6. AOACSPIFANMethods-2018Awards

264

only. When evaluating after an analysis run, defer to 3.c), below. When the system is equilibrated, order the sequence with the four standard levels (high to low recommended), blanks, samples, and then bracketing standard injections. Repeat calibration after approximately 20 sample injections. g) Following the sample analysis, rinse the analytical column with a solution of approximately 25% methanol in water. For longer storage, the column is best left in 60% to 80% organic, preferably acetonitrile. h) Resolution: In the Laboratory Control sample, the resolution of the Tryptophan peak from the small peak following shall be > 1.25. i) USP Tailing: The USP tailing factor may be calculated within the chromatography software and should be < 1.59 for Tryptophan and < 1.26 for 5-methyl-tryptophan. Excessive tailing indicates the need for a new column to restore expected performance. j) Residual Error of Linear Regression: The residual error for any one point of the four point linear regression is not to exceed 5%. In the event of excessive error, repeat the analysis with fresh standard solutions

G. Calculations

1. Manual Calculations a) Calculate concentrations of the tryptophan standards (mg/L) using the following equations: (1) Tryptophan Stock Standard Solutions: TRP (mg/L) = Weight TRP (g) x 1000 mg Volume (L) g Example:

Tryptophan Weight = 0.0100 grams. Dilution = 100 mL volumetric flask.

0.0100 g x 1000 mg =100 mg 0.1 L g L Tryptophan Concentration (stock) = 100 mg/L

(2) Diluted Tryptophan Standard Solutions:

TRP (mg/L) = Stock Conc. (mg/L) x Dilution

Where: Dilution = Volume Stock (mL) Final Volume (mL)

Example:

Conc. of Stock: 100 mg/L (tryptophan) Dilutions: made in 50 mL volumetric flasks.