UNITED STATES ENVIRONMENTAL PROTECTION AGENCY
NATIONAL EXPOSURE RESEARCH LABORATORY
CINCINNATI, OH 45268
Method 445.0
In Vitro Determination of Chlorophyll a and Pheophytin a
in Marine and Freshwater Algae by Fluorescence	office of
RESEARCH AND DEVELOPMENT
Revision 1.2
ERRATA SHEET
Sectionl.4 - References numbered 5-8 should be numbered 6-8.
Section 12.2 - Equation for calculating the "corrected" concentration of chlorophyll a in the whole water
sample is as follows:
Cs c = Ci. „ X extract volume (L) X DF
Sample volume (L)
Where, Cs c = corrected chlorophyll a concentration (|ag/L) in the whole water sample
extract volume = volume (L) of extract prepared before dilution
Section 13.4.1 - The following has been added to the second paragraph.
The reported p-EDLs reflect between-lab variability and extraction variability. There was,
however, a major flaw in the study design. Even though the concentrations used could be easily determined
by fluorometry after appropriate dilution, we did not specify to the labs the dilution factor to be used.
Because of that, each lab diluted at their discretion. Dilution factors ranged from 10-2000. The
"observed" concentration by each fluorometer was not the reported concentration used in the multi-lab
statistical analysis. Since p-EDLs are based on an estimate of variance (standard deviation) of the reported
concentration in the extract, the p-EDL for fluorometry is not reflective of the concentration actually
observed by the instrument. Since all the participants used different dilution factors there was no way to
correct the determined p-EDLs for the fluorometric techniques. It is safe to say that the statistically
determined p-EDLs are at least 1000 times too high. Still, fluorometry yielded the lowest p-EDLs. The p-
EDLs for the other methods are valid.
Table 4 - The following has been added to footnote 5.
This is due to a flaw in the study design and not due to any inherent limitations of fluorometry.
Please see Section 13.4.1 for a discussion of the determination of p-EDLs. Single-lab EDLs may be 1000
times lower than the p-EDLs reported here.
Tables 4-9 - The following footnote has been added.
Reported concentrations (ppm) are for the 10 mL extraction volume and not the concentrations in
the whole water sample. Using the notation of Section 12 of the method, this would be CE c.

Table 9 - Eighth column, last value should be 0.675

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