustment to
the experimental temperatures.
TABLE 5
TOTAL MILLIGRAMS NITROGEN ASSIMILATED BY ALGAL
CULTURES AT DIFFERENT TEMPERATURES
AND CO2 ADDITIONS
Temp.
15
20
30
Percent C02^dded"
11
16
11
32
IT
11
1
3
1
-43-

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Algal Species. During the spring of 1969, unlalgal
cultures of 30 species of algae were obtained from the culture
collection of the University of California at Davis. These
algae were then cultured in drainage water, with and without
P and Fe additions, and their rates of nitrate uptake compared
to that of Scenedesmus from the outdoor growth unit. Of the
species tested (including green, blue-green, and diatoms),
six grew well in sump water plus nutrients, and one grew in
the P-deficient medium. The six species included three green
algae -- Ankistrodesmus, Eucapsls, and Gleocapsls; and three
blue-green algae Anacystis, Oscillatoria, and Anabaena. By
far the best growth and nitrogen assimilation were noted in
Ankistrodesmus and Anacystis cultures; however, growth and
nitrogen assimilation were not appreciably better than those
of Scenedesmus. With no apparent advantage to be gained from
using these algae, no concerted effort was made to develop
large-scale cultures of the species tested.
Water Source. In a study of this type, where the
water comes from an Isolated system, one of the questions
raised concerns the applicability of the data to an area-
wide system. Although such combined drainage was not avail-
able during the project, samples of other tile systems were
collected and tested for their algal stimulatory potential.
As the partial chemical analyses of the samples shown in
Table 6 indicate, the sumps differed considerably in nitrate
and electrical conductivity (EC). The differences in nitro-
gen concentration were nullified by bringing all sumps to
about 55 mgN/1 as nitrate. Part of this study was performed
to determine whether or not iron and phosphorous additions
were needed to produce the algal biomass required to assimi-
late the nitrogen in each of these waters. The results
showed that both elements were necessary and that there was
TABLE 6
CHEMICAL ANALYSES OF VARIOUS SUMPS
Concentration - mp;/l
Total KJeldahl|N03-N|N02-W[P81t-P|Total Fe
Sump
umho/cm^
DPS 1367
HMV 7016
GSY 0855
BVS 7402
(Alamitos)
5890
0.28
45
3880
0.30
55
8100
0.25
16
5200
0.40
13
0.003 0.06	0.03
0.003 0.00	0.04
0.004 0.00	0.07
0.006 0.038	0.05
-44-

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little or no algal growth without their additio	mefcer
The curves in Figure 17 show changes in
readings with time for two of the sumps:
7402 (Alamitos). Data from HMH 7016 and DPS 1367 were similar
to those of Alamitos and were omitted from the S^aph for
reasons of ease of reading. The curves illustrat
differences in the rate of nitrate uPta^e, with the ;alg;3
in GSY 0855 water removing all the original nitrogen in
7 days. The algae grown in the other drainage w	-ration
about 12 days tS remove the 55 mgN/1. As the
of nitrate in a flask reached zero, an additional1 20 iwater
N03-N were added. In this particular study GSY 0855 water-
was "respiked" three times, and the cultures from
three sumps only once. By the end of day	5
growing in GSY 0855 water had asslrn^la5dmJ1	assimi-
milligrams NO3-N, as compared to only 60	* q855
lated in the Remaining sumps. The celis grown in aSYO55
tile drainage were practically all one-celled Scenedesmus,
whereas a considerable number of Oscilla or
appeared in the other water samples.
ORIGINAL SAMPLES
RESPIKED SAMPLES
o 6OO-1
ALAMITOS (BVS 7402)
GSY 0855
NOTE: 4% CO,
400H
3mg/l Fe
CONTINUOUS LIGHT
200-1
lL
20
DAYS AFTER INOCULATION
FIGURE 17-comparison of nitrogen assimilation by scenedesmus
CULTURES USING WATER FROM THREE DIFFERENT TILE
DRAINAGE SYSTEMS
-45-

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The water for the previous study was collected in
early September when sump flows were characteristic of sum-
mer irrigation practices, especially rice flooding. Another
series of samples from the same tile systems was collected
in October when field irrigation had been discontinued. The
plots of nitrate meter reading versus time for cultures grown
in the three tile drainages are snown in Figure 18. Complete
nitrate assimilation in all the sumps required about 12 days
or about the same length of time required by cultures in
DPS 1367, HMH 7016, and BVS 7^02 water in the previous study.
Results using water from GSY 0855 were similar to those from
the other sumps. Algae grown in a composite sample from the
four systems to which iron and phosphorus had been added but
which contained only about 30 mgN/lj assimilated essentially
all of the nitrogen by day 7. The difference in time needed
to reach zero nitrate between the composite and the individual
sumps undoubtedly was caused by the original difference in
nitrogen content of the samples. The individual drainage
samples were all brought up to about 55 mgN/l as nitrate.
NOTE:
5% C0Z
2	mg/ I PO4-P
3	mg/1 Fe
CONTINUOUS LIGHT
More studies of this type are needed for confirma-
tion; however, seasonal variation in the algal growth poten-
tial of individual tile systems does occur, at least in labo-
ratory cultures. The water used in this investigation
appears typical
of other systems,
and the nutrient
additions re-
quired for growth
of Scenedesmus
probably willbe
required in a
combined system.
Miscel-
laneous Llghtbox
Studies. Addi-
tional lightbox
studies were de-
signed to answer
specific questions
about occurrences
In the outdoor
growth units. The
results of these
studies will be
presented in later
discussions.	n 5	4 e
DAYS AFTER INOCULATION
FIGURE 18-COMPARISON OF NITROGEN ASSIMILATION
IN VARIOUS TILE DRAINAGE SYSTEMS - OCTOBER SAMPLE
6OO-1
500
400-
uj 300-
200-
I OO-
PS 1367
ALAMITOS
(BVS 7402
HMH 7016
MIXTURE
-46-

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The use of laboratory batch assays Jf"
valuable information about such Items as nutri	various
ments, desirable algal species, and comparison .	tlnff
waters. However, some discretion is needed in .^
the results of such studies, especially with r^P	.
nutrient additives. Because environmental conditions :in
laboratory cultures are near optimum, S^ov,th r_
and the effect of minor elements can ^e" b d^*^Lre 
Outdoor cultures are often limited by ^S^t or P noten-
therefore, the algal cultures do not ac^leve.m!lJ1^w^fc n
tial growth and the effect of a minor element ^masked.
This condition appeared to be especially tr	orac-
element necessary for maximum nitrogen assimilation in priac
tically every lightbox study but only seasonally required in
outdoor cultures.
Minipond Studies
The miniponds were con^ructed during Listed*!^
of 1968 and were placed in operation that May. fisted in_
Table 7 are the minipond runs, their d f, t'three runs
spective areas of emphasis. During th	until
operational Problems,we^efl^e^ntrSl of'the ponds was achieved,
fall that adequate physical "krol o	before September
Because of this, results of	This section deals
1968 will not be emphasized in ^hi P *hvslcal ana bio-
with the effect of nutrient additions and physical and dio
logical parameters on the uptake of
n 4 4-	Five nutrients which, based on
TtF1S44gimFdrainage, theoretically limited
the composition of e tile drigrowth units -- phos-
algal growth were added to the out ^ rese and potas-
phorus, iron, inorganic carbon 100^;, Addition of the
alum. Of these five nutrients, 01nly lt en assimilation,
first three enhanced algal row^J*"strabie effect. With all
Manganese and potassium had no deJ	maintain a culture of
these additions, an attempt was made to	in	_
Scenedesmus quadrlcauda. The specific eiiecra
ent are described in the following P 6
Phosphorus. Preliminary
had indicated that the Phospboro^al growth. In the first
drainage water was limiting to aLf~xJ? weeks duration, the
three minipond runs, all of four to six *aluated# The
effect of four levels of P addition wio-ure 19, demonstrated
results of these studies, shown m Fig tdQor cultures of
that phosphorus also was necessary in centratlon Was not
Scenedesmus. The optimum P^osP^orou oval lncreased with
determined. In runs 1 and 2, hi^rate ever^ ln run 3#
phosphorous additions up to ^m6/* r* phosphorus had been
only the cultures to which 0.5 mg/1 o P ^ control which
added assimilated more nitrogen than	these three runs,
contained no additional phosphorus. Durxi &
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TABLE 7
MINIPOND RUNS CONDUCTED AT
THE IAWTC, 5/15/68 to 12/31/69
Run No. 	Dates	;
1	5/15 to 6/28/68
2	7/1 to 7/23/68
3	8/5 to 9/17/68
4A	9/17 to 12/7/68
4B	12/8/68 to 1/25/69
5	2/10 to 4/7/69
6	4/16 to 6/27/69
7	6/27 to 7/29/69
8	8/8 to 9/29/69
9A	10/4 to 11/19/69
9B	11/19 to 12/31/69
Main Areas of Study
Phosphorous addition, depth,
detention time, fish, soil
ponds
Phosphorous addition, depth,
detention time, fish, soil
ponds
Mixing, P addition, detention
time, depth, fish, soil
ponds
Mixing, fish, soil ponds
Mixing, fish, soil ponds
Mixing, depth, detention
time, soil ponds, fish
C02 addition, Pe addition,
depth, soil ponds
CO2 addition, Fe addition,
biomass control, detention
time, depth, soil ponds
CO2 addition, Pe addition,
biomass control, detention
time, depth, soil ponds
COg addition, Fe addition,
biomass control, detention
time, depth, soil ponds
CO2 addition, Fe addition,
biomass control, detention
time, depth, soil ponds
-48-

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influent flows were not
regulated with the desired
accuracy; therefore, the
contradictory results in
the third run were assumed
to be caused by opera-
tional problems.
Beginning in the
fall of i960 and through
July 1969, approximately
2 mg/1 P (added as
H3PO24) were added to the
storage pond. During the
remainder of 19&9, all of
the miniponds except one
received daily P additions.
Total nitrogen assimila-
tion for the control pond
in one minipond run and a
pond at the same depth,
detention time, and so
forth, are shown in
Figure 20. The original
inoculum for both of these
ponds had been grown in
a phosphorus-rich medium
(drainage water plus
2 mg/1 P), and laboratory
analyses of the filtered
inoculum indicated that
the residue contained
about 4 percent phosphorus.
Additional analyses of
filtered and unfiltered
pond samples indicated that
a substantial portion of
the phosphorus in the resi-
due was composed of col-
loidal precipitates caused
by high in-pond pH values.
The data in Figure 20 show
that for a two-month period
the nitrogen assimilation
in the non-P pond was con-
sistently 10 to 20 per-
cent lower than that in the
pond receiving supplemental
phosphorus. By the middle
of December, this dif-
ference had increased to
loo-.
RUN I
RUN 3
. 40-
PO4-P ADDITION (mg/1)
FIGURE 19 - THE EFFECT OF PHOSPHATE ADDITION
ON NITRATE ASSIMILATION IN OUTDOOR GROWTH UNITS
100 -r
C 50
2mfl/l P04-P
NO ADDED	! \
P4 \	>
s /	x
s /
V
OCT
DEC
JAN
FIGURE 20-COMPARISON OF NITROGEN ASSIMILATION IN MINIPONDS
WITH AND WITHOUT THE ADDITION OF 2mg/1 PO4-P
-49-

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30 to 40 percent, perhaps
because all excess cellu-
lar phosphorus, and all
precipitated phosphorus
in the pond, had been used.
The phosphorus-deficient
cultures were pale green
and tended to settle more
rapidly than those re-
ceiving P. Microscopi-
cally, the algae seemed
to be viable although
they lacked the dark green
color of typical healthy
cells.
Iron. In almost
all of the lightbox
studies, the addition of
2 to 4 mg/1 iron was neces-
sary for maximum nitrogen
removal. Results of the
first test of this element
in the miniponds, per-
formed in the spring of
1969, indicated that sub-
stantially more nitrogen
could be removed by cul-
tures with 6 mg/1 Pe added
(using PeCl3 as the
source) than by controls
with no Fe added
(Figure 21). The increase
in removal amounted to
about 3 mg/1 N at the de-
tention times tested. The
iron-supplemented cultures
were characterized by a
dark, luxuriant green
color, and microscopic ex-
amination of a pond sample
showed large numbers of
empty cell walls, indi-
cating rapid cell division.
As in the lightbox cultures,
the use of EDTA seemed to
decrease the amount
6 mq/l Fe
0 mg/1 Fe
8-1		1	r	
0	5	10	15
DETENTION TIME (DAYS)
FIGURE 21 - EFFECT OF FE ADDITIONS ON ALGAL NITRATE
ASSIMILATION IN OUTDOOR GROWTH UNITS IN LATE SPRING
TOTAL N
14-
2 12-
CO
8
10-
z
IRON ADDITION (mg/1)
FIGURE 22-EFFECT OF FE ADDITION
(3mg/0 ON ALGAL NITROGEN ASSIMILATION
IN OUTDOOR GROWTH UNITS IN LATE SUMMER
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of iron required for maximum effect, but its use was discon-
tinued because of the dissolved organic nitrogen contributed
by the compound. (Since EDTA contains about 10 percent
nitrogen, the daily addition of 25 mg/1 EDTA resulted in an
increase of about 2 mg/l in effluent dissolved organic
nitrogen.) Wide use of this chelating agent in an operating
treatment plant probably would not be economically feasible.
From July through December 1969, one minipond was
always operated without added iron while the remaining ponds
usually received a daily dose of 3 mg/1 Fe. The addition of
iron had little apparent effect on nitrogen uptake. Illus-
trated in Figure 22 are the results of a typical minipond
run during this period. Although the average total nitrogen
assimilation was slightly higher when iron was added, the
increase was less than 0.05 mg/l, and was of doubtful sig-
nificance. Analyses for total (particulate plus dissolved)
and dissolved iron in this and other runs provided inter-
esting results. Since the dissolved iron in all samples,
including one containing tile drainage only, and another
from a pond which received 3 mg/1 Fe were essentially the
same, the solubility of the element must have been extremely
limited. Conversely, total iron concentrations increased in
those ponds receiving added iron with the result that more
than 30 mg/1 Fe was present in many samples, presumably ac-
cumulating in the ponds as an iron phosphate precipitate.
Because of the low solubility of Fe in tile drainage, the
method of addition used in the studies reported herein was
extremely inefficient and should receive more consideration
before being used in future studies. A beneficial side
effect resulting from iron addition was that the FeCl3 acted
as a coagulant, thereby reducing the amount of coagulant
required in separation studies. This effect is discussed
further in the harvesting portion of the report.
Carbon Dioxide. Addition of CO2 was first evalu-
ated early in the summer of 1969* with rather spectacular
results. One hundred percent CO2 was added to two miniponds
by bubbling the gas into the intake side of the mixing pumps
during the afternoon mixing session. Results obtained with
one of the ponds are illustrated in Figure 23. Almost im-
mediately after CO2 injection, inorganic nitrogen and ortho-
phosphate concentrations decreased to near zero. For the
next few days the culture assimilated all the available
inorganic nitrogen and then turned brown and appeared to die.
Similar results were noted in the other minipond and in the
RGP. In the second minipond, the period of complete assimi-
lation was about twice as long as it was in the previous
pond. Injection of CO2 was discontinued when the culture
began to turn brown. Within four weeks after cessation of
CO2 injection, the algae (Scenedesmus) had regained their
-51-

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20-|
o>

ro
O
Z
h-
z
Ul
3
_J
li.
li.
UJ
no2-n

INJECTION OF C02
r2
NO3-N
1
20
DAYS OF RUN
01
E
O
Q.
I
csi
O
UJ
3
U
40
FIGURE 23-EFFECT OF C02 ADDITION ON EFFLUENT QUALITY
IN OUTDOOR GROWTH UNITS IN LATE SPRING

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original color and metabolic activity. Monitoring the in-
pond pH indicated that the pH level never fell below seven,
although it may have been much lower at the point of
injection.
The minipond runs following the initial C02 studies
all Included various levels of CO2 addition  100 percent,
50 percent, 5 percent, atmospheric air, and natural aeration.
In general, the results of these runs indicated that the
addition of CO2 increased nitrogen uptake and that a 5 per-
cent C02 concentration was preferable to 100 percent.
Table 8 contains average nitrogen assimilation data from
TABLE 8
AVERAGE PERCENT TOTAL NITROGEN ASSIMILATED FROM
20 mg/1 INFLUENT (MINIPOND RUN 8)
Detention Time : 		Type Addition
	Days	: 5ff CO2 : 100ft CO2 : Air ; Natural Aeration
3	43	52	46
5	64	62	60	49
8	78	74	70
minipond run 8, which lasted from August 6 through
September 29, 1969. These data show that at a three-day
detention time nitrogen assimilation was low regardless of
the availability of C02* indicating that detention time, not
carbon, was the limiting factor. At five days detention time,
5 percent CO2, 100 percent C02, and atmospheric air were all
somewhat more effective than was natural aeration. At eight
days detention time, about 8 percent more nitrogen was re-
moved in cultures receiving 5 percent CO2 than in those
receiving atmospheric air only.
Another example, based on minipond run 9A
(October 4 through November 19, 1969) is illustrated in
Figure 24. In this run, when the detention period was held
at 11.4 days, an average of about 80 percent of the influent
nitrogen was assimilated by cultures injected with 100 per-
cent and 5 percent CO2; whereas only about 55 percent was
removed by cultures nor receiving CO2 enriched air. When the
detention period was reduced to eight days, nitrogen uptake
seems to have been enhanced only when 5 percent CO2 was
added. With further reduction of detention time to five
days, assimilation was not affected by the presence or ab-
sence of CO2 enrichment. An interesting aspect of the ex-
periment in which 100 percent CO2 was added to one of the
-53-

