Identification—
A:
Determine the absorbances of Assay preparation B at 455 nm and at 483 nm: the ratio of the absorbance at 455 nm to that at 483 nm is between 1.14 and 1.18.
B:
Determine the absorbance of Assay preparation B at 455 nm and that of Assay preparation A at 340 nm: the ratio of the absorbance at 455 nm to that at 340 nm is not less than 1.5.
Assay—
[NOTE—Perform this procedure in subdued light, using low-actinic glassware.
]
Assay preparation A—
Transfer about 50 mg of Beta Carotene, accurately weighed, to a 100-mL volumetric flask, dissolve in 10 mL of acid-free chloroform, dilute with cyclohexane to volume, and mix. Pipet 5 mL of this solution into a 100-mL volumetric flask, dilute with cyclohexane to volume, and mix.
Assay preparation B—
Pipet 5 mL of Assay preparation A into a 50-mL volumetric flask, dilute with cyclohexane to volume, and mix.
Procedure—
Determine the absorbance of
Assay preparation B, at the wavelength of maximum absorbance at about 455 nm, with a suitable spectrophotometer, using cyclohexane as the blank. Calculate the quantity, in mg, of C
40H
56 in the portion of Beta Carotene taken by the formula:
20,000 A X /250,
in which
A X is the absorbance of
Assay preparation B; and 250 is the absorptivity of pure beta carotene.
;)
,
731
—
Dry it in vacuum over phosphorus pentoxide at 40
for 4 hours: it loses not more than 0.2% of its weight.