Identification—
To about 0.5 g of it in a beaker add 5 mL of water and 10 mL of a mixture of water, 0.2
M potassium chloride, and 0.2 N hydrochloric acid (135:50:13), and mix. Add 5.0 mL of 0.5
M sodium nitrite and 5.0 mL of 0.02
M sodium tungstate, and mix. Heat at 55
to 60
for 15 minutes, allow to cool, and filter. To the filtrate add 10 mL of 1 N sodium hydroxide: a bright yellow color is produced (
presence of capsaicin).
Assay—
Mobile phase—
Prepare a mixture of methanol and 2% acetic acid (56:44), filter through a 0.5-µm or finer porosity filter, and degas. Make adjustments if necessary (see
System Suitability under
Chromatography 
621
).
Standard preparation—
Prepare a solution of
USP Capsaicin RS in methanol having a known concentration of about 0.5 mg per mL. Filter a portion of this solution through a 0.2-µm porosity filter, and use the filtrate as the
Standard preparation.
Assay preparation—
Transfer about 1000 mg of Capsicum Oleoresin, accurately weighed, to a 100-mL volumetric flask, dissolve in and dilute with methanol to volume, and mix. Filter a portion of this solution through a 0.2-µm porosity filter, and use the filtrate as the Assay preparation.
Chromatographic system
(see
Chromatography 621)—The liquid chromatograph is equipped with a 280-nm detector and a 3.9-mm × 30-cm column that contains packing L1. The flow rate is about 2 mL per minute. Chromatograph the
Standard preparation, and record the peak responses as directed for
Procedure: the tailing factor is not more than 2.0; and the relative standard deviation for replicate injections is not more than 2.0%. Chromatograph the
Assay preparation, and record the peak responses as directed for
Procedure: the relative retention times are about 0.9 for nordihydrocapsaicin, 1.0 for capsaicin, and 1.6 for dihydrocapsaicin; and the resolution,
R, between the nordihydrocapsaicin peak and the capsaicin peak is not less than 1.2.
Procedure—
[NOTE—Use peak areas where peak responses are indicated.
] Separately inject equal volumes (about 10 µL) of the
Standard preparation and the
Assay preparation into the chromatograph, record the chromatograms, and measure the responses for the three major peaks. Calculate the percentage of total capsaicins in the portion of Capsicum Oleoresin taken by the formula:
(CP / W)(rU / rS),
in which
C is the concentration, in mg per mL, of
USP Capsaicin RS in the
Standard preparation;
P is the designated purity, in percentage, of
USP Capsaicin RS;
W is the weight, in mg, of Capsicum Oleoresin taken to prepare the
Assay preparation;
rU is the sum of the peak responses for nordihydrocapsaicin, capsaicin, and dihydrocapsaicin obtained from the
Assay preparation; and
rS is the peak response obtained from the
Standard preparation.