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ponds was the sudden
decline in nitrogen
uptake after
November 9 (Figure 24).
At this time the rate
of algal growth in the
pond seems to have de-
clined, although the
culture appearance
remained green and
healthy. This pond
continued to receive
100 percent C02 until
December but never re-
gained its former as-
similation rate.
The appar-
ently active toxic
effect of 100 percent C02 demonstrated in the early summer
of 1969 was never again noted in subsequent runs. Possible
reasons for this effect may have been toxicity of the gas
itself or toxicity of material brought into solution by the
lower pH values. When the apparent toxic effect was first
noted in the outdoor cultures, samples of pond water and
CO2 were taken into the laboratory for lightbox culture
studies. No toxicity was evident when filtered pond water
was assayed for this factor with both 4 percent and 100 per-
cent C02. Another possible explanation for the die-offs was
based on the hypothesis that the algae died because of their
rapid growth rate resulted in nutrient starvation. This
hypothesis was tested by allowing laboratory cultures to
remove all of the available nitrogen from the medium and
leaving them in this nutrient-deficient condition for up to
10 days (with continuous lighting). At the end of this
starvation period, the cultures were given nitrogen, iron,
and phosphorus. The algae readily incorporated the additional
nitrogen and assumed a healthy appearance. The true cause of
the apparent CO2 toxicity is still unresolved and, since the
effect was never repeated after the first few occurrences,
the question may never be answered. Minipond runs in late
1969 did indicate that the addition of CO2 was required to
achieve maximum nitrogen assimilation in the outdoor cultures,
but that the quantity of CO2 which is optimum will depend on
season and drainage water quality.
Mixing. The exact role that mixing plays in outdoor
cultures may be a combination of various effects includings
(l) moving the algae into the light; (2) reducing anaerobic
(00-1
5% C0Z~^	\
50-
COMPRESSED A\R
h-
O
25
24
OCTOBER	NOVEMBER
FIGURE 24-EFFECT OF DIFFERENT LEVELS OF C02 ADDITION ON NITROGEN
ASSIMILATION IN PONDS AT 11.4 DAYS DETENTION TIME
-54-

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sludge banks caused by
settled algae; (3) re-
moving the algae from the
system by keeping them in
suspension and thus avail-
able for discharge in the
effluent; (4) replenish-
ing of carbon dioxide by
increasing the surface
area exposed to the atmos-
phere; and (5) by stirring
the bottom deposits,
making essential nutrients
more available. In the
first two minipond runs,
mixing was limited to
sweeping the ponds twice
daily. In these studies,
actively growing cultures
of Scenedesmus often com-
pletely settled from sus-
pension during the after-
noon, presumably as a
result of entrapment in
precipitated calcium and	FIGURE 25- EFFECT OF CONTINUOUS MIXING
magnesium salts at pH	ON NITROGEN ASSIMILATION, MP RUN 3
values above 10. In
minipond run 3, power and
equipment became avail-
able for mixing six miniponds. The results, shown in
Figure 25, indicated that the amount of nitrogen assimilated
could be significantly increased by continuously mixing the
cultures.
By the fall of 1968, all but two of the miniponds
had mixing systems, and in run 4 various durations of mixing
were investigated. The average percentages (each number is
an average of three ponds) of total nitrogen assimilated for
approximately 60 days operation are plotted in Figure 26.
The two- and four-hour mixing periods took place in the
afternoon, and the 12-hour period occurred during daylight
hours. The results showed that total nitrogen assimilation
was almost doubled by the optimum amount of mixing and that
4 and 24 hours had approximately equal effects. The unex-
plained decrease in nitrogen removal in cultures mixed 12
hours daily noted in the individual weekly averages was
unexplained but statistically significant. During the run,
total nitrogen removals were low in all the ponds; therefore,
the study was repeated on a smaller scale in May and June,
1969. The main emphasis at that time was placed on deter-
mining what portion of the daylight hours was optimum for
S 80-
2
8
H-
Z
UJ
3
60-
40-
5 DAYS
2.75 DAYS
g
H 20H
u.
o
DETENTION TIME-DAYS
iu
u
OC
K
1
24
HOURS OF DAILY MIXING
-55-

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mixing the cultures. The results of the four mixing periods,
four hours in the morning, four hours in the afternoon, 15
minutes of each daylight hour and for 14 daylight hours, are
shown in Figure 27. After the initial period of adjustment,
30-
tii
20
24
HOURS OF DAILY MIXING
FIGURE 26-EFFECT OF MIXING DURATION ON AVERAGE TOTAL
NITROGEN ASSIMILATION IN MINIPOND RUN 4A
up to about May 20, four-hour mixing in the afternoon con-
sistently increased the amount of nitrogen incorporated by
the algal cultures. Pull daylight mixing, either continu-
ously or for 15 minutes of each hour, was detrimental to
nitrogen assimilation. Again, in July 1969 cultures con-
tinuously mixed during daylight hours assimilated less
nitrogen (65 percent) than did cultures with afternoon mixing
only (85 percent), both at eight days detention time.
Based on the information from the runs cited above,
the standard mixing regimen imposed was four daylight hours.
For convenience in sampling, this period was divided into two
intervals  0800 to 0o30 hours and 1200 to 1530 hours. One
effect of this regimen Imposed on the miniponds was to peri-
odically change the type of flow through the system. Tracer
-56-

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z
o
MIXING SCHEDULE
1500-1930-
0600-1030
0600-2000
I5min/hr -0600-2000-
UJ
cc
ui
CL
10
15
20
25
30
MAY
JUNE
FIGURE 27-EFFECT OF TIME OF MIXING ON TOTAL NITROGEN ASSIMILATION
studies showed that the pond influent went through the pond
as a discrete entity (plug flow) when the mixing pumps were
off. During mixing periods, the ponds acted as almost
completely mixed vessels.
The average in-pond mixing velocities were deter-
mined by measuring the velocity at various points in some
selected ponds. These results showed that the average velo-
cities varied from about 0.25 feet per second (fps) in the
16-inch pond to about 0.50 fps in the 8-inch ponds. These
velocities did not prevent the algae from settling, especially
in some stagnant areas of the pond. The two ponds that were
modified to eliminate these stagnant spots (see Figure 5) had
higher velocities than comparable unmodified ponds, and they
were relatively free of algal sludge banks. In spite of the
increased mixing efficiency, no significant difference occur-
red in rates of nitrogen assimilation between algal cultures
grown in the two types of pond.
After the addition of CO2 became routine, one mini-
pond was left without C02 and without mixing. The data from
this unit, when compared to a similar pond that was mixed,
showed comparable effluent dissolved nitrogen levels, a find-
ing that indicated mixing was not always essential to algal
growth. Preliminary evaluation of the effect of C02 addition
to a nonmixed pond has indicated that CO2 alone may be ade-
quate for achieving the desired rates of nitrogen uptake.
The mixing requirements of an algal stripping system will
receive more emphasis in the 1970 operational studies.
-57-

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Detention Time. Detention times as used in this
study are theoretical hydraulic terms computed by the
formula:
9 = V
F
where 0 is the detention time, P the flow, and V is the
volume of the container. Three separate dye tracer studies
indicated that the true and the theoretical hydraulic deten-
tion times were in reasonable agreement. For example, the
calculated theoretical detention time for one of the ponds
was 5 days and the measured true detention time about 5.2
days. Because of pond design, mixing, and natural algal
settling rates, cell detention time was significantly longer
than the hydraulic detention time. In one instance, when
the two were compared by use of tracer studies, the values
were about 30 days for the cells, as compared to 5 days for
a hypothetical water molecule (periods of mixing, 0800 to
0830 hours and 1200 to 1530 hours).
The effect of detention time can be best evaluated
from results obtained after July 19&9 because standard mix-
ing, nutrient additions, and detention times were used during
this period. Detention times of 3, b 8* H.5> and 16 days
were studied, although only three of these times were com-
pared in any particular run. An example of a detention time
study is illustrated in Figure 28. The ponds were operated
on a batch basis until August 15> 1969* when the desired
detention times were set. After reaching steady-state, the
pond with an eight-day detention time consistently assimi-
lated 80 percent or more of the influent nitrogen. At five
days, steady-state was reached at a level less than 75 per-
cent, and at three days the level was closer to 50 percent
total nitrogen uptake. The sudden decrease in removal that
occurred about September 12 may have been caused by a sudden
temperature drop during this period. (See Figure 48.)
The overall effect of hydraulic detention time is
best illustrated in Figure 29, in which the data from four
minipond runs from July through December 1969 are summarized.
The data are averages of total nitrogen assimilations for
12-inch ponds in which C02 was the only other variable tested,
i.e., depth, mixing, etc., were identical. As will be shown
in later sections, detention times can also be affected by
physical changes in the system. The influent total dissolved
nitrogen concentration was about 20 mg/l during this period.
The results demonstrate clearly that detention times must be
varied seasonally to achieve the desired algal growth. The
average maximum removals of dissolved influent nitrogen for
each run (which included start-up) were consistently near
-58-

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t
r-20
I00n
8 DAYS-
5 DAYS-
z 75-
3 DAYS

	\
u 25-
30
20
AUGUST
SEPTEMBER
DETENTION TIME - DATE
FIGURE 28-EFFECT OF DETENTION TIME ON NITROGEN ASSIMILATION, 5% C02 ADDED
80 percent, while in the actual runs the steady-state levels
were in the 85- to 95"Percent range, in these 12-inch depth
ponds, eight days was the shortest detention time which would
allow for the assimilation of an average of 80 percent of the
influent nitrogen. Although not shown in this figure because
of differing mixing schedules and the absence of carbon di-
oxide, data for May and June also indicated that detention
times of less than eight days were too short for maximum
nitrogen assimilation.
Required detention time for a given rate of nitro-
gen assimilation is intimately related to water temperature
and must be adjusted to ensure that the rate of nitrogen
assimilation is not significantly different from nitrogen
input. Smooth curves of the maximum and minimum daily
minipond temperatures are shown in Figure 30. Actual maxi-
mum and minimum water temperatures recorded were 98F
(36.6C) and 3^F (l.lC) respectively. The average maximum
temperature for each run was calculated from these curves,
(Because the curves had generally similar shapes, minimum
temperatures could also have been used.) In Figure 31, shows
the average maximum temperatures versus the detention time
required to remove about 16 mg/l of the total iijfluent
nitrogen.
-59-

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MINIPOND RUN No. AND DURATION
8	H H	9A	H
98
note: numbers above bars indicate theoretical
HYDRAULIC DETENTION TIME
60-
1
1.5
11.5
8
I
JULY
AUG
SEPT	OCT
MONTH
NOV
DEC
FIGURE 29-EFFECT OF DETENTION TIME ON NITROGEN REMOVAL-
SUMMARY OF DATA FROM 12 INCH PONDS FROM JULY-DECEMBER 1969
Prom these data it appears that detention time was indirectly
proportional to maximum average pond temperature between 15
and 25C, the range of temperatures during the selected
study period. With standardization of pond operation and
shorter run times during the operational studies, such a
curve will be extended to cover the entire range of temper-
atures found on the west side of the San Joaquin Valley.
Culture Depth. Water depths of 8, 12, and 16
inches were studied as part of the investigation. Depth
primarily affects amount and quality of available light
energy received by algal cultures and indirectly affects
mixing. As shown previously, the average water velocity in
the 16-inch deep pond was about 0.25 fps as compared to
0.5 fps in an 8-inch pond. Experiments conducted during the
period of May through December 1969 were taken as the source
of data for studying this variable, because of the uniform
operating conditions during that time. Because of the limited
pond availability, only the 12-inch depths were replicated in
any particular run. The data for the 8- and 16-inch depths
were from single ponds. In all minipond runs, nitrogen
-60-

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assimilation
in the 8- and
16-inch cul-
ture depths
differed dis-
tinctly. The
curves in
Figure 32 show
the typical
effect of
depth. All
these ponds
were operated
at eight days
detention time
and received
5 percent CO2.
The overall
average assim-
ilations of
total influent
nitrogen were
80, 69, and
55 percent for
8, 12, and 16
inches,
respectively.
JULY	AUG	SEPT	OCT	NOV	DEC	JAN
MONTH
FIGURE 30-MAXIMUM AND MINIMUM MINIPOND TEMPERATURES-JULY TO DECEMBER 1969
16-]
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O
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8-
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10	15	20	25
AVERAGE MAXIMUM POND TEMPERATURE
	1	
30
FIGURE 31-PLOT OF DETENTION TIME REQUIRED TO ASSIMILATE
APPROXIMATELY 80% OF THE TOTAL NITROGEN vs AVERAGE RUN TEMPERATURE
-61-

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All of
the average total
assimilations for
each depth were
plotted for the
five minipond
runs, Figure 33*
and the slopes
(change in percent
removal per change
in depth.
A^R/AD) of the
straight lines
from the percent-
age at 8 inches
to that at 16
inches were calcu-
lated, and the
values listed in
Table 9 were ob-
tained. The data
show that the
slope of the line increased as the days became shorter, and
that the difference in nitrogen assimilation between algae
cultures grown at 8 and 16 inches of depth was more pro-
nounced when available light energy per day increased.
In Figure 3^ the slopes from Table 9 have been
plotted as a function of the average daily solar radiation
during each of the runs. A straight line fitted by eye
indicates that the effect of depth within the limits tested
was directly correlated to average dally solar radiation.
As illustrated in Figure 35* one effect of de-
creased depth is to lessen the detention time required for
algal assimilation of the available nitrogen. In the four
runs shown, the average amounts of nitrogen assimilated
were practically identical; however, in each run the neces-
sary detention times were three to four days shorter in the
8-inch culture depths. The savings in detention time are
about balanced by the additional land area required for
treatment at the shallower depth.
Soil Ponds. The two ponds which had layers of
soil on the bottom received a minimum of operational atten-
tion, consisting mainly of regulation of the influent flow
and addition of phosphorus, and they were never drained or
refilled. In spite of, or perhaps because of, the lack of
attention, both ponds maintained cultures of organisms which
removed surprisingly large amounts of nitrate-nitrogen.
Both ponds had a mixed biota consisting of algae, zooplankton,
100-,
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NOTE! DETENTION TIME, 8 DAYS WITH 5% C02
15
OCTOBER
25
14
NOVEMBER
20
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5 DAYS D.T.
8 DAYS D.T
DATES	OF RUN
D	MAY-JUNE
	JULY
O	AUG-SEPT
X	OCT-NOV
A	NOV-DEC
D.T.	DETENTION TIME
\ s8DAYS
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\ 8 DAYS -l^^X
x DT Sa
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N.
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11 DAYS D.T.
s
"A
12
POND DEPTH - INCHES
-1
16
FIGURE 33-AVERAGE TOTAL NITROGEN ASSIMILATION FOR
VARIOUS CULTURE DEPTHS, MINIPOND RUNS 6-9B
TABLE 9
SLOPE OF VARIOUS DEPTH VERSUS PERCENTAGE REMOVAL CURVES
Run
Approximate :
 Dates of Run :
Slope
(A#R/AD)*
6
May-June
1.5
7
July
1.5
8
August-September
2.5
9A
October-November
3.1
9B
November-December
5
~Percent change in removal per change in depth from 8 to
16 inches.
-63-

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higher aquatic
plants, bacteria,
and flying insect
larvae (mainly dip-
teran). Dissolved
oxygen determin-
ations made at dif-
ferent times of the
year indicated that
the soil ponds were
always aerobic, at
least above the
soil-water inter-
face. The amount of
algae was usually
low (less than 100
mg/1), and the algae
were often of the
flagellated variety,
e.g., Euglena and
Carteria. An indi-
cation that the
nitrate removal
E
NE 80 Ch
z> 600-
o 400'
NOTE:
CHANGE IN PER CENT REMOVAL AS DEPTH
CHANGES FROM 8 TO 16 INCHES
t.O	2.0	3.0	4.0	5.0
SLOPE OF LINES (A%R/CHANGE IN DEPTH+) FROM TABLE 9
FIGURE 34-SLOPE OF DEPTH vs PER CENT REMOVAL CURVES COMPARED
WITH AVERAGE SOLAR RADIATION DURING CORRESPONDING RUN
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mechanism was not entirely algal was provided in a study in
which a sample of the soil, water, and organisms was en-
closed in a darkened bottle and floated in the pond. At the
end of one week, the original 21 mg/1 of total dissolved
nitrogen had been reduced to 14.5 mg/l ( JO^-N, 12 mg/1;
N02-N, 2 mg/1; and NJto+Organic-N, 0.5 mg/l). Because light
was not available during the incubation period, the nitrogen
reduction was assumed to be the result of anaerobic bacterial
decomposition. The dissolved oxygen content of water in the
bottle decreased from about an initial 12 mg/l to a non-
detectable amount at the end of the test period. The organic
carbon source was presumably the product of algal and bac-
terial decomposition, as well as extracellular organic
products of algal metabolism.
A comparison of nitrogen removal data from one of
the soil ponds with a minipond of comparable depth and de-
tention is shown in Figure 36. The remaining soil pond
followed essentially the same pattern as the one shown and
was omitted for graphical clarity. Also included in the
figure are the removal data for the best 12-inch depth non-
soil pond in each minipond run. These data support the fol-
lowing conclusions: soil pond cultures consistently removed
more nitrogen than did a mixed minipond at a similar deten-
tion time, and the removals in the soil ponds were often
higher than in the best routinely operated minipond.
25i
SOIL POND
'r-
MINIPOND ON SAME SCHEDULE
AS SOIL PONDS
STANDARD MINIPOND WITH
.HIGHEST NITROGEN ASSIMILATION
10-

DEC
JAN
MAR
JUL
NOV
FEB
AUG
JUN
APR
MONTHS
MAY
SEP
FIGURE 36-COMPARISON OF TOTAL NITROGEN ASSIMILATION IN ONE SOIL POND WITH A COMPARABLY
OPERATED MINIPOND AND STANDARD MINIPOND WITH HIGHEST NITROGEN ASSIMILATION
-65-

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The operational studies In "1970 were designed to
define the mechanisms involved in reduction of nitrogen
in these ponds, which we have called "symbiotic ponds".
The nitrogen removal mechanisms for these units appear to
resemble that in grass plots described by Williford and
Cardon (1971) in which water flowing through fields of
water grass showed marked nitrogen reduction. In the grass
plots, relatively little planktonic algal growth took place;
dissolved oxygen concentrations were always above zero; and
no organic carbon was added. Net influent nitrate concen-
trations ranging from 30 to 300 mg/1 (as NO3) were reduced
by amounts ranging from 50 to almost 100 percent. The
authors tentatively concluded that the reduction was the
result of anaerobic denitrification in local strata near or
at the soil-water interface.
Blomass Control. The term biomass control is used
to describe the process of removing settleable algae and
other settleable materials which tend to accumulate in the
ponds during long periods of operation. Mixing schedule,
algal settling, and pond design each contributed to the
problem of accumulation of algal cells in the small outdoor
ponds. The possibility that the older cells might be detri-
mental to growth of the younger cells was considered in this
study. The detrimental effect could result from light ex-
clusion by older, nonreproducing cells, by the production
of some autoinhibitory substance, or by competition for
scarce nutrients. Although not a. part of the algal biomass,
particles of suspended inorganic precipitates also tended
to accumulate in the ponds and substantially reduced light
penetration into the cultures. To determine whether or not
such effects were present and their influence, a settling
tank (described previously) was attached to the mixing pump
of one of the two redesigned miniponds, while the other pond
was operated as a control. As shown by the curves in Figure
37> the settling tank reduced the volatile solids within the
pond by about 50 to 60 mg/1 during minipond run 9-A.
The total nitrogen removal for the two ponds is
shown in Figure 38* both at eight days detention time and
with 5 percent CO2 addition. The average total removal for
the pond with biomass control was about 80 percent; for the
control pond, 6l percent. During this run the best of the
remaining 12-inch ponds also removed about 80 percent but at
a detention time of 11.5 days. In another run involving
five days detention time, the pond with the settling tank
removed an average of 71 percent compared to 59 percent re-
moved by the control. These data indicate that accumulation
must be prevented in outdoor cultures, and that the construc-
tion of larger units may have to Include some mechanism for
controlling the accumulation of inactive algae in the pond,
-66-

