美国药典(USP)附录-英文.docVIP

171 VITAMIN B12 ACTIVITY ASSAY USP Reference Standards 11— USP Cyanocobalamin RS. Assay Preparation— Place a suitable quantity of the material to be assayed, previously reduced to a fine powder if necessary and accurately measured or weighed, in an appropriate vessel containing, for each g or mL of material taken, 25 mL of an aqueous extracting solution prepared just prior to use to contain, in each 100 mL, 1.29 g of disodium phosphate, 1.1 g of anhydrous citric acid, and 1.0 g of sodium metabisulfite. Autoclave the mixture at 121 for 10 minutes. Allow any undissolved particles of the extract to settle, and filter or centrifuge, if necessary. Dilute an aliquot of the clear solution with water so that the final test solution contains vitamin B12 activity approximately equivalent to that of the Standard Cyanocobalamin Solution which is added to the assay tubes. Standard Cyanocobalamin Stock Solution— To a suitable quantity of USP Cyanocobalamin RS, accurately weighed, add sufficient 25 percent alcohol to make a solution having a known concentration of 1.0 μg of cyanocobalamin per mL. Store in a refrigerator. Standard Cyanocobalamin Solution— Dilute a suitable volume of Standard Cyanocobalamin Stock Solution with water to a measured volume such that after the incubation period as described for Procedure, the difference in transmittance between the inoculated blank and the 5.0-mL level of the Standard Cyanocobalamin Solution is not less than that which corresponds to a difference of 1.25 mg in dried cell weight. This concentration usually falls between 0.01 ng and 0.04 ng per mL of Standard Cyanocobalamin Solution. Prepare a fresh standard solution for each assay. Basal Medium Stock Solution— Prepare the medium according to the following formula and directions. A dehydrated mixture containing the same ingredients may be used provided that, when constituted as directed in the labeling, it yields a medium comparable to that obtained from the formula given herein