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-NO BIOMASS CONTROL
BIOMASS CONTROL
FIGURE 37-COMPARISON OH VOLATILE SOLIDS IN MINIPONDS
WITH AND WITHOUT BIOMASS CONTROL
such as in-pond settling
areas with sludge re-
moval, or other suitable
methods. By providing
control of excess bio-
mass and inert materials
the required detention
times for maximum nitro-
gen assimilation may be
lowered significantly.
The data from this
study of biomass control
indicated that, during
the period from July
through November, the
rate of nitrogen assimi-
lation in the biomass-
controlled ponds were
comparable to other 12-
inch ponds which were
being operated with
about three days of
additional deten-
tion time. Thus
with biomass control,
five days may be .
the minimum summer '
detention time to
achieve 80 percent
or more assimilation.
Addition
of Fish. Some pre-
liminary studies
indicated that fish
may possibly en-
hance the growth of
algae in tile drain-
age waters and aid
algal assimilation
of nitrogen. Their
beneficial effect
was thought to be due in part to their stirring action and
in part to their production of some waste product essential
to algal metabolism. For these reasons, and because fish
might also add a true recreational value to drain waters,
a quantity of Sacramento blackfish weighing from 0.5 to
1.5 pounds each were obtained in May 1968 and placed in
some ponds during the first four minipond runs. Some ponds
received 2.5 pounds of fish and some received 5 pounds of
S ,0<>i
5
s

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fish. This particular species was selected for Its reported
active consumption of planktonic algae.
The fish survived in the miniponds, although some
mortality was noted. Unfortunately, no record of growth was
maintained; thus we do not know if the fish actually gained
weight in the ponds. The fish did have a beneficial effect
on nitrogen uptake. In Figure 39* which summarizes their
impact in the first four runs, the best minipond without
fish is compared to a pond with comparable operating cri-
teria containing fish. In all four runs, the algae in the
ponds with fish assimilated more nitrogen than did the con-
trol ponds. The increased nitrogen assimilation attributed
to the presence of fish ranged from 13 to 43 percent with an
average of 28 percent. After minipond run 4a, all fish were
removed from the miniponds, chiefly because iron additions
had proved to be more effective at increasing nitrogen re-
moval but also because the screens covering the influent to
the mixing pumps (to keep the fish out) were reducing the
mixing efficiency. Large algal ponds can apparently support
fish growth and fish apparently do enhance algal growth and
nitrogen assimilation.
lOOn
75-
FISH
25-
CONTROL
AUG
SEPT
MAY
JUNE
JULY
OCT
1968
FIGURE 39-PROPORTIONATE INCREASED NITROGEN ASSIMILATION
ATTRIBUTED TO FISH, BY SEASON
-68-

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Rapid Growth Pond
This pond was of limited value as a research tool
because of the lack of a comparable unit to use as control;
consequently, it was used principally as a demonstration
unit and a source of algae for separation studies. During
the first few months of operation we were unable to maintain
Scenedesmus cultures and the pond was reseeded twice, once
in the summer of 1968 and again in January 1969. The pond
normally was operated with a 12-inch water depth and at vary-
ing detention times, on the basis of minipond data. During
these first few months, the pond occasionally was operated
at less than 20 mg/1 influent nitrogen and with different
mixing cycles. After January 1969, the pond was operated
for almost five months without being drained and with little
variation in operating procedures.
Plots of effluent nitrogen, influent nitrogen, and
detention time for the RGP during the calendar year 1969,
are presented in Figure 40. The period up to June 12 was
characterized by a relatively stable effluent that averaged
about 5 mg/1 total N (75 percent assimilation). On June 12,
100 percent C02 was added to the pond, whereupon the effluent
inorganic nitrogen decreased to near zero. Ten days later
the pond turned brown and nitrogen removal ceased. On July 7
INFLUENT NITROGEN
rZO
20-]
EFFLUENT NITROGEN
DETENTION TIME
-I0P
10-
T
OCT
NOV
DEC
JUN
SEP
FEB
MAR
AUG
MAY
JUL
APR
JAN
MONTHS
FIGURE 40-OPERATION OF RAPID GROWTH POND DURING 1969
-69-

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the pond was drained and cleaned, and on July 14 the pond
was refilled and Inoculated with the contents of two of the
minlponds. The 100 percent C02 was again added but with
slightly different results than before. Scenedesmus began
to grow actively but were soon replaced by diatoms and blue-
green algae, which floated to the surface one afternoon and
were removed by surface overflow. The Scenedesmus then
returned in late July and have since remained as the domi-
nant algae. The possible explanations for this change in
species composition could be the apparent C02 toxicity
discussed earlier In the section on addition of C02 to the
miniponds.
After July 1969 we were hesitant to try C02 again
because the Scenedesmus culture in the pond was the principal
source of material for the separation studies; and, inasmuch
as the addition of 100 percent CO2 to the Pond might have
proved lethal or at least Inhibitory to the population of
Scenedesmus, no CO2 was added to the pond thereafter. Also,
after July the pond was often operated to conform to the
needs of the individual separation studies. For example,
occasionally the pond was mixed all day to provide a uniform
algae supply even though full-day mixing had been found to
reduce nitrogen assimilation. Because of this, the nitrogen
uptake figures are minimal approximations of the unit's
potential. The 1970 operational studies will provide a more
realistic evaluation of the potential nitrogen removal capa-
bilities in this pond.
Biological and Chemical Observations
Blomass Production. As used in the present study,
the term biomass refers to the results of volatile suspended
solids determinations. In many algal growth studies, cell
counts, packed cell volume, or light transmittance (or
absorbance) are used to estimate changes in biomass. The
primary interest at Pirebaugh was in biomass because of its
possible effect on the growth system and on the market value
of the algae produced; therefore, estimates of biomass pro-
duction were needed. Volatile suspended and total suspended
solids of algal cultures can often be used interchangeably
since the ash portion of algae is usually only 10 to 15 per-
cent of the total weight. At Pirebaugh the volatile portion
was usually only 50 to 70 percent of the total suspended
material. The nonvolatile solid material consisted of in-
soluble complexes of magnesium, calcium, phosphorus, potassium,
and iron resulting from the high pH levels attained in the
growth units, as well as of some windblown soil particles.
Although CO2 addition to the growth units did lower the pH,
-70-

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apparently no signifi-
cant change occurred
in the percentage of
volatile material in
the effluent solids
(Figure 4l).
The mixing
system finally selected
for general use in the
outdoor growth units
caused obvious diurnal
fluctuations in the
effluent volatile
solids. A typical
diurnal cycle is illus-
trated in Figure 42.
In this particular in-
stance, the 0830 vola-
tile solids (normal
sampling time) of
374 mg/1 indicated an
unrealistically high
value for the amount of
biomass produced by the
system. Graphical in-
tegration of this curve
indicated that the aver-
age volatile solids
concentration for the
24-hour period was ap-
proximately 120 mg/l.
Because of the diurnal
fluctuation caused by
algal settling, the total
pond biomass increased
indicating that it was
accumulating at a rate
faster than that at which
it was being removed.
Volatile solids concen-
trations of more than
500 mg/1 often were noted
in the growth units and
the solids seldom reached
steady-state conditions.
Actual biomass
production during a 19-
day period of August-
September 1969 was calcu-
lated for a minipond in
70
60
8
LJ
I
<
O
>
UJ
o
g 50
CL
	,	1	I
0	5	10
PER CENT CARBON DIOXIDE ADDED
FIGURE 41-AVERAGE AND RANGE- PER CENT
VOLATILE SOLIDS OF TOTAL SUSPENDED SOLIDS AT
DIFFERENT LEVELS OF C02 ADDITION
-MIXING PERIODS
400
r 300-
\
o
E
(/)
Q
d 200-
 100-
0600
1200
2400
SAMPLING TIME
FIGURE 42- DIURNAL FLUCTUATIONS IN EFFLUENT
VOLATILE SOLIDS
-71-

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run 8. Effluent (0830
samples) volatile solids
and total nitrogen are
shown in Figure 43. An
overnight study of the
pond during this period
indicated that the aver-
age volatile solids in
the effluent were about
one-half of the 0830
values. Both curves in
Figure 43 were divided
into 48-hour intervals
and the average effluent
nitrogen and volatile
solids calculated for
each interval. During
this 19-day period, an
average of 240 mg/1
volatile solids was dis-
charged in the effluent
of the pond. From this
figure, the actual pro-
duction of algae per
liter of influent can be
calculated from evapo-
ration data by assuming
that one-fourth of the
influent was lost through evaporation. Based on this calcu-
lation, about 180 grams of volatile material were shown to
be produced from each liter of influent. If it is further
assumed that algae are about 90 percent volatile, then
the actual algal production was approximately 200 mg/1.
For the 19-day period, an average of approximately
17 mg/1 of influent nitrate as N was used in the production
of the biomass. Chemical analyses showed that the volatile
solids were about 8.3 percent nitrogen. These data result
in the following nitrogen balance:
Cl should = Cde + CAE - (cAi " cAe) + Cx
185 should - 23 + 146 + 6 + Cx
185 =* 175 + cx
where Cj * grams N entering system
Cde 58 grams N dissolved in effluent
CAE " grams N in effluent algae
VOLATILE
SOLIOS
w) 300-
EFFLUENT
NITROGEN
o
o
fx.
H
2
UJ
3
-J
a
UJ
3
o
(/>
(/)
5
g
t-
FIGURE43-EFFLUENT NITROGEN AND VOLATILE SOLIDS
USED IN BIOMASS PRODUCTION FIGURES
-72-

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Cai = grams N in in-pond algae-initial
CAe = grams N in in-pond algae-final
Cx = grams N not accounted for
Although an exact balance was not obtained, the difference,
Cx, could be in the range of experimental and sampling error,
or could be a measure of nitrogen lost by some mechanism such
as denitrification. Based on the information accumulated
thus far, it is estimated that the overall biomass yield for
the algal system will be on the order of 1 ton of volatile
solids per million gallons of waste plus 1/2 ton of inert
materials.
Algal Genera Noted. The number of algal genera
which were noted in the pond cultures was relatively low.
The only genera that dominated the cultures are shown in
Table 10. As previously noted Scenedesmus quadricauda was
originally introduced, but occasionally Scenedesmus dimorpha
was noted although never as the dominant algae. Scenedesmus
was typically in the two- or four-celled stage, although in
very active cultures the alga took a unicellular form with
four small spines. The soil ponds went through several algal
successions, with some flagellated algae (usually Carteria,
Phacus, or Euglena) as the dominant genera. The soil ponds
also often contained large numbers of an extremely small uni-
cellular green algae, either Chlorella or Nannochloris.
TABLE 10
DOMINANT ALGAL GENERA AT THE IAWTC
Genus
Type
Scenedesmus
Oscillatorla
Dysmorphococcus
Schroederia
Lagerheimia
Navicula
green
blue-green
green
green
green
diatom
Scenedesmus was originally seeded into the RGP in
the spring of 1968 and dominated the cultures until early
summer of that year. At that time, species of other genera,
mainly of Dysmorphococcus and Oscillatoria, began to dominate
the cultures and, by August, Scenedesmus had almost entirely
-73-

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disappeared. The RGP was then drained, cleaned, and refilled,
and another 100 pounds of Scenedesmus sp. obtained from the
Richmond Field Station were added. Prom then on, Scenedesmus
was the dominant species in the RGP, although the culture
was killed by CO2 in the summer of 1969 and was reseeded from
the miniponds. Summer conditions appeared to be particularly-
suitable for growth of the blue-green alga, Osclllatorla.
During both 1968 and 1969, this alga was prevalent in some
minipond cultures. Scenedesmus reestablished dominance with
the onset of cooler weather.
All of the algae listed in Table 10, with the ex-
ception of diatoms and Lagerheimla, appeared capable of re-
moving large quantities of nitrate-nitrogen. The appearance
of large numbers of diatoms or Lagerheimla was usually ac-
companied by a decrease in nitrogen assimilation.
Predatory Organisms. Relatively little emphasis
was placed on this aspect of pond ecology. However, during
the spring and early summer of 1969, some data were collected
on potential algal predators. Rotifers were by far the most
common of such organisms with populations of 30,000 organ-
isms/liter noted on occasion. In ponds with actively growing
cultures of algae, rotifers were always present in numbers
fewer than 500/liter. Large numbers of rotifers usually
were found in ponds with senescent or dying algal populations,
but it is not possible to state whether rotifers preceded or
followed the senescent conditions.
Planktonic crustaceans (Copepoda, Daphnla, etc.)
were notably absent from the algal ponds, and only in the two
soil ponds were these organisms prevalent. Although usually
fewer than 1,000 Copepod nauplii and adults per liter appeared
in the cultures, on one occasion more than 5,000 of these
organisms per liter were noted in a soil pond.
Dissolved Effluent Nitrogen. In general, the dis-
solved effluent nitrogen contained only nitrate, nitrite,
and organic nitrogen. Ammonia, if detectable, was almost
always less than 0.1 mg/1, compared to an undetectable amount
occurring in the influent. Biological activity in the mini-
ponds did cause dissolved organic nitrogen to increase from
about 0.4 mg/1 to 0.7 - 0.8 mg/l. Dissolved organic nitrogen
usually was higher in the soil ponds than in the remaining
miniponds. In minipond run 8 conducted during August and
September 1969, dissolved organic nitrogen in all the mini-
ponds was consistently above 1 mg/1 and occasionally greater
than 1.5 mg/1.
The concentration of nitrite-nitrogen followed a
consistent pattern in the outdoor growth studies. After the
ponds were filled, nitrite generally increased to 1 to 2 mg/1
-74-

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and then, as the nitrate decreased, nitrite followed the
same trend. This pattern is illustrated in Figure 44, which
shows a minipond with a high percentage of total nitrogen
removal. The ponds with low effluent nitrate usually also
had a low nitrite concentration.
On several occasions, effluent nitrate-nitrogen
concentrations were determined at two- or three-hour inter-
vals for a 36-hour period. The variations in concentration
were beyond the limits of the accuracy of the analysis; thus,
a diurnal change in the concentration of this constituent
could not be demonstrated.
Dissolved Oxygen. This parameter was not routinely
monitored, although determination of dissolved oxygen con-
centration was a part of most diurnal studies. An example of
the diurnal fluctuations in dissolved oxygen in four mini-
ponds (data from September 10-11, 196Q) are shown in
Figure 45. Ponds 1, 4, and 22 each showed approximately 75
percent total nitrogen assimilation, whereas pond 10 showed
only about 40 percent. The greatest fluctuation was noted
in pond 22, an unmixed soil pond. Undoubtedly this change
occurred because the afternoon mixing tended to deoxygenate
the pond water in the remaining ponds. The 5 mg/1 minima
noted in ponds 1, 4, and 22 were near the minimum concentra-
tions recorded in any of the overnight studies.
It is interesting to note that the deoxygenation
factor for the ponds is on the order of 1.25 mg/l/hr, whereas
the oxygenation factor is a net of 2.5 mg/l/hr, or a gross of
57. C02f 3.0 mg/l Fe
0 DAYS DETENTION TIME
2 mg/l PO4-P
AUGUST
SEPT
days of run
FIGURE 44-CHANGE IN NITROGEN FORMS DURING MINIPOND RUN
-75-

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TABLE 11
EFFECT OF CARBON DIOXIDE AND AERATION
ON ALKALINITY IN MINIPOND RUN 8
Pond
No.



Addition:
Detention
time
(Days)

a/12/69

8/2b/fo9


9/5/59

pH :
Alkalinity+
 
 
:pH :
Alkalinity+
 
 
:pH :
Alkalinity+
12
100$ C02
5
9.3
185
7.0
529
7.75
500
H
5$ CO2
5
9.25
164
7.9
428
7.8
369
20
Control
5
9*2
168
8.8
297
8.6
305
RGP
No air
10
9.2
197
8.9
280
8.75
235
Influent


7.6
374
7.5
373
7.5
373
+ mg/1 as CaC03

-------
about 4 mg/l/hr. If an oxygen : algae ratio of 1.5 is assumed,
the rate of algae growth is 2.66 mg/l/hr, or about 32 mg/l/12
hr-day. At this rate about eight days would be required in
September to attain the 240 mg/1 algae required for 90 percent
N assimilation. This detention agrees with the required de-
tention times illustrated in Figure 29.
pH and Alkalinity. As noted earlier, growth of
algae usually affects the pH and alkalinity of systems ex-
posed to the atmosphere. A typical minipond without C02
addition would reach an afternoon pH of more than 10 and bi-
carbonate alkalinity approaching zero, as calculated by the
table in Standard Methods. During this period of low bicar-
bonate alkalinity, algal growth probably was carbon limited.
Some miniponds also showed a diurnal change in total alka-
linity caused by precipitation and redissolving of calcium
carbonate. The precipitation caused the pond surfaces to
collect a scale, which was made up of a complex of phosphorus,
magnesium, and calcium carbonates. Alkalinity samples had to
be filtered to eliminate a sliding end-point caused by the
presence of colloidal calcium carbonate in the sample.
As shown in Figure 46, the addition of 100 percent
CO2 had the effect of providing the algae with bicarbonate
alkalinity. In ponds to which C02 was added, the diurnal pH
change was minimized and the pH usually never reached more
than 8.5 to 9.0; bicarbonate alkalinity was also present at
all times of the day. A further effect of added CO2 is shown
in Table 11. With the lowering of the pH level brought about
by the C02, the calcium carbonate scale went back into solu-
tion, which in turn raised the total alkalinity. In the ex-
ample shown, there was greater than 100 mg/1 (as CaC03) more
effluent alkalinity by the end of the run than in the
influent. This particu-
lar minipond run was
started with a culture
which had not received
C02 previously; thus, the
initial pond alkalinities
had been lowered by
CaC03 precipitation. The
data for pond 20, the
control pond, indicates
that the cultures were not
very active (low pH and
alkalinity), and this ob-
servation is verified by
the nitrogen removal data.
Only about 50 percent of
the total influent nitro-
gen was removed in this
minipond.
>- 20-
MP-4
1700	2200	0300	0800	1300
SAMPLING TIME
FIGURE 45-DIURNAL FLUCTUATIONS IN DISSOLVED OXYGEN
-77-

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RGP Evaporation
and Temperature Data. A
weather station located
in the rapid growth pond
provided information on
daily rates of evapora-
tion loss and temperature
change during 1969. Evap-
oration data was used to
obtain mass balances of
water and dissolved nutri-
ents passing through the
pond.
The RGP, filled
to a depth of 12 inches,
contained 81,000 gallons
of water, or slightly
less than 7,000 gallons
per inch of depth.
Figure 47 shows that the
minimum daily evaporation
loss was nearly -0.06 inch,
which occurred in February
1969 (an atyplcally rainy
month) and the maximum
daily evaporation loss was
0.4 inch in August. An
evaporation of 0.4 inch of
pond water represents a
volume loss of about 2,800
gallons. Total evapora-
tion loss for the entire
year was about 67 inches
of water.
Detention times
during the period of maxi-
mum evaporation were about
8 days, which required an
influent flow of about
10,000 gallons of drainage
water per day. Because of
evaporative losses of
2,800 gallons per day, the
effluent flow was only
about 70 percent of the
influent. The numbers
shown for nitrogen removal
were based on effluent
concentrations compared to
10
8 300
o
O
(/)
<

200
-J
<
-J
< 100
LlJ
O
03
tr
NOTE: DETENTION TIME-8 DAYS
INJECTION j
/
\	/ /-CONTROL
\ /vj
w-
1200	2400
SAMPLING TIME
1200
FIGURE 46- DIURNAL CHANGES
IN BICARBONATE ALKALINITY
+0,4QOi
+0.300-
(/)
Ui
X
o
2
z
2 +0.200

UJ
>J + O.J 00-
f
UJ
UJ
<3
<
01
UJ
>
<
-O.tOO
i
h
J FMAMJJASOND
MONTH (1969)
FIGURE 47-AVERAGE DAILY EVAPORATION IN
RAPID GROWTH POND DURING 1969
-78-

-------
influent concentrations, without regard to total pounds of
nitrogen entering and leaving the system. This type of
measurement was used because an effluent standard of 2 mg/l
total dissolved nitrogen was in the original study objective.
Computation of percent assimilation based on weights of nitro-
gen in the influent and effluent would yield higher rates of
removal efficiency, especially during periods of maximum
evaporation.
Presented in Figure 48 are the daily maximum and
minimum pond temperatures observed in the RGP during 1969.
During the months of May through early September, the maxi-
mum pond temperatures were consistently in the 80 to 90F
(27 to 32C) range. The highest water temperature recorded
during the entire period was 92P (33C) -- about 6 to 8P
(4C) cooler than the maximum minipond temperature. The
minimum temperature was 38^ (3C) in late December. About
September 15 there was a sudden 9 to 10F (6C) decrease in
temperature from about 85 to 87P (about 30C) down to 75 to
78F (about 24C). Although temperatures were still quite
warm, a decline in nitrogen assimilation accompanied the
drop in almost all of the minipond growth units. Examination
of the RGP nitrogen removal as graphed in Figure 39# shows
that a similar decline in nitrogen uptake occurred; and, as
the pond warmed up, the nitrogen removal was resumed at its
former rate. This example was included to Indicate that
shallow algal cultures respond to changes in temperature.
MAXIMUM

  A
f
. V.' 
X * " t\
MINIMUM-
//! ..
r35
30
)
20 
r>
h-
<
QC
U
Q.
10 W
>-
 
15 '25 ' 5 ' 15^*25^"5 15 '25 ' 5 ' 15 '25 1 5 ' 15^5 5 15 '25 ' 5 ' 15 '25 ' 5 '15 '25 '5 15^25' 5 '15 "25 ' 5' T5'25 1 5"rT5125
JAN FEB MAR APR MAY	JUN JUL AUG	SEP OCT NOV DEC
MONTH
FIGURE 48- DAILY MAXIMUM AND MINIMUM TEMPERATURES IN RAPID GROWTH POND DURING 1969
-79-

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Algal Harvesting
Laboratory
Lime, alum, and ferric sulfate were primary coagu-
lants evaluated during the studies on harvesting of algae.
The concentration required of each of these chemicals to re-
move the desired amount of algae varied according to the
operating conditions of the growth units and was relatively
independent of the algal concentration. The range in con-
centrations is illustrated in Figure 49 in which is shown
the amount of each reagent needed to remove at least 90 per-
cent of algae from rapid growth pond samples. Although all
three chemicals proved to be effective coagulation aids,
each had distinctive characteristics which added to or de-
tracted from its usefulness.
The concentration of lime needed for the desired
level of removal varied from 20 to 200 mg/1 added as a 16,000
to 32,000 mg/1 slurry; however, during normal pond operation
(growth at or near the optimum level for maximum nitrogen
assimilation) the required concentration was 20 to 40 mg/1.
Calcium hydroxide raises the pH of a solution and causes algal
flocculation by precipitation of mineral salts at elevated
pH values; hence any addition to the growth unit that lowers
pH will tend to increase the amount of lime needed for floc-
culation. A comparison of two miniponds, one receiving C02
(pH of 8.0) and one not receiving CO2 (pH of 9.8) showed that
the lime requirement was Increased tenfold by the addition of
carbon dioxide. Overnight studies involving the use of lime
as a flocculent indicated that little diurnal change occurred
in chemical requirements, although the best separation with
lime took place in the afternoon when pond pH was maximum.
The high pH values associated with the addition of
lime result in the precipitation of potential inorganic algal
C 200-,
o
E
~ 160-1
|
Z
40-

	LIME
	ALUM
+ Fez(S04)3
+   
 +  
t,. .t
5 lis 251 5 15 sfe'i iFis' 5 15 25' 5 lE^TTl5l5T5_i5_Sr5_l5 251 5 15 25' 5 15 25' 5 IS 25' 5 15 25' 5 15 25'
MAR APR MAY JUN jut AUG SEP 0CT NOV DEC JAN FEB MAR
1969	MONTHS	1970
FIGURE 49 - CONCENTRATIONS OF MINERAL COAGULANTS REQUIRED FOR AT LEAST 90% ALGAL REMOVAL, JAR TESTS
-80-

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nutrients, particularly phosphorus. Analysis of water samples
before and after algal separation showed almost complete phos-
phorous removal, as well as removal of about one-half of the
original magnesium and total alkalinity. Although total hard-
ness was reduced by one-third, the calcium concentration was
lowered only slightly, presumably because the calcium carbon-
ate precipitate remained in the supernatant as a colloidal
suspension. The pH levels of a lime-treated effluent may also
be potentially detrimental in receiving waters because most
discharge requirements call for pH levels in the 6.5 to 8.5
range. If such discharge requirements are set for the plant
effluent, then pH adjustment by aeration, acid addition, etc.,
will be necessary. Some very preliminary work on the time
required for reaching a pH equilibrium by atmospheric exchange
indicates that it will be about two to three days. If this
time interval is adequate, the pH of the treated drainage
water would probably reach an acceptable range during transit
from the treatment plant to the Delta discharge point.
Aluminum sulfate, the second compound, causes coagu-
lation by the action of simple metal ions, e.g. (AlH20)6+3,
and by the formation of hydrolysis products which may be
highly charged, multinuclear, hydroxometal complexes (Stumm
and Morgan, 1962). Alum is normally more effective at low
pH levels. However, at Firebaugh the results indicated that
the cost of adjusting the pH to the optimum level was gener-
ally higher than the cost of the additional coagulant required
to achieve the same flocculation. Some preliminary work also
indicated that pH adjustment plus rapid mixing resulted in
release of soluble cellular nitrogen, thus lowering the over-
all efficiency of the nitrogen removal process. Alum concen-
trations to bring about 90 percent removal of algae ranged
from 5 to 200 mg/l, with 20 mg/l necessary when the pond was
operated at the best conditions for nitrogen assimilation.
Although pH values were lower when carbon dioxide was added,
alum requirements were slightly higher with than without the
C02. Alum flocculation removed dissolved phosphorus but was
not accompanied by a change in pH to an unacceptable level.
The third compound receiving extensive testing was
analytical grade ferric sulfate, its effective concentration
ranged from 1 mg/l to about 140 mg/1 (as Fe2(S04)3), depending
mainly on the presence of ferric chloride in the rapid growth
pond. As stated earlier, ferric chloride, 3 to 6 mg/l, was
added to the growth unit to enhance algal growth. It can also
be used as a coagulant. The dramatic effect of the addition
of iron to the RGP on required dosages of all three chemicals
is illustrated in Figure 50. Without any additional chemical
the removal level was increased from 40 to 70 percent, pre-
sumably as a result of coagulation by the ferric chloride
added for growth. The effective level of Fe2(S04)3 addition
-81-

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was reduced from 1^0 mg/1 to 5 mg/1. The concentrations of
both lime and alum were also reduced, with alum being effec-
tive at about 20 mg/1 and lime at 40 rng/l. The effect of
iron added to the RGP in terms of reduction in required
dosage for coagulation can also be seen in Figure 49. Al-
though only 1 mg/1 was added to the pond daily, the concen-
tration of total iron in the pond tended to increase
gradually. The growth studies indicated that as a necessary
addition to the algal ponds iron will provide a dual benefit.
Shown in Table 12 is the effect of adding lime,
alum, and iron on the cost of removing 90 percent of the
algae from one million gallons of plant effluent. (This
does not include the cost of the PeCl^ needed for algal
growth.) Figures in the table show that the cost of these
chemicals for separation of algae may be extremely low and
that ferric sulfate was the cheapest of the primary floccu-
lents tested. Additions of 1 to 5 mg/1 of this compound
would probably have little effect on the algae's potential
market value.
ioo! P- "grrrrS
?vV/
"&	A-	-A
20-
BEFORE FeCI3 ADDITION
AFTER
LIME
ALUM
Fez (504)3
SAMPLE DATA
BEFORE AFTER
DATE	JAN. 9,1970...MAR. 11,1970
pH	Q-8	9.85
VOLATILE
SOLIDS	175	265 mg/1
TEMPERATURE	10" C	I 1.5 C
0+
	,	
40
~ I20 ~ i^o
CONCENTRATION OF COAGULANT (mg/1)
~80~
200
~240
FIGURE 50-CONCENTRATIONS OF LIME, ALUM AND FERRIC SULPHATE
REQUIRED FOR 90% ALGAE REMOVAL - BEFORE AND AFTER
ADDITION OF FERRIC CHLORIDE TO RAPID GROWTH POND
-82-

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In addition to the mineral coagulants, approximately
60 polyelectrolytes were tested for their effectiveness as
primary coagulants, or in combination with lime, alum, or
ferric sulfate. A list of the common names of these com-
pounds, along with data on their effectiveness, either alone
or in combination with a mineral coagulant, is shown in
Table 13. Of the 60 polyelectrolytes, the 17 compounds listed
in Table 14 were found to be effective coagulants and their
costs were economically competitive with mineral coagulants
alone. In general, required concentrations of polyelectro-
lytes less than 10 mg/1 effected coagulation. If iron was
added to the growth pond, the effective concentration was
often as low as 0.05 to 0.25 mg/l. The list of polyelec-
trolytes in Table 13 includes some compounds that were
evaluated when iron was not added to the pond, and the results
may be somewhat misleading in that iron addition affects the
performance of the polyelectrolytes. Many of these polyelec-
trolytes will be tested again during calendar year 1970 to
determine whether their effectiveness is influenced by the
addition of iron to the growth units. One compound, Cat-Floe
(Calgon Corp.) was not beneficial in early testing, but with
the addition of iron to the RGP, more than 90 percent of the
algae could be removed with 0.5 mg/1 of this compound. The
cost of treating one million gallons of algal pond effluent
at this concentration of Cat-Floe would be about $2,00.
TABLE 12
ESTIMATED COST OF LIME, ALUM, AND FERRIC SULFATE
TO REMOVE 90 PERCENT TOTAL SUSPENDED SOLIDS
(Dollars/million gallons)
Date
Volatile
Solids
(rcgA) _
Alum
Lime
11/19/69
250
11.30
16.80
12/03/69
200
12.30
14.60
12/11/69
100
14.00
20.00
1/09/70
175
13.30
20.80
1/16/70
71
11.90
15.90
1/23/70
91
12.70
8.20
2/11/70*
114

3.50
2/18/70
210
13.00
5.^0
2/25/70
417
2.35
1.30
3/11/70
265
3.46
3.28
3/18/70
229
3.50
3.23
Ferric
Sulfate
14.40
25.00
23.80
22.30
24.40
11.00
1.50
0.25
0.40
0.62
Commenced 1 mg/1 daily addition of Fe+-H- to"
pond on 1/26/70.
-83-

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TABLE 13
RESULTS OF LABORATORY TESTS OF THE EFFECT OF
VARIOUS POLYELECTROLYTES OK FLOCCULATION AND
SEDIMENTATION OF ALGAL CULTURES AT THE IAWTC
/ Tested
Effect on Flocculationi/ Cone. Range
Compound	Company	Alone:With Alum:With Lime (mg/l)
Cat-Floe
Calgon Corp.
B
B
B
1-10
ST-260
Calgon Corp.
N
N
N
0.25- k
ST-266
Calgon Corp.
N
N
N
1-10
ST-269
Calgon Corp.
N
N
N
0.25- k
ST-270
Calgon Corp.
N
N
N
0.25- 1+
Drevfloc-3
Drew. Chem. Corp.
N
N
N
0.25- 5
Hercofloc 810
Hercules, Inc.
B
B
B
0.25-10
Hercofloc 8lU
Hercules, Inc.
B
B
B
0.25-10
Kelgin W
Kelco Company
N
N
N
0.25-10
Kelcosol
Kelco Company
N
N
N
0.25-10
Kelzan
Kelco Company
N
N
N
0.25-10
Kelco SCS MV
Kelco Company
SB
SB
SB
0.25-10
Kelco SCS HV
Kelco Company
SB
SB
SB
0.25-10
Ferrifloc?/
Tennessee Corp.
B


5-60
Chitosan
Chem. Res. Lab, Inc.
N
N
N
1-10
Gendriv 162
General Mills, Inc.
B
N
B
1-10
Genfloc 139
General Mills, Inc.
N
N
N
0.5 - 5
Genfloc lUo
General Mills, Inc.
N
N
N
0.5 - 5
Genfloc 155
General Mills, Inc.
B
B
B
0.5 - 5
Genfloc 156
General Mills, Inc.
B
B
B
0.5 - 5
Genfloc 292
General Mills, Inc.
H
B
B
0.06- 2
Polyhall M-293
Stein-Hall
N
N
N
O.25-IO
Polyhall M-U02
Stein-Ha11
N
N
N
O.25-10
Polyhall M-603
Stein-Hall
N
N
N
O.25-IO
Jaguar MDD
Stein-Hall
N
N
N
O.25-IO
Jaguar MAL-22-A
Stein-Hall
N
N
N
O.25-IO
Primafloc C-3
Rohm and Haas
B
B
B
1-10
Primafloc C-5
Rohm and Haas
B
B
B
1-10
Primafloc C-7
Rohm and Haas
B
B
B
0.5 - 5
Zeta floe WN-2
Narvon Mines, Ltd.
N
N
N
5-80
Zeta floe C
Narvon Mines, Ltd.
N
N
N
5-80
Zeta floe 0
Narvon Mines, Ltd.
N
N
N
5-80
-84-

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TABLE 13 (Continued)
Compound
Company-
Effect on Flocculationi/
Alone:With Alum:With Lime
Tested
Cone. Range
(-mg/l)
Nalcolyte 603
Nalco
Chemical Co.
N
B
N
1-10
Nalcolyte 610
Nalco
Chemical Co.
N
N
B
1-10
PEI 6
Dow Chemical Co.
N
N
B
0.5 - 5
PEI 12
Dow Chemical Co.
N
N
B
0.5 - 5
PEI 18
Dow Chemical Co.
N
N
B
0.5 - 5
PEI 600
Dow Chemical Co.
B
B
N
1.0 -10
SA 1188-1A
Dow Chemical Co.
N
N
N
1.0 -10
SA 1188-1F
Dow Chemical Co.
N
N
N
1.0 -10
SA 1569
Dow Chemical Co.
N
B
N
1.0 -10
NC 1621
Dow Chemical Co.
N
B
N
1.0 -10
Purifloc A-21
Dow Chemical Co.
B
B
N
0.5 - 5
Purifloc A-22
Dow Chemical Co.
B
B
N
0.5 - 5
Purifloc C-31
Dow Chemical Co.
B
B
N
1-10
Purifloc C-32
Dow Chemical Co.
B
B
N
0.5 - 7
Purifloc NP-10
Dow Chemical Co.
B
B
N
0.5 - 5
Wisprofloc-202/
Ionae Chemical Co.
N
N
N
1-10
Wisprofloc-P
Ionae
Chemical Co.
N
N
N
1-10
Burtonite #78
Burtonite Co.
N
N
N
1-10
Magnifloc 530C
Amer.
Cyanamid
N
N
N
0.06- 2
Magnifloc 837A
Amer.
Cyanamid
N
N
N
0.5 - 5
Magnifloc 521C
Amer.
Cyanamid
B
B
B
0.06- 2
Magnifloc 82OA
Amer.
Cyanamid
B
N
B
0.06- 2
Magnifloc 83 5A
Amer.
Cyanamid
B
N
B
0.06- 2
Magnifloc 836A
Amer.
Cyanamid
B
N
B
0.06- 2
Magnifloc 86OA
Amer.
Cyanamid
N
N
N
0.06- 2
Magnifloc 87OA
Amer.
Cyanamid
N
N
N
0.25- U
Magnifloc 900N
Amer.
Cyanamid
N
N
N
0.06- 2
Magnifloc 95N
Amer.
Cyanamid
N
N
N
0.06- 2
Magnifloc 985N
Amer.
Cyanamid
N
N
N
0.06- 2
Magnifloc 990N
Amer.
Cyanamid
N
N
N
0.06- 2
1/ The effect has been denoted as N - no help. B - beneficial; and SB -
slightly beneficial. A beneficial effect was shown if the polyelectro-
lyte aided flocculation and sedimentation with no consideration to the
economic aspects of its use.
2/ Not a true polyelectrolyte, but a crude iron sulfate ore.
2/ Not commercially available in this country.
-85-

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TABLE 14
POLYELECTROLYTES WHICH WERE EXPERIMENTALLY BENEFICIAL
AND ECONOMICALLY FEASIBLE AIDS IN ALGAL SEPARATION




Effective
(mg/l) :
Primary Coagulant
Manufacturer
Name
:Concentration Range
I
Aluminum:
Calcium



1/
2/
3/
None:
Sulfate:
Hydroxide
Dow Chemical Co.
PEI 600

5-10
0.5-4.0

X
X

Dow Chemical Co.
C-31

0.5-5
0.25-4.0

X
X

Dow Chemical Co.
C-32

0.5-5
0.5-1.0

X
X

Nalco
Nalcolyte
603

4-8


X

Nalco
Nalcolyte
610


4-6


X
Rohm and Haas
Primafloc
C-3

4-8


X

Rohm and Hsas
Primafloc
C-5
4-8
0.25-4.0

X
X

Rohm and Haas
Primafloc
C-7
5-10
O.25-I.O

X
X

Hercules, Inc.
Hereofloc
8l4
1-3
0.5-1.0
0.5-1.0
X
X
X
General Mills, Inc.
Glenfloc
155

0.25-1.0
0.25-1.0
X
X
X
General Mills, Inc.
Glenfloc
156

0.25-1.0
0.25-1.0
X
X
X
General Mills, Inc.
Glenfloc
162

0.25-1.0
0.25-1.0
X
X
X
American Cyanamid Co.
Magnifloc
521C*

0.25-1.0
0.25-1.0
X
X
X
American Cyanamid Co.
Magnifloc
820A**


0.25-1.0


X
American Cyanamid Co.
Magnifloc
835A**


1-2


X
American Cyanamid Co.
Magnifloc
836A**


1-2


X
Calgon Corp.
Cat-Floe

0.05-
04 0.05-1.0

X
X
X
1/ Alone
2/ With alum
3/ With lime
*C - cationic
**A - anionic

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Polyelectrolytes reduce the zeta potential, a term
involving the net particle charge and the distance the charge
affects the surrounding medium (Pair, et al, 1968), thus
allowing colloids to approach each other "(Black, et al* 1965).
The polymer molecules also aid in the formation oT~three di-
mensional algal-polymer matrices by attaching themselves to
the surface of algal cells (Tenney, t al, 1969). Algae
generally have a net negative surface cTiarge (Ives, 1956).
Tenney, e_t al (1969) found that only cationic (positively
charged) polyelectrolytes effected algal flocculation. At
Pirebaugh cationic polymers were generally more effective;
however, three anionic compounds (Magnifloc 820A, 835A, and
836A) did cause coagulation and floe formation when used with
lime. Tenney, t al, (1969) also noted that polymer effective-
ness was affected By pH (more effective at lower pH ranges),
algal concentration, and algal growth phases . Their observa-
tion that there was an optimum polyelectrolyte concentration
was not demonstrated at the IAWTC; however, the range of tested
concentrations was relatively narrow, and we may have missed
the decline at higher levels of polymer since we tried to stay
within an economic range.
The use of polyelectrolytes to flocculate algal sus-
pension was often the most economical means of achieving algal
removal. However, their use may be limited if the algal
product is to be used as an animal food. Studies need to be
conducted to determine whether cationic polyelectrolytes are
toxic to animals. Preliminary tests run by Golueke, et al
(1964), in which the cationic polyelectrolyte Sondellite was
used, showed that single massive dosages of the compound had
no demonstrable effect on weanling rats. Daily dosage at a
level comparable to that which the animals would receive from
Sondellite-flocculated algae resulted in no apparent symptoms
for a period of two weeks, the duration of the test. Another
possible problem which might arise through the use of poly-
electrolytes would be the rise in effluent dissolved nitrogen
caused by the nitrogen in the polyacrylamide and polyamide
compounds. At the effective concentration used at Flrebaugh.
this problem should not be significant.
Pilot Scale Separation Studies
Sedimentation Tank. This unit was not evaluated
as a separation device per se, but was used to control bio-
mass levels in the rapid growth pond. Water from the RGP
was pumped into the unit at flows which gave theoretical
hydraulic detention times in the tank of two to six
hours. The algal sludge from the unit ranged up to 3 to 5
percent solids, depending on retention time of the solids.
The sun-dried algal product from this unit provided a
-87-

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source of algae for companies interested in studying the
market potential of the product. From March 10 through
March 14, 1970, a total run time of 95 hours of sedimentation
in this unit resulted in the removal of about 50 pounds
(22.7 Kg) of suspended solids from the RGP. Detention time,
within the range of two to six hours, did not have any apparent
effect on sedimentation.
The composition of the sedimentation tank effluent
may be more significant than the actual production of an algal
product. The suspended solids in the Influent and algal sludge
were about 50 to 70 percent volatile material (algae)j whereas
the effluent suspended solids were often as high as 90 percent
volatile and consisted of the younger, actively growing
Scenedesmus cells. The tank thus helped in the removal of the
older and heavier cells, as well as the suspended chemical
precipitates and clay particles from the growth pond.
Shallow Depth Sedimentation Tank (Water Bo.v). The
shallow depth sedimentation tank was operated on-line with
the rapid growth pond in a system in which growth pond ef-
fluent passed through the unit and then was discharged from
the site. The algal slurry was used in the testing of the
effectiveness of various dewatering devices. As described
previously, separation was effected by coagulation-
flocculation, with subsequent filtration of supernatant to
remove cells not trapped in the algal floe.
The percent transmittance of light through effluent
and influent samples over a three-month period is shown in
Figure 51. The influent varied from about 150 to 700 mg/1
total suspended solids. After the first of February 1970, the
the growth pond was swept daily; thereafter, the suspended
solids concentration in the influent to the sedimentation
tank were consistently in the 300 to 500 mg/l range. During
this period of the study the effluent from the Water Boy
(after the mixed-media filter) usually contained less than
10 mg/l volatile solids (less than 20 mg/1 total solids).
Influent flows varied from 1.4 to 3 gPm with resulting
hydraulic detention times of 93 to 44 minutes. At the flow
rates tested, removal of algae was relatively independent of
influent solids in the range of 150 to 700 mg/l.
The effluent data shown in Figure 51 were products
of both sedimentation and filtration. Samples of the effluent
of the sedimentation chamber indicated that more than 90 per-
cent of the suspended solids were removed by coagulation-
sedimentation and that the filter acted as a final polishing
device. The mixed-media filter remained in the Water Boy
until May 12, 1970> at which time the entire filter was re-
moved because of plugging of the pores by what appeared to be
-88-

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IOO-i
\ X
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TABLE 15
WATER BOY OPERATIONAL CRITERIA AND
RESULTS FOR MAY 1970
Date :	:	 Chemicals	:	:
of : Plow Rate : Fe2(S0i|)3 : Cat-Floe : Percent:Total Solids
Sample: GPM : mg/1 : mg/1 :Removal:in Eff. mg/l
1	2.0	3.0	1.5	100	0
2	2.5	2.5	0.6	100	0
3	2.5	3.0	0.7	100	0
4	2.5	3.0	0.6	100	0
5	2.5	3.5	0.5	100	o
6	3.0	3.5	0.6	100	0
7	3.0	3.5	0.4	100	0
8-12	- - - - Shut Down - Removed Sand Media  -
13	2.5	3.0	0.7	97	21
14-15 2.0	3.0	0.7	97	21
16	2.0	3.0	0.8	99	7
17	2.0	4.5	0.7	99	7
18	2.5	3.0	0.6	99	7
20	2.5	0.8	0.7	97	22
21	2.5	2.0	0.7	98	15
22	3.0	2.5	0.6	98	15
25	3.0	2.0	0.4	97	27
26	3.0	2.5	0.4	98	18
27	3.0	4.5	0.4	99	9
28	3.0	5.0	0.3	98	18
29-31 3.0	5.0	0.4	100	o
four different coagulants  alum, lime, Ferrifloc, and Cat-
Floc  on two different days. On both days, ferric sulfate
was the most economical of the compounds tested. In the
test, the Fe2(S04)o was derived from Ferrifloc, and the dos-
age was measured as pounds of total material (i.e., Ferrifloc)
which Included waters of hydration, other compounds, etc.
Laboratory comparisons (jar tests) of analytical grade Fe2
(S04)3 and Ferrifloc indicated that the latter material was
the more effective of the two materials.
The Water Boy was routinely backwashed every 48
hours and the algal slurry was sampled for total suspended
and volatile solids. Some of the results of these analyses
are given in Table 17. The slurry taken directly from the
tubes contained from 1.1 to 1.6 percent solids with 50 to 60
percent of the solids as volatile material. Overnight set-
tling of the slurry increased the percent solids to 4 to 6
percent; however, tests of settling as a function of time
-90-

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TABLE 16
COST COMPARISONS OF FOUR COAGUL/ PS
(Dollars per Million Gallorr
1 
 *
: Cost for :

: : Cost
:40 mg/1 Effluent:

: :for 90$
:Total Suspended :
RGP
Date :Coagulant :Removal
: Solids i
Variables
5/20/70 A12(S02|)3
8.83
14.45
PH
9.45
Ca(0H)2
6.22
7.02
Temp.
15.5C
Fe2(SOl03
0.77
0.77
Total
Solids
726 mg/1
Cat-F'loc
2.17
5.93
Settleable
Solids 14.0 ml/1
5/27/70 Al2(SOi4)3
2.59
9.26
PH
9.35
Ca(0H)2
3.06
3.88
Temp.
17.1C
Fe2(S0/j )3
0.16
0.48
Total
Solids
999 mg/l
Cat-Floe
1.44
3.87
Settleable
Solids 19.0 ml/1
TABLE 17
COMPOSITION OF SLURRY FROM WATER BOY
:Total Suspended
Date : Solids - mg/1
Volatile Solids
mg/1
Percent
Volatile
3/11/70
3/12/70
3/16/70
3/17/70
3/18/70
3/19/70
3/20/70
3/21/70
3/22/70
3/23/70
3/2V70
3/25/70
16,000+
44,600
14,200
56,000
16,200
47,700
13,900+
42,300
11,200+
33,500
14,800+
37,900
8,4oo
22,800
6,800
24,000
7,600
22,400
8,600
26,300
6,400
20,000
8,600
18,700
53
51
48
43
47
47
62
62
57
60
58
57
+ = Slurry directly from unit,
o = Slurry after settling for 24
hrs. and decanting off liquid,
-91-

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indicated that 80 percent of the settling occurred in the
first six to eight hours. The settled slurry normally was
held in a collecting sump to provide material for the de-
watering studies.
Upflow Clarifier. The upflow clarlfier used de-
tention times of about one hour to remove up to 95 to 100
percent of the suspended solids and produced a slurry con-
taining 2 to 10 percent solids. The only chemical additive
tested was sodium hydroxide, and the required concentration
varied from 100 to 200 mg/1.
Although the unit did do an effective job of re-
moving the algae, many operational problems arose which pre-
vented long run times. Ihe most serious of the problems was
caused by the consolidation of the algal mat, which increased
hydraulic pressure on the mat and caused sloughing of the
algal floe. The algae were then carried over with the efflu-
ent. When this occurred, normal operation of the unit did
not resume until the mat was built up again. Until this
problem is solved, the upflow-clarifier type of separation
device cannot be used as a reliable method of removing algae.
The upflow clarifier will be remodeled in an attempt to
eliminate plugging.
Upflow Sand Filter (Sanborn Filter). Preliminary
studies on the Sanborn filter constructed by Bohna began in
June 1969 with an algal suspension from the rapid growth pond
as the test medium. Although the early studies showed that the
filter could separate the algae from the liquid phase, some
problems soon developed, especially those concerned with back-
washing of the unit. In the first runs, the filter was plugged
after relatively short run times, and the algal concentration
in the backwash was low. The low concentration in the backwash
may have been caused by too large a backwash volume and by the
plugging due to biological fouling. Washing the sand bed with
dilute sulfuric acid did not lead to an increase in run time.
However, treatment with dilute hypochlorite solution did remove
the fouling materials.
Personnel from Bohna assisted in testing the filter
for four days in the fall of 1969. As shown in Table 18, at
a flow rate of 1 gpm, the unit consistently removed more than
90 percent of the influent solids. The loading on the surface
of the filter during this test period ranged from 107 to
595 mg/ft^/min with an average of 364 mg/ft^/min. The varia-
tion in influent concentration was the result of the mixing
cycle used in the RGP. Run length was limited by the maximum
design pressure of the unit and varied from 40 to 100 minutes,
-92-

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TABLE 18
SUMMARY OF OPERATIONAL DATA FOR
SANBORN FILTER, 11/17-21/69
Run No.
: Feed
:Concentration
: Suspended
: Solids, mg/l
:Filtrate:Percent:
:Concen- :Removal:
jtration : Susp. :
: mg/l :Solids :
Backwash
Volume
(Gal.)
: Backwash
: Cone. Total
: Susp. Solids
: (mg/l)
11/18/69-1
156
3.7
97.8
2.1
6,500
ll/l8/69-2
113
0.5
99.6
1.7
7,200
11/18/69-3
549
1.4
99.8
2.4
11,600
ll/l8/69-4
556
2.8
99-5
2.5
17,700
H/18/69-5
572
11.9
98.0
3.0
13,700
ll/19/69-l
142
1.7
98.8
1.9
6,360
11/19/69-2
565
0.7
99.9
2.5
11,100
11/19/69-3
395
14.5
96.5
3.8
6,680
11/19/69-^
629
1.7
99.8
4.25
5,330
ll/20/69-l
138
1.6
98.6
2.7
4,880
11/20/69-2
1*56
0.2
99.6
2.6
4,330
11/20/69-3
548
0.5
99.9
1.8
10,700
11/20/69-4
615
2.4
99.62
2.4
10,600
11/20/69-5
293
15.9
95.2
3.3
5,780
ll/21/69-l
211+
19.0
92.0
3.2
4,260
11/21/69-2
214
16.2
93.0
1.8
5,230
-93-

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depending on the solids
concentration in the
influent. The rela-
tionship between run
time and solids concen-
tration is shown in
Figure 52. Above 400
mg/1 suspended solids
the run time was uni-
formly near 40 minutes.
Routine operation of an
algal stripping plant
would probably be char-
acterized by a more
uniform solids concen-
tration, and thereby a
stabilization of the
run time.
The data in
Table 18 also show the
concentration of algae
in the backwash. In
this particular study,
the unit was manually backwashed when the pressure differen-
tial across the filter reached 20 pounds per square inch (psi).
Based on a one-run study of the effect of the backwash volume
of two gallons (0,5 gallon/ft2) was selected as the optimum
quantity. Concentration of suspended solids in the backwash
was roughly proportional to the initial feed concentration
and varied from 0,4 to 1.7 percent solids. These concentra-
tions could probably be increased to 3 to 5 percent with
smaller backwash volumes and automated control of the back-
washing cycle.
Another intensive five-day study of the Sanborn
filter was conducted in March 1970, at which time flow rates
of 1.0 and 1.25 gpm were tested. The one-gpm throughput used
in the previous study corresponds to the manufacturer's design
criterion of 0.25 SPm P*1 square foot of filter surface. In
this second run, the unit was operated on a continuous basis.
The results were similar to those obtained in the November
study with about 95 percent solids removal and backwash con-
centrations of 1.2 to 2.7 percent solids. After five days
of operation, solids buildup within the unit appeared to
cause a gradual decrease in run time and the buildup appeared
to be significantly faster at the higher influent rate. Pre-
liminary estimates showed that the expected filter life would
be about 200 days at 1 gpm, compared to about 50 days at
1.25 gpm.
-94-
I lO,
100-
70-
60-
50-
20-
10-
100 150 200 250 300 350 400 450 500 550 600 650 70O
TOTAL DISSOLVED SOLIDS (mg/l)
FIGURE 52- RELATION BETWEEN SOLIDS LOADING AND RUN LENGTH,
SANBORN FILTER

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Vacuum Filter. Preliminary opera!.icn of the
Eimco vacuum filter yielded disappointing esults. No build-
up of algal cake occurred on the filter boj.f, and the con-
centration of algae in the effluent was orly slightly less
than that in the influent. Microscopic examination of the
algae and of the belt material indicated that the mesh
openings in the belt were too large to retain the small (10
to 25 micron) algal cells. Based on these results, a multi-
filament nylon belt (Eimco No. NY-319F) with a pore size
similar to that of the algae was substituted for the existing
porous polypropylene belt. The unit was then evaluated as
a means of dewatering a concentrated algal slurry.
The first three-day run with the new belt was com-
pleted in January 1970. The feed concentration was about
3,000 mg/1 (03 percent solids). Although this concentration
of algae was considerably lower than the planned 3 percent
solids from the concentration step, it did permit an estima-
tion of the unit's effectiveness at lower algal loadings.
The unit appeared to take up water at a fixed rate of about
1 gpm, resulting in a suspended solids loading on the belt
of about 3i720 mg/ft2/min. Only one belt speed, 2.9 ft/min
(or 2.8 ftVmin), was evaluated. During this test period,
the unit produced sludge containing 18 to 25 percent solids
and removed 90 to 95 percent of the influent suspension,
with an average concentration of solids in the effluent of
about 300 mg/1. The high removal rates were obtained with
a vacuum of 15 to 20 inches of mercury (Hg). When the vacuum
was lowered to 5 inches of Hg, less than 20 percent of the
algal solids were removed (influent  3,880 mg/1; effluent 
3,080 mg/1).
A second run was completed in which an algal slurry
of about 30,000 mg/l (3 percent solids) was used. The re-
sulting loading on the belt was 17,500 mg/ft2/min. The
vacuum in this run was 20 to 25 inches of Hg, or about 5
inches more than in the previous test. The unit removed
about 90 percent of the influent solids and produced an algal
product containing 24 percent solids. In both of these tests,
effluent from the unit contained algal concentrations higher
than allowable and would thus have to be recycled to the
head of the sedimentation area.
Microscreen. The Zurn microscreen tested was sup-
plied with screens of two pore sizes, 25 and 35 microns.
Operation of the unit soon showed that algae were passing
through the finer screen. Removals up to 30 percent were
obtained, but most of this was due to algal settling in the
influent and effluent chambers. A more complete evaluation
-95-

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of this unit will require smaller pore-size screens, and
perhaps testing of different pore sizes in series.
Centrifuges. Two of the centrifuges tested at
Firebaugh, the continuous solid bowl type and the nozzle
type, were not particularly efficient at either primary
concentration or dewatering. With the solid bowl centrifuge,
algal slurries with concentrations of 3 to 22 percent solids
were obtained, but the highest removal rate was only 28
percent. A major difficulty with the nozzle-type centrifuge
was the plugging of the nozzle openings and consequent
failure of the machine to remove algae. The plugging did
not appear to be caused by the algae but by colloidal clay
particles in the water.
A third type of machine, the De Laval self-cleaning
centrifuge, was evaluated as both a primary concentration and
as a dewatering device. Used as a primary concentrator, the
unit removed up to 95 percent of the influent (concentration,
800 mg/1). The flow through the unit was 6 gpm and the ef-
fluent slurry contained about 10 to 12 percent solids. Oper-
ation was continuous under these experimental conditions.
A problem in plugging was encountered because of incomplete
discharge of the material in the bowl. This problem was re-
solved through the use of a so-called "double shoot", in
which the machine was programmed to partially discharge the
contents followed by a complete ejection. Using this double-
shoot concept, a product with up to 17 percent solids was
obtained, but the percent removal dropped to the low 80's.
The self-cleaning centrifuge was also tested with
an influent containing 20,000 to 30,000 mg/1 influent solids.
In this test the flow rate was 2.75 gpm> while the slurry
contained about 10 percent solids. At this loading, the
centrifuge removed more than 98 percent of the influent sus-
pended solids.
Oswald and Golueke (1968) reported on the use of
centrifugation to separate sewage-grown algae. With a 30-
inch bowl industrial centrifuge, they were able to remove 64
to 84 percent of the influent algae (200 mg/l), depending on
the throughput rate. Power requirements varied inversely with
the original algal concentrations. They concluded that the
use of centrifugation to separate algae was limited by the
high initial cost of the units and by operational power re-
quirements, although development of new power sources (atomic)
and changes in centrifuge technology might make the use of
centrifuges economically feasible in the future. Centrifuges are
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desirable concentrating and dewatering devices because of
their simplicity of operation and high product quality.
Drying
Algae samples for distribution normally were ob-
tained by draining the contents of the sedimentation tank
onto a 2-foot by 2-foot section of cloth supported on a wire
frame. A 2-inch layer of 5 percent solids algae sludge dried
to about 85 percent solids in two to three days. On one
occasion an attempt was made to quantify the drying rates of
the algal slurry on various types of surfaces. Three layers
of algae, 0.5* O.75, and 1.25 inches thick:, were tested on
12-inch diameter circles of asphalt, cloth, black plastic,
and sand. Table 19 shows the number of hours required for
the sludge to go from about 6.4 percent solids to 80 percent
solids. The sand drying method was by far the fastest, but
it also resulted in a product having a considerable amount
of entrapped sand particles. The asphalt surface would
probably be the most practical because the algae could be
applied and scraped off with less difficulty. The foregoing
study was conducted in May 1970 and will be repeated to de-
termine seasonal variations in drying time,
A dewatered sample of algae sent to the De Laval
Corporation for spray drying contained 10 percent solids and
was dried to 90 percent solids with no difficulty. Operators
of the unit did not expect any problems when drying an algal
sludge containing 20 percent solids.
Regrowth Studies
Laboratory studies designed to determine whether
nitrogen removal actually lowered the potential of tile
TABLE 19
NUMBER OP HOURS REQUIRED TO REACH 80$ SOLIDS,
FROM ORIGINAL SAMPLE CONTAINING 6.4# SOLIDS
:	 Type of Surface	
Slurry Depth (inches) : Asphalt : Cloth:Black Plastic: Sand

0.5
46
36
42
24

0.75
No sample
43
No sample
50

1.25
72
65
73
80
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drainage water to stimulate or support additional algal
growth in waters of the Sacramento-San Joaquin Delta were
integral parts of the nitrogen removal investigation. The
objectives of these regrowth studies were to determine if:
(1)	tile drainage stimulated growth of algae in Delta waters;
(2)	if the treatment processes studied at Firebaugh lowered
the stimulatory effect; and (3) if the waste did prove stimu-
latory, could the effect be attributed to any particular
constituent (or constituents)? Some of the preliminary re-
sults of these studies were presented by Brown, ej; al, 1969,
and a comprehensive final report will be published In 1971.
A review of results pertinent to algal stripping will be
discussed here.
Fourteen separate algal assays were conducted during
a one-year period at EPA (Alameda) and DWR (Bryte) labora-
tories. The general procedure in these assays included col-
lecting water samples from the Sacramento-San Joaquin Delta
near Antioch (site of proposed drain discharge) and trans-
porting the samples to the laboratory where they were mixed
with various percentages (from 1 to 50 percent) of treated
(algal growth and harvesting) and untreated tile drainage.
Triplicate flaslcs of the different mixtures were then incu-
bated at 24+ 1C and under continuous fluorescent lighting
(approximately 300 to 400 ft-c). No additional algal seed
was added. Growth responses of the Indigenous algae were
then followed by observing daily changes in in vivo chloro-
phyll fluorescence as measured directly In aTluorometer
without chlorophyll extraction. The samples were Incubated
until the chlorophyll levels had peaked. The peak values of
the mixtures of tile drainage and Delta water were compared
with those of controls, which contained only Delta water.
A statistical examination of the data indicated that in vivo
fluorescence was directly correlated to cell number (r =
0,69) and that either peak fluorescence or change in fluores-
cence could be used to determine the extent of growth response
in the cultures.
Figure 53 Illustrates the results of a typical
batch algal assay of an algal pond effluent with a relatively
high inorganic nitrogen concentration of 3.7 mg/l. In this
particular experiment, all additions of tile drainage stimu-
lated algal growth in the Delta waters; although the peak
fluorescence values for any particular dilution were lower
with treated effluent than with untreated tile drainage. The
results also show that the maximum algal growth occurred when
0,8 mg/l nitrogen was added. Above this level, some other
nutrient (or light) presumably became limiting. Cell counts
made at the beginning and end of the assay Indicated that the
cultures retained the original proportion of algal groups
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>^J0.87)
20CH
z
3
Ul
o
z
K
O
3
100-
(0.37)
(0.32)
"A
100% DELTA
98% DELTA
95% DELTA
90% DELTA
ALGAE POND WATER ADDED (TREATED)
TILE DRAINAGE ADDED (UNTREATED)
NOTE : TOTAL INORGANIC NITROGEN (mgN/l) IN
THE SAMPLES ARE IN PARENTHESES
ADJOINING THE APPROPRIATE CURVE
Tl ME - DAYS
FIGURE 53-ALGAL GROWTH RESPONSES OF VARIOUS COMBINATIONS OF UNTREATED AGRICULTURAL
TILE DRAINAGE AND "ALGAL STRIPPED" WATER MIXED WITH SAN JOAQUIN RIVER WATER (DELTA)
(75 percent diatoms, 25 green and blue-green) during the in-
cubation period, regardless of the composition of the culture
medium.
The results of all assays using algal system efflu-
ent showed that treatment by such a process reduced the bio-
stimulatory nature of the tile drainage. When the dissolved
inorganic nitrogen content of the effluent was reduced to
2 mg/1 or less, additions of as much as 20 percent of algal
system effluent usually had little stimulatory effect on
algae in Delta waters. Peak fluorescence values of Antioch
controls (incubated samples of Delta water only) were rela-
tively stable throughout the 12-month period, although the
initial levels varied seasonally. Samples collected in the
winter and early spring contained 0.5 to 1.0 mg/1 inorganic
nitrogen and had low initial fluorescence levels; whereas,
samples collected during the summer and fall had little
inorganic nitrogen (0.2 mg/l), and the initial fluorescence
level was similar to the final level. At the low inorganic
nitrogen concentration found in water from the western Delta
during the summer months addition of nitrogen-rich tile
drainage caused stimulation of algal growth in laboratory
cultures.
In many of the assays, the treated tile drainage
was restored to its original nitrogen level by the addition
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of analytical grade potassium nitrate. Comparison of fluo-
rescence values in cultures grown in the "respiked" drainage
with those grown in untreated tile effluent did not show any
significant difference in stimulatory effect; the data
indicated that nitrogen was the important biostimulatory
substance in the drainage, at least with respect to the con-
ditions of the test. In many of the assays, water treated
by bacterial denitrification and algal stripping were com-
pared for their effect on algal growth. Denitrification
results in the removal of nitrogen only; whereas, theoret-
ically, algal stripping leads to the removal of nitrogen as
well as other nutrients essential for algal growth. Statis-
tical analyses of the results demonstrated that, if the
nitrogen levels of the effluent were comparable, growth re-
sponses in the cultures were similar. That is, effluent
nitrogen level, not the method of treatment, was the important
factor.
Botulism Studies
The results of botulism studies conducted by
California Department of Fish and Game are contained in a
progress report titled "Applied Botulism Research Including
Management Recommendations , published by the Wildlife Manage-
ment Branch in January 1970. The study was initiated to
determine whether algal growth units would provide a suitable
habitat for the bacterium, Clostridium botulinum. Type C.
This organism, which often occurs in shallow bodies of water
in the San Joaquin Valley, can cause waterfowl mortalities,
where the necessary ecological conditions are present. The
most significant such condition is complete anaerobiosis,
although Clostridium botulinum can survive in the presence
of oxygen if an electron acceptor or reducing substance is
also present. The required anaerobic condition in waterfowl
areas normally is satisfied by the remains of dead organisms,
either plant or animal, that are killed as the water level
in the area is raised and lowered. Carcasses of birds, fish,
and mammals are the primary means whereby the botulism bac-
terium is perpetuated, because they provide anaerobic con-
ditions necessary for its growth. The botulism-infected
maggots living in the carcasses are ingested by healthy water-
fowl, which then sicken and die. The botulism outbreak is
then continued as the bacteria multiply in the carcasses of
the recently killed waterfowl.
Fish and Game personnel made periodic trips to the
Firebaugh site and sampled for the presence of Clostridium
botulinum with consistently negative results. Injection of
pure toxin into the pond water showed that the toxic effects
of the material disappeared within a period of time varying
from a few minutes to four hours.
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Although Clostridium botulinum was never observed
in the ponds at the IAWTC, even in the pond containing soil
from a known botulism area, the report concluded that the
shallow algae ponds could be potential problem areas if
proper precautions were not exercised. It was recommended
that the ponds be constructed with levee slopes of 30 or
more and that carcasses of birds, fish, or mammals be removed
immediately. If these recommendations are followed, water-
fowl will be able to use the algae growth ponds as resting
areas without suffering detrimental effects.
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CHAPTER V - DISPOSAL
The use of photosynthetic organisms to remove
nutrients from agricultural drainage water produces an algal
by-product. Estimates for an algal stripping plant to treat
the total tile drainage from the San Joaquin Valley in the
year 2000 indicate that between 66,000 and 83,000 tons of
algae will be produced annually, or about 200,000 cubic yards
of material which would require disposal each year. This
material may prove to be a valuable by-product and thus
defray part of the total treatment cost. Actual project
research into the disposal possibilities was limited to
reviewing literature, making an economic evaluation of poten-
tial markets, and supplying product samples to interested
companies. Thirty-five companies with a potential interest
in an algal product were contacted to determine the extent
of their interest. These included feed and grain dealers,
representatives of the chemical industry, and companies
distributing or producing soil supplements. About one-third
indicated they would like samples for further evaluation.
These samples were provided. The results of their studies
have not been received as yet. In the following paragraphs,
some of the possible uses will be discussed, along with
estimated market potentials.
Animal Food Supplement
Marine algae have often been used to provide food
for humans and livestock, especially in the Asiatic areas of
the world. Recently, interest has been shown in the possible
uses of unicellular algae as a protein source, especially
for livestock feeding. This interest arises from the fact
that algal production is a more efficient use of land than
is conventional farming (in terms of biomass produced per
unit area) and substrates for algal growth are often readily
available in the form of domestic sewage and other liquid
wastes. Usually these substrates are fairly complete and
need only minimal supplemental growth factors. Although
algae produced from these systems may have a variable compo-
sition, normally their protein content is quite similar to
that of commonly used protein supplements, soybean meal and
cottonseed meal, for instance. An idea of the composition
of an algal product can be obtained from the results of
analyses described in the following paragraphs.
Hintz, et al, (1966) reported on the analysis of
several samples oFHsewage-grown mixtures of Scenedesmus and
Chlorella. The results of these analyses are shown in"
Table 20. The composition is similar to that reported by
other authors, although the composition varies with the
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TABLE 20
COMPOSITION OP ALGAE GROWN ON SEWAGE3' b
4

Component :
: Number of
s Samples

#

Percent
Crude Protein
25
50.93
+ 0.68
Crude Fiber
25
6.20
+ 0.41
Ether Extract
25
6.01
+ 0.40
Ash
25
6.24
+ 0.74
Cellulose
10
3.33
+ 0.25
Lignin
10
4.21
+ 0.30
Calcium
10
1.93
+ 0.19
Phosphorus
10
2.22
+ 0.10
Silica
10
1.73
+ 0.21
Magnesium
Composite
1.60

Potassium
Composite
0.92

Iron
Composite
0.23

Sodium
Composite
0.23

Zinc
Composite
0.18

Aluminum
Composite
0.12

Manganese
Composite
0.03

Copper
Composite
0.01

Lead
Composite
0.01

Molybdenum
Composite
Not detected
Carotene
3
221.4
 59.2
a - Hintz, et al, 1966
b - Percent dry matter
c - Standard error
d - jugm/gm
environmental conditions under which the algae are grown.
The carbohydrate (fiber) portion of the cell is fairly
stable; however, the fatty acid (ether extract) or lipid
components are particularly variable. If algae are grown
in a low nitrate medium, the cell develops fatty acid
storage products at the expense of protein production.
Milner (1948) reported that the protein fractions of
Chlorella pyrenoldosa varied from 7.9 to 46.4 and the lipid
fraction varied from 20.2 to 75.5 percent of the total cell
material, depending on environmental conditions. The pro-
duction of fats might lower the value of algae as a food
supplement, but this should not occur in the algae stripping
process where nitrogen is seldom a limiting factor.
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Foree and McCarty (1968) summarized the results of
various investigators reporting on the composition of fresh-
water algae. Table 21 illustrates the concentration of the
various algal components. The percentage of ash was highest
in the two species of Scenedesmus analyzed and was about 15
percent of the dry weight. Scenedesmus also had a higher
proportion of protein, about 50 percent of the dry weight,
than the other algae cultures listed.
During the feasibility studies at Firebaugh,
relatively little analytical work was performed on the compo-
sition of the algae. Organic nitrogen concentrations ranged
from about 2 to 12 percent of the volatile solids of algae
produced in outdoor growth units, although it was normally
in the range of 7 to 9 percent. Total phosphorus was usually
1 to 3 percent of the volatile solids. The difference between
the algal product collected at Firebaugh and the algae shown
in Table 21 was the high percentage of ash, usually about 30
to 50 percent of the total suspended material in the Firebaugh
product. Analysis of the ash showed the existence of signifi-
cant concentrations of carbonates, iron, phosphorus, chloride,
sulfate, and magnesium. The primary anionic constituent was
carbonate. The ash was present when ponds were operated at
pH values near neutrality but was lowered to about 10 percent
nonvolatile material when in-pond sedimentation was included
in the pond design.
Amino acid profiles were run on two samples of
oven-dried algae by means of automated column chromatographic
analyses. The results are given in Table 22. Both samples
contained all ten of the amino acids essential for animal
growth. The values as determined by analyses are in reason-
ably close agreement with those reported by Fisher and Burlew
(1953) for pilot plant cultures of Chlorella pyrenoldosa.
Samples of the algal product and tile drainage water
were analyzed for the presence of pesticide by gas chromato-
graphy. Neither the algae nor the water contained detectable
amounts of organic phosphorus pesticide, but the tile drain-
age had about 10 parts per trillion (ppt) of dieldrin. The
dried algae product had 75 parts per billion (ppb) of uniden-
tified chlorinated hydrocarbons, probably the decomposition
products of chlorinated hydrocarbon pesticides.
Several feeding experiments have been reported in
which algae have been used in place of the normal protein
supplement. Hintz, et al (i960), fed various percentages of
sewage-grown algae (mTxe^r culture of Scenedesmus and Chlorella)
to swine, cattle and sheep. Results obtained in this study
showed that to ensure consumption the algae had to be pelleted
with other feeds and that ruminants could digest about
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TABLE 21
PROTEIN, CARBOHYDRATE, AND LIPID CONTENTS OF SOME FRESHWATER ALGAE
(From Foree and McCarty, 1968)
Algal	: (% Dry Wt.) : Number of :	Ash-Free Dry Weight) :
Species	: Ash	: Cultures : Protein : Carbohydrate : Lipid :	Reference
Chlorophyta
Chlorella pyrenoidosa
5.60
13
39-6
19.3
4l.l
Spoehr & Milner (1949)
Chlcx*ella pyrenoidosa

17
37.4
34.0
28.6
Ketchum & Redfield (1949)
*Chlorella pyrenoidosa

8
16.1
35.0
48.9
Aach (1952)
*Chlorella vulgaris

5
19.2
53.6
27.2
Fogg & Collyer (1953)
Chlorella vulgaris
12.40
1
48
32
20
Ketchum & Redfield (1949)
Scenedesmus obliquus
15.33
2
51
23
26
Ketchum & Redfield (1949)
Scenedesmus basilensis
14.34
1
53
20
27
Ketchum & Redfield (1949)
Stichococcus bacillaris
9.61
6
4l
38
21
Ketchum & Redfield (1949)
Stichococcus bacillaris
8.87
2
42.5
32.1
25.4
Milner (1953)
Chlamydomona s sp.
4.74
1
36.3
58.2
5.5
Milner (1953)
~Carbohydrate fraction calculated assuming 5$ ash and protein + carbohydrate + fat =
100$ ash-free dry weight.
Values presented are mean values for the number of cultures indicated.

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TABLE 22
AMINO ACID COMPOSITION OF OVEN-DRIED RGP PRODUCT ALGAE
	Sample 1 - 10/15/69	:	Sample 2- 1/3/70
Amino Acid :Percent of Total :mg N/mg Algae:Percent of Total :mg N/mg Algae
: Amino Acid Cone.: x 10"	: Amino Acid Cone.: x 10"^
Lysine .
7.6
3.2
5.7
1.1
Histidine^/
2.2
1.3
1-9
0.5
Argininel/ .
Tryptophani/
7.8
1.1
5.5
0.3
5-1
1.0
1.6
0.1
Aspartic Acid
10.5
2.If
11.9
1.3
Threonine!/
4.9
1.3
5 .4
0.6
Serine
4.7
1.4
6.0
0.8
Glutamic Acid
10.5
2.2
13.1
1.2
Proline
5.7
1.5
5.2
0.7
Glycine
6.6
2.7
7.1
1.3
Alanine
9-5
3.2
8.4
1.3
Valinei/
5.8
1.5
6.1
0.7
Methionine!/
1.8
0.5
1.8
0.2
Isoleucinei'
3.7
0.9
4.2
0.4
Leucinei/ .
8.8
2.1
8.4
0.8
Tyrosine^/ .
Phenylalanine^/
3.5
0.6
3.4
0.2
4.8
1.0
5.6
0.5
Total
99.5
31.6
100.3
13.3
-'Essential amino acid.
20 percent more of the crude protein than could swine. The
study concluded that, although algae were not a high energy
food, its high protein content and available minerals made
It a potentially useful food supplement. Erchul and Isenberg
(1968) evaluated the protein quality of various algal bio-
masses produced at a water reclamation experimental pilot
plant. The products were mainly a mixture of Chlorella and
Scenedesmus fed to rats to compare protein efficiencyratios
(PER} to casein, soybean, and fish meal supplements. The
algal biomass with the highest PER compared favorably with
soybean meal. The variation in PER was apparently due to
differences in digestibility.
Leveille, et a_l (1962), reported on the use of
various algae (Chlorella, a mixture of Chlorella and
Scenedesmus and Sponglococcum) as the sole protein source In
rat and chick feeding studies. All the algae were inferior to
soybean meal as a source of protein. The data showed them to
be deficient in methionine for rats and chicks and the mix-
ture to be deficient in glycine for chicks. Grau and Klein
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unpublished progress report, 1956) fed sewage-grown algae
Scenedesmus-Chlorella mixture) to chicks and found that
growth was generally slower when algae was used in place of
soybean meal. The use of alum-flocculated algae appeared to
be especially detrimental to chick growth when the concentra-
tion of aluminum in the feedstuff exceeded 0.5 percent of the
chickens' total food intake.
Researchers from the North American Aviation
Company (1967) reported on the feeding of mixtures of
Scenedesmus, Closteridlum, and Chlorella (grown on sewage)
to chickens as a substitute for soybean meal. Their results
indicated that the protein value and feeding efficiencies
of algae were comparable to soybean meal. The algae may
actually be preferred by the chicken rancher because the
algal pigment, xanthophyll, gives a deeper golden color to
the skin, meat, and egg yolks. The algae grown at Firebaugh
may be more suitable for poultry production because the salts
included with the algae might provide some essential nutrients
to the fowl. (Most poultry diets contain a small percentage
of certain salts.)
Information in the preceding paragraphs indicates
that algal meal may find a market as a substitute for such
protein supplements as soybean, cottonseed, or fish meal,
especially in poultry production, although more data are
needed. The next step was to determine whether the potential
market could absorb the production of the proposed algal
stripping plant. In Table 23 the estimated maximum and mini-
mum algal production is given for each five-year interval
from 1975 to 2000. The data are based on projected drainage
flows from the entire San Joaquin Valley and estimates of
algal production per unit flow. The basic assumption is that
the total drainage from the Valley will eventually require
treatment. The data indicate that peak annual production may
be between 66 x 103 and 83 x 103 tons of dry algae. Accord-
ing to the California Department of Agriculture (1965)* the
California poultry industry used more than 560 x 103 tons of
protein supplement in 1963> while the total used by all feed-
ing operations was almost one million tons. The prices paid
for these animal feed supplements varv from $8o/ton (cotton-
seed meal) and $l00/ton (soybean meal) to $110/ton (meat and
bone meal). If, as the tests indicate, algae can readily be
substituted for soybean meal, the value of the by-product
will be as shown in Table 23.
Use of Algae as a Soil Conditioner
Another market which can absorb the production of
a full-scale algal stripping plant is the retail market for
soil supplements to lawns, golf greens, etc. Its high
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TABLE 23
ESTIMATED ALGAL PRODUCTION BY AN ALGAL STRIPPING PLANT,
1975-2000
Year :
Thousands
of Algae
of Tons :
per Year :
Approximate Value as
Substitute for Soybean Meal

Maximum :
Minimum :
Maximum :
Minimum
1975
13,300
8,410
$1,330,000
$ 841,000
1980
27,200
18,000
2,720,000
1,800,000
1985
44,300
29,610
4,430,000
2,961,000
1990
62,000
42,400
6,200,000
4,240,000
1995
75,300
54,100
7,530,000
5,410,000
2000
82,610
65,510
8,261,000
6,551,000
nitrogen content and various salts In the product, combined
with its slow rate of decomposition, make algae a desirable
lawn conditioner. The current wholesale price of a soil
conditioner manufactured in San Jose, California, is about
$100/ton. This figure is the cost of 50-pound sacks of the
product intended to retail at $l40/ton, 1970 prices, in
Fresno, California. Railroad freight agents were hesitant
about quoting shipping costs: however, the maximum prices
would be as shown in Table 24 (shipped in 100-pound sacks).
The exact shipping costs, which will be determined when algal
product is given a freight classification, will have to be
deducted from the value of the algae.
Sewage plant operators have tried to develop mar-
kets for their dried activated sludge, which is mainly com-
posed of nonbiodegradable polysaccharide, protein, fat, and
inorganic complexes (Hurvitz, 1957). According to a report
by Foster D. Snell, Inc. (l9o9), the market for this product
has decreased because such material is not competitive with
other nitrogen sources, and many plants are unable to recover
freight costs, let alone process costs. An algal product
with higher nitrogen content (6 to 8 percent) than activated
sewage sludge (l to 2 percent) may find a more receptive
market.
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TABLE Zk
RAIL FREIGHT RATES FOR 100 LB. ALGAE PACKAGES
FROM SAN FRANCISCO, CALIFORNIA
 
Destination : Amount Per 100 Lbs. : Amount Per Ton
Los Angeles
$1.30
$26.00
Portland
1.76
35.20
New York
4.31
86.20
Kansas City
3.23
64.60
Fresno
.82
16.40
Tacoma, Washington
1.98
39.60
Atlanta, Georgia
3.9^
68.80
Miscellaneous Possible Markets for an Algal Product
Some other possible markets exist for algae, but
current technological difficulties preclude their consider-
ation as important market outlets. A brief outline of several
of these uses follows:
1.	Some interest has been expressed In
feeding algae to organisms reared commercially in
aquacultures. These organisms include freshwater
fish and marine bivalves (clams and oysters).
Development of these aquatic farms is now mostly
in the talking and planning stages and could develop
into a substantial market for algae.
2.	Borgmann and Feeney (1948) isolated a
sterol from Scenedesmus obliquus which had charac-
teristics identical to those of chondrillasterol, a
sterol isolated from a sponge. This algal sterol
is one of the few naturally occurring compounds
having the necessary configuration to serve as a
starting point in the manufacture of cortisone, an
important drug. More research will be required to
determine whether this sterol is present in commer-
cially valuable quantities in Scenedesmus quadricauda
used in this project.
3.	Use of algal protein in the adhesive
industry has been suggested as a market for the
product. Samples of Chlorella sent by Fisher and
Little (1953) to a private laboratory to investigate
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the possible uses of the protein fraction in algae
showed that the protein was difficult to separate
and that the molecular weight was too low to serve
as a substitute for casein. Most chemical adhesive
manufacturers are using synthetic compounds for
adhesive production and show little interest in
natural organic sources.
Use of Algae to Produce Methane Gas
Suggestions have been made that an algal product
can be converted to usable energy through methane fermentation,
with subsequent use of the methane gas to provide heat and/or
electric power. In a comprehensive paper on this process,
Oswald and Golueke (i960) concluded that this could be a
practical means of converting solar energy to thermal or elec-
trical energy. The problem with this type of system in tile
drainage is the high concentration of sulfate present. This
amounts to about 3*000 mg/1 in the Alamitos tile system.
Foree and McCarty (1968) studied the anaerobic decomposition
of algae and concluded that, as long as dissolved sulfate is
available, organic matter decomposes through the reduction of
the available sulfate ion. Further stabilization of the
material then occurs by methane fermentation, as long as the
sulfide concentration is not toxic to the bacteria. Little
or no methane was produced in the decomposition of algal
cultures containing about 2,000 mg/1 dissolved sulfate. The
presence of this inhibitory action may preclude the use of
this method of algal disposal in the projected agricultural
wastewater treatment plant, but an investigation would be re-
quired to ascertain the feasibility, methodology and problems
associated with such a system.
-Ill-

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CHAPTER VI - PROCESS EVALUATION
The studies at the IAWTC have shown that the algal
stripping process can be a reliable means of reducing the
nitrogen concentration in agricultural tile drainage. Because
the algal process involves more than one step, and because
each step may have alternatives, this section of the report
will assign tentative estimates of nitrogen removal at each
step, as well as total nitrogen removal by the system. Also,
the section will include some preliminary design criteria
and estimates of treatment costs based on facilities built
and operated according to these criteria.
Removal Efficiencies
As shown in Figure 29, the algal cultures assimi-
lated an average of 80 percent of the total influent nitrogen
(80 percent of 20 mg/l, or about 16 mg/l) from July through
December 1969. This period, which covered the temperature
range noted at the site, was selected because of standardized
conditions in operation of the growth units. This 80 percent
average is somewhat misleading because it includes data from
the start-up period of the run  algae growing at steady-
state conditions assimilated about 90 percent, or 18 mg/l,
of the influent nitrogen with an effluent of 2 mg/l, or less,
total nitrogen. The amount of nitrogen assimilated by the
algae increased steadily as the study progressed, probably
because of more complete understanding of the system and
natural selection of a strain of algae more adapted to the
waste and to the environment of the San Joaquin Valley. The
average amounts of Influent nitrogen assimilated by the
(best) pond during steady-state conditions for all minipond
runs are shown in Figure	During 1969 from 16 to 21 mg/l
of nitrogen were assimilated, as compared to 7 to 10 mg/l in
the earlier runs. Preliminary (1970) operational data obtained
at the time this report was being prepared indicate that the
algal system can remove 27 to 28 mg/l from a 30 mg/l nitrogen
influent.
Based on data from the feasibility phase of the
project, minimum nitrogen assimilation (20 mgN/l influent)
will be 16 mg/l and maximum assimilation about 18 mg/l. If
95 percent of the algae can be separated from the liquid
phase (a level consistently achieved by the flocculation-
sedimentatlon unit tested at the IAWTC), then the total nitro-
gen remaining in the effluent would be on the order of:
-113-

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Nrp = Nj) + where NT = total effluent N; Np = dissolved
effluent N; and NA = nitrogen in effluent algae or
{% algae remaining)(original algal conc.)(# N in
algae)
Maximum Nrj. = 4 mg/l + (0.05 remaining)(250 mg/l)
(0.08 N in algae)
= 5 mg/l N,
Minimum = 2 mg/l + (0.05 remaining)(250 mg/l)(o.08 N
in algae)
= 3 mg/l N
"" C7T
r
nser
15T~
(3)	(9A)
(IaI
Tib) "i?\"
... ! <
4B	4B
	 1968
	 1969
NOTE: NUMBERS IN PARENTHESIS ARE RUN NUMBERS
1	1	1	1	1	,	1			1	1	
FEB MAR APR MAY JUN JUL AUG SEP OCT NOV DEC JAN
MONTH '
FIGURE 54-NITROGEN ASSIMILATION IN VARIOUS MINIPOND RUNS
-114-

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These figures were based on 20 mg/l total N In the influent.
Thus, overall nitrogen removal efficiencies will range from
75 to 85 percent. Depending on water temperatures, growth
unit detention times of 5 to 16 days will be required to
achieve these removal rates.
Process Configuration
From the data contained in this report, a tentative
process configuration can be drawn. The configuration is
preliminary in that data from the operational phase of the
study were not considered. Figure 55 illustrates the flow
diagram of an algal stripping plant. Shallow growth units
are followed by a sedimentation process which removes 95
percent of the suspended biomass. The effluent from the
sedimentation tanks leaves the plant site and the algal slurry
goes to a vacuum filter for dewatering to about 20 percent
solids. The effluent from the dewatering device is recycled
to the head of the sedimentation area while the sludge is
dried to 85 to 90 percent solids by air or flash driers. The
dried product is then re-
moved from the site to be
sold as a marketable by-
product.
Figure 56 Illus-
trates a schematic design
of an algal growth unit.
The flow enters and leaves
the unit by gravity
through concrete pipes.
Iron, phosphorus, and
CO2 are added to individ-
ual growth units at the mixing pumps. Outdoor propeller-
type pumps perform four hours mixing per day. To reduce the
required number of mixing pumps, it is proposed that the
ponds be constructed in units of 12 with a central mixing
station capable of mixing two ponds at a time for four hours.
This type of design is shown in Figure 57 A series of auto-
mated valves would rotate mixing to the desired ponds. An
in-line sedimentation area included in the design of the
growth unit will remove much of the heavier suspended solids.
Cost Estimates
The feasibility studies conducted at the IAWTC were
not designed to provide definitive cost data for an algal
stripping system; however, some preliminary process cost esti-
mates can be made. These estimates will undoubtedly be revised
as more data become available from 1970 operational studies.
F#,P,

PLANT	
INFLUENT
SEDIMENTATION
GROWTH
PLANT
EFFLUENT
FIGURE 55-FLOW DIAGRAM OF ALGAL STRIPPING PLANT
-115-

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This is especially true in
the case of pond mixing.
Results demonstrated that in-
organic carbon addition may
be more important than mixing.
Predicted seasonal
variation in flows and nitro-
gen concentrations of tile
drainage from the San Joaquin
Valley (see Figure 58) and
climatic factors such as
temperature and sunlight indi-
cated that April was the criti-
cal month for sizing the growth
units. During this month, flows
will be high, nitrogen con-
centration should be about 30
mg/l, and sunlight and temper-
ature conditions will not be
optimum.
Several factors were
used in obtaining these esti-
mates:
1.	All costs were
based*on January 1970 dollars.
2.	Debt service
was calculated for 50 years at
5 percent.
3.	Costs per million
gallons treated were calculated
by dividing the total annual
cost (debt service plus oper-
ations and maintenance costs) fo
the estimated annual flow of the
FLOW DIRECTING-
VANES
IN-LINE
SEDIMENTA-
TION
TANK
o )u-
200'
INFLUENT
STRUCTURE
EFFLUENT
STRUCTURE-
PLAN VIEW
FIGURE 56-ALGAE GROWTH POND
> a full-capacity plant by
San Luis Drain.
4.	An engineering and contingency factor of 57
percent was assumed for capital cost items whose design and
operation was based on experimental data obtained at the
treatment center. A lower engineering and contingency
factor was used for items common to waste treatment facili-
ties and for those items which can be purchased from the
manufacturer. These engineering and contingency factors are
shown in Table 27.
5.	The land required for a full-capacity plant is
to be purchased at the beginning of the project at $500 per
acre.
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IN-UNE SEDIMENTATION TANK
3
5!
"8
100 TYPICAL
V;
POND NUMBER
MIXING PUMPS
FIGURE 57- ONE OF THREE 12-POND GROUPS CONSTRUCTED PER PHASE
-117-

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20i
r40
I
H"
H
00
t

o
<
z>
z
z
<
_J
2
o
-
li.
o
z
llj
o
cr
UJ
L
V*^-NITROGEN
FLOW
20 w
FEB MAR APR MAY JUN JUL AUG SEP OCT NOV DEC
JAN
FIGURE 58-PREDICTED SEASONAL VARIATION OF TILE DRAINAGE FLOW a NITROGEN
CONCENTRATIONS FROM SAN JOAQUIN VALLEY, CALIFORNIA

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6.	The laboratory and office buildings and main-
tenance and storage areas are basically the same as those
at the San Jose, California, Sewage Treatment Plant. Capital
costs for these facilities are based on information published
by Guthrie (1969).
7.	Electric power co3ts were calculated on the
basis of $.01/kw-hr.
8.	General plant operation and maintenance (o&M)
costs were based on curves published by Smith (19&7 ) for
trickling filter O&M. It was assumed that the general plant
0&J4 costs included replacement of power costs for all plant
operations except for the items indicated in Table 26.
9 The cost of reaeration was taken from figures
published by Smith (1967).
The individual ponds are designed according to the
criteria presented in Table 25, with a mean channel length
of approximately 6,500 feet. The influent and effluent struc-
tures, the in-line sedimentation tank, and flow-directing
vanes are all made of concrete, in addition, wherever a con-
crete structure and the growth pond proper meet, a 20-foot
concrete apron is built. Although an in-line sedimentation
tank was not built into the rapid growth pond operated at the
IAWTC, evaluation of a sedimentation unit used in conjunction
with the growth pond indicated that a device of this type
would be beneficial to algal growth. The in-line sedimenta-
tion system included in this proposed pond design is routinely
used in operations requiring sedimentation facilities but may
not be the best solution of the problem of removing excess
suspended material from the growth unit. With the exception
TABLE 25
ALGAE STRIPPING DESIGN CRITERIA
April Growth Pond Hydraulic Detention Time	8 days
Growth Pond Depth	1 foot
Growth Pond Width	200 feet
Mixing Velocity	1 fps
Duration of Mixing	4 hrs/day
In-line Sedimentation Tank
Surface Loading	5*000 gpd/ft^
Detention Time	15 minutes
Growth Chemical
Pe	2 mg/l
P	2 mg/l
CO2	800 lbs/million gal.
-119-

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of the structures and aprons, the pond bottoms are assumed
to be 22 percent soil cement and 78 percent compacted native
soil by volume. The ratio is intended to prevent seepage into
or out of the ponds. The levee which divides the pond is 2
feet high and has a crest width of 8 feet wide and 1.5-to-l
side slopes. It is constructed from the soil excavated from
the influent and effluent structures and the in-line sedi-
mentation tank. The flow-directing vanes are 2 feet high and
8 inches thick and built on 50-, 100-, and 150-foot radii.
Ponds are separated from each other on the long sides by a 3-
foot high levee with a 20-foot crest and 1.5-to-l side slopes.
The surface loading on the flocculation-sedimentation
tank is considered to be 900 gpd/ft^, with 10 mg/l ferric
sulfate and 0.5 mg/l cationic polyelectrolytes (Calgon - Cat-
Ploc) as the chemical additions. Loading on the vacuum filter
is 0.2 gpm/ft2, with capital and 08M cost estimates supplied by
the Eimco Corporation. One-half of the sludge from the vacuum
filter goes to 1,000 pounds H20/hr driers, and the remaining
half is air-dried. Cost estimates for the driers were provided
by the De Laval Separator Company.
Table 26 lists the capital costs of the various com-
ponents of the algae stripping system and Table 27 illustrates
the total costs of these components for treating a million
gallons of tile drainage. The estimated net cost of treat-
ment Is $135 per million gallons, which includes the
recovery of some money through the sale of an algal product.
The product recovery figure was based on producing 1400 pounds
of algae per million gallons of water and selling the product
at $60 per ton.
The costs shown in Tables 26 and 27 were obtained by
assuming that the plant would be built to treat the estimated
maximum annual quantity of about 50,000 million gallons for the
San Luis Drain. Because the estimated ultimate annual flow
of the facility will not be attained until abotit 30 years from
the time the initial drainage reaches the treatment plant,
another approach would be to construct the plant in stages
paralleling the buildup of drainage flows. For a treatment
plant constructed in phases of equal size (each additional
phase to be constructed during the year when drain flow reaches
90 percent of existing plant capacity), the average net cost
for treatment over the first 50 years of operation would
probably be higher than the estimate shown above. This in-
creased price is because part of the plant would be idle at
all times.
These preliminary cost estimates were obtained to
determine which aspects of the process were the most costly.
With this information, the operational studies of 1970 were
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TABLE 26
TREATMENT COSTS FOR ALGAE STRIPPING
Dollars Per Percent
Million	of
Item		Gallons Total Cost
Capital
Production
Growth Pond
55
30
In-line Sedimentation Tanks
5
3
Chemical Peed Pumps
1
1
Mixing System
39
22
Harvesting


Separator
1
1
Dewatering
1
1
Drying
6
3
Other


Land, Building, Miscellaneous
6
3
Annual O&M Costs


General
21
12
Drying, Dewatering & Growth Chemicals
27
15
Mixing Power
16

TOTAL
$177/mg
Minus By-product Income
-42

Net Cost of Treatment	$135/Milllon gallons
directed toward reducing the cost of the more expensive items
(mixing, land preparation, etc.)* while maintaining the maximum
level of nitrogen removal.
The cost estimate presented in this section was
based on high-rate algal growth units. The other algal system
studied at the site  the "symbiotic", soil-lined ponds 
has not had a co3t estimate, primarily because of the unknown
nature of the systems involved. Data obtained during the 1970
operational studies will provide some information on the ques-
tions of reliability and the mechanisms involved. With this
data cost estimates can be prepared. Because the system is
simple and because it requires no mixing and little chemical
addition for growth, costs for this process will be consider-
ably lower than for the high-rate algal growth system.
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TABLE 27
CAPITAL COST FOR ALGAE STRIPPING
Number
Item
Capital Expenditure - 1970 Dollars
1
2
3
4
5
6
7
8
9
10
Production
Growth Ponds
In-line Separation Tanks
Chemical Feed Pumps
Mixing System
Harvesting
Separator
Dewatering
Drying
Other
Land Acquisition
Buildings
Miscellaneous
$ 50,900,000
5,000,000
125,000
37,500,000
250,000
1,250,000
6,125,000
5,000,000
750,000
157,000
Total
$107,057,000
Notes: (l) Items Nbs. 2,3,5,6,7 and 9 include a 25 percent engineering and
contingency factor.
(2)	Items Nos. 1, 4 and 10 include a 57 percent engineering and
contingency factor furnished by the U. S. Bureau of Reclamation,
in its "reconnaissance" estimate.
(3)	No engineering and contingency factor was applied to land cost.

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CHAPTER VII - AREAS FOR FUTURE INVESTIGATION
Many of the questions concerning + chnical feasi-
bility have been answered by the study at 17 L'ebaugh and other
answers will result from current operational studies. There
are, however, areas of interest which cannot be fully investi-
gated during the current project and which warrant further
study. The proposed studies will be more meaningful when a
combined drainage facility provides larger quantities of water.
Among the areas of interest are:
1.	A redesigned algal system which may reduce the
land area and modify the type of growth units necessary for
nitrogen removal.
The basic idea for this type of system was derived
from personal communication with Dr. William J. Oswald,
Professor of Environmental Health Sciences and Sanitary
Engineering at the University of California, Berkeley. The
type of unit suggested by Dr. Oswald would consist of a number
of pond in series, each containing different depths of water.
The firat pond would be relatively deep, with light as the
factor limiting algal growth. This pond would only receive
phosphorous addition and would not require mixing. The next
pond would be shallower but light would still be the only
limiting factor. The last pond in the series would be similar
to the rapid growth pond used in the present study (shallow,
with mixing, etc.), and nitrogen would be the constituent
limiting growth. Nitrogen assimilation in the earlier ponds
would permit the use of short detention times in the final
unit. Theoretical considerations indicate that a system of
this type could reduce the land area required in a conventional
system as much as 50 percent.
2.	Use of the drain as a treatment system has been
described by Goldman, e_t al (1969), in which the drainage
canals and holding reservoTr are part of the process.
Because of nutrients contained in the water, the
optical clarity of the drainage and the use of relatively
shallow, lined canals, some natural algal growth (either
planktonic or sessile), estimated at 15 to 60 mg/l, will occur
to remove some nitrogen. If this growth is encouraged by
nutrient additions and/or modification to the canal to in-
crease turbulence, as much as 100 mg/l of algae could possibly
be grown in the canal prior to Kesterson Reservoir (a holding
reservoir). This could remove about 8 to 9 mg/l nitrogen.
Then if Kesterson were redesigned to include both holding
capability and improvements for increasing algal growth, it
could conceivably remove the remaining nitrogen. A separation
-123-

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plant would then be placed at the outlet of the reservoir.
If these facilities are included in the drainage system,
considerable savings in land costs may be realized.
3. Use of the algae growing capability of the drain
and of Kesterson Reservoir with the anaerobic denitrification
filter process as a final polishing step.
In the anaerobic system, an organic carbon source is
added to the drainage and the water passed through a column
containing some type of aggregate. Anaerobic bacteria
attached to the media reduce the nitrate to nitrogen gas,
which is evolved to the atmosphere. Studies at Firebaugh
have demonstrated that algae entering the columns can increase
dissolved organic and ammonia nitrogen, presumably by bacte-
rial decomposition of algal protein. Since algae undoubtedly
will grow in the drain, some provision for their removal will
have to be incorporated in the design of an anaerobic system.
If algal growth is encouraged, the dual system may reduce
costs and provide a more reliable system for nitrogen removal.
An unpublished communication from Dr. Oswald contains refer-
ence to the following possible benefits from the dual system:
a.	Back-up coverage is provided in case of
breakdown of one of the systems.
b.	The algal system can remove ammonia
nitrogen produced by decomposition of bacterial
dells in the filter.
c.	Recreational use may be provided by the
ponds.
-12k-

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ACKNOWLEDGMENTS
The following list includes the personnel actively involved in
the algae stripping project at the IAWTC.
Algae Stripping Fro.ject Under the Direction of
Louis A. Beck .... Engineer, California Department of Water Resources
Donald Swain 	 Engineer, U. S. Bureau of Reclamation
Percy St. Amant 	 Engineer, Environmental Protection Agency
Conducted by
Randall L. Brown . . Biologist, California Department of Water Resources
Bruce A. Butterfield . Engineer, California Department of Water Resources
Joel Goldman 	 Engineer, California Department of Water Resources
James F. Arthur 	 . Biologist, Environmental Protection Agency
James R. Jones			Engineer, U. S. Bureau of Reclamation
Assisted by
Robert G. Seals 	 Chemist, Environmental Protection Agency
William R. Lewis . . . Chemist, California Department of Water Resources
Dennis L. Salisbury 	 ..... Engineering Technician,
California Department of Water Resources
William L. Baxter . . . Laborer, California Department of Water Resources
Clara P. Hatcher 	 Laboratory Aid,
California Department of Water Resources
Elizabeth J. Boone 	 . Laboratory Aid,
California Department of Water Resources
Linda S. Harrington 	 Laboratory Aid,
California Department of Water Resources
Consultants to the Pro.iect
Dr. William J. Oswald	University of California, Berkeley
Dr. Clarence G. Golueke 	 University of California, Berkeley
Dr. Perry L. McCaxty 	 Stanford University, Palo Alto
Report Prepared by
Randall L. Brown 	 California Department of Water Resources
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42.	North American Aviation, Inc. 1967. A study of the use of
biomass systems in water renovation. Pinal Report.
43.	Osterlind, S. 1950. Inorganic carbon sources of green
algae. II - Carbonic anhydrase in Scenedesaus quadrlcauda
and Chlorella pyrenoidosa. Physiologic Plantarum. 3:^30-434.
44.	Oswald, W. J. 1963. Light conversion efficiency of algae
grown in sewage. Trans. Amer. Soc. Civil Engrs. Vol. 128
part III: 47-83.
45.	Oswald, W. J., D. G. Crosby and C. G. Golueke. 1964.
Removal of pesticides and algal growth potential from
San Joaquin Valley drainage water (A feasibility study).
Unpublished report submitted to the Calif. Dept. of
Water Resources.
46.	Oswald, W. J. and C. G. Golueke. i960. Biological trans-
formation of solar energy in Advances in Applied Microbiology.
Academic Press Inc., New YorkT Vol. II: 223-2bl.
47.	Oswald, W. J. and C. G. Golueke. 1968. Harvesting and
processing of waste-grown microalgae. In Algae, Man and the
Environment. D. P. Jackson (ed.). Proceedings of a Symposim
held June I8-30 1967. PP. 371-389.
48.	Oswald, W. J. and H. B. Gotaas. 1955. Photosynthesis in
sewage treatment. Proceedings - Separate No. 636. Amer.
Soc. Civil Engrs.
49.	Retovsky in Malek, I. and Z.Pencl. 1966. Theoretical and
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Academic Press. New York.
50.	Sawyer, C. N. 1952. Some new aspects of phosphates in
relation to lake fertilization. Sewage and Industrial Wastes.
Vol. 24, No. 6:768.
-129-

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51.	Schuler, J. F., V. M. Diller, and H. J. Kerslen. 1953*
Preferential assimilation of ammonium ion by Chlorella
vulgaris. Plant Physiol., 28:299-303.
52.	Shelef, G., W. J. Oswald, and C. G. Golueke. Light inten-
sity and nitrogen concentration as growth limiting factors.
U. of Calif. Sanit. Engr. Res. Lab. Report No. 6o-4.
53.	Smith, R. 1967. A compilation of cost information for
conventional and advanced wastewater treatment plants and
processes, IJSDI, FWPCA. Cincinnati, Ohio.
54.	Spoehr, H. A. and H. W. Milner. 19^9 The chemical com-
position of Chlorella; Effect of environmental conditions.
Plant Physiol., 24:120-149.
55.	Stumm, W. and J. J. Morgan. 1962. Chemical aspects of
coagulation. J. Amer. Water Work Assoc., 54:971.
56.	Tenney, M. W., W. P. Echelberger, Jr., R. G. Schuessler,
and J. L. Paroni. 19^9. Algal flocculation with synthetic
organic polyelectrolytes. J. of Applied Microbiol.
Vol. 18, No. 6:965-971.
57.	Williford, J. W. and D. R. Cardon. 1970. Potential changes
in the nitrogen concentration of drainage water during
transport. Report No. 4. Agricultural Wastewater Study
Group.
58.	Witt, V. M. and J. A. Borchardt. i960. The removal of
nitrogen and phosphorus from sewage effluents through the
use of algal culture (Scenedesmus and Chlorella in mixed
or unialgal cultures). J. Biochem. Microbiol. Technol.
Engr. 2:187-203.
59 Zabat, M., W. Oswald, C. Golueke, and H. Gee. 1970. Kinetics
of algal systems in waste treatment, phosphorus as a growth
limiting factor. SERL Publication, University of California,
Berkeley.
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PUBLICATIONS
SAN JOAQUIN PROJECT, FIREBAUGH, CALIFORNIA
1968
"Is Treatment of Agricultural Waste Water Possible?"
Louis A. Beck and Percy P. St. Amant, Jr. Presented
at Fourth International Water Quality Symposium,
San Francisco, California, August 14, 1968; published
in the proceedings of the meeting.
I22
"Biological Denitrification of Wastewaters by Addition of
Organic Materials"
Perry L. McCarty, Louis A. Beck, and Percy P.
St. Amant, Jr. Presented at the 24th Annual Purdue
Industrial Waste Conference, Purdue University,
Lafayette, Indiana. May 6, 1969*
"Comparison of Nitrate Removal Methods"
Louis A. Beck, Percy P. St. Amant, Jr., and Thomas A.
Tamblyn. Presented at Water Pollution Control Federa-
tion Meeting, Dallas, Texas. October 9, 1969.
"Effect of Surface/Volume Relationship, C02 Addition, Aera-
tion, and Mixing on Nitrate Utilization by Scenedesmus
Cultures in Subsurface Agricultural Waste Waters
Randall L. Brown and James F. Arthur. Proceedings
of the Eutrophication-Biostimulation Assessment
Workshop, Berkeley, California. June 19-21, 1969*
"Nitrate Removal Studies at the Interagency Agricultural
Waste Water Treatment Center, Firebaugh, California"
Percy P. St. Amant, Jr., and Louis A. Beck. Presented
at 1969 Conference, California Water Pollution Control
Association, Anaheim, California, and published in the
proceedings of the meeting. May 9, 1969.
"Research on Methods of Removing Excess Plant Nutrients from
Water"
Percy P. St. Amant, Jr., and Louis A. Beck.
Presented at 158th National Meeting and Chemical
Exposition, American Chemical Society, New York,
New York. September 8,-1969.
"The Anaerobic Filter for the Denitrification of Agricultural
Subsurface Drainage"
T. A. Tamblyn and B. R. Sword. Presented at the 24th
Purdue Industrial Waste Conference, Lafayette, Indiana.
May 5-8, 1969.
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PUBLICATIONS (Continued)
SAN JOAQUIN PROJECT, FIREBAUGH, CALIFORNIA
1969
"Nutrients in Agricultural Tile Drainage"
W. H. Pierce, L. A. Beck and L. R. Glandon. Presented
at the 1969 Winter Meeting of the American Society of
Agricultural Engineers, Chicago, Illinois.
December 9-12 1969 
"Treatment of High Nitrate Waters"
Percy P. St. Amant, Jr., and Perry L. McCarty.
Presented at Annual Conference, American Water Works
Association, San Diego, California. May 21, 1969.
American Water Works AssociationJournal. Vol. 61.
No. 12. December 1969. pp. 659~bt>2.
The following papers were presented at the National Fall
Meeting of the American Geophysical Union, Hydrology Section,
San Francisco, California. December 15-lo, 1969. They
are published in Collected Papers Regarding Nitrates in
Agricultural Waste Water. USDI, FWQA, #13530 ELY
December 19^9
"The Effects of Nitrogen Removal on the Algal Growth
Potential of San Joaquin Valley Agricultural Tile Drainage
Effluents"
Randall L. Brown, Richard C. Bain, Jr. and Milton G.
Tunzi.
"Harvesting of Algae Grown in Agricultural Wastewaters"
Bruce A. Butterfield and James R. Jones.
"Monitoring Nutrients and Pesticides in Subsurface Agricultural
Drainage"
Lawrence R. Glandon, Jr., and Louis A. Beck.
"Combined Nutrient Removal and Transport System for Tile
Drainage from the San Joaquin Valley"
Joel C. Goldman, James F. Arthur, William J. Oswald,
and Louis A. Beck.
"Desalination of Irrigation Return Waters"
Bryan R. Sword.
"Bacterial Denitrification of Agricultural Tile Drainage"
Thomas A. Tamblyn, Perry L. McCarty and Percy P.
St. Amant.
"Algal Nutrient Responses in Agricultural Wastewater"
James F. Arthur, Randall L. Brown, Bruce A. Butterfield,
Joel Co Goldman.
-132-

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I Accession Number
Subject Fiold& Group
05 D
SELECTED WATER RESOURCES ABSTRACTS
INPUT TRANSACTION FORM
r I Organization
_3 [ Department of Water Resources
San Joaquin District
6 |T,"#
	 REMOVAL
OF NITRATE BY
AN
ALGAL SYSTEM
| Q | Authors)
-' Brown,
Randall L
16
Project Designation
13030 ELY


21
Note


Available from Department of Water Resources
Post Office Box 2385
Fresno, California 93723
HJ
221C"a"Agricultu,ral Wastewater Studies
Report No. 13030 ELY V71 - 7
^	Pages 132. Figures 58. Tables 27, References 59	
22 I Descriptora (Starred First)
-' *Agricultural Wastes, *Water Pollution Control, Biological Treat-
ment, Nitrates, Treatment Facilities
25 j ldentifiera (Starred First)	~
*Algae Stripping, Scenedesmus, Algal Growth and Harvesting
3ZJ
I Abstract
An algal system consisting of alga* growth, harveatlng and disposal was evaluated as a possible means
Of removing nitrata-nitrogen from subsurface agricultural drainage in the San Joaquin Valley of California,
the study of this assimilatory nitrogen reaoval process was Initiated to determine optimum conditions for
growth of the algal blomass, seasonal variations in assimilation rates, and methods of harvesting and
disposal of the algal product, a aecondary objective of the study was to obtain preliminary ooat esti-
mates and proceaa design.
Ihe growth studies showed that about 75 to 90 percent of the 20 mg/l influent nitrogen was assimilated
*>7 shallow (12-lneh culture depth) algal cultures receiving 2 to 3 ag/1 additional iron and phosphorus
*nd  mixture of 5 percent C02. Theoretical hydraulic detention times required for theae assimilation
frates varied from 5 to 1< days, depending on the time of the year. The total nitrogen removal by the
*lgal system, assuming 95 percent removal of the algal eells, ranged from 70 to 85 percent of the influent
nitrogen.
*he most economical and effective algal harvesting system tested was flocoulatlon and sedimentation
followed by filtration of the aedlment. the algal oake from the vacuum filter, containing about 20 percent
"olidB, waa then air- or flaah-dried to about 90 percent aollda. The market value for thla product aa a
Protein supplement was estimated to be about $80 to $100 per ton.
Abstractor Brown
'"""'""'""Department of Water Resources
WR: 102 (REV. JUUY !>
WR SIC
WATER RESOURCES SCIENTIFIC INFORMATION CENTER
U.S. DEPARTMENT OF THE INTERIOR
WASHINGTON, D. C 20240
* OJ 19fl9-359399

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