19.5.04
Preparation of Test Solution
AOAC Official Method 986.37
Chlorpropamide in Drug Tablets
Liquid Chromatographic Method
First Action 1986
Final Action 1992
A. Principle
Chlorpropamide is dissolved in mobile phase and determined by
liquid chromatography with UV detection at 240 nm.
B. Apparatus
(a) Liquid chromatograph.—With sampling valve introducing
20 µL in jec tions, UV de tec tor op er at ing at 240 nm, and
recorder/integrator.
(b) Column.—Reverse phase octadecylsilane (ODS), 5–6 µm
diameter spherical particles, 4.6 mm × 25 cm.
(c) Filters.—Polyvinylidene fluoride (PVDF) hydrophilic,
0.45 µm porosity.
C. Reagents
(a) Mobile phase.—52/48 ratio of aqueous/organic phases:
(1) Aqueous.—Acetic acid–H2O (1 + 99). (2) Organic.—LC grade
CH3CN.
(b) Chlorpropamide standard solution.—Transfer ca 50 mg,
accurately weighed, USP Chlorpropamide Reference Standard to
100 mL volumetric flask and dissolve in mobile phase. Dilute
quantitatively to final concentration of ca 0.05 mg/mL in mobile
phase.
(c) Resolution solution.—Chlorpropamide +
p-chlorobenzene-sulfonamide (PCBS) (ca 0.05 mg/mL of each) in
mobile phase.
D.
Transfer accurately weighed portion of finely ground tablets
equivalent to 45–55 mg chlorpropamide to 100 mL volumetric flask.
Add ca 70–80 mL mobile phase and shake thoroughly 6–8 min (or
sonicate 3–4 min) and dilute to volume with mobile phase. Dilute
quantitatively to final concentration ca 0.05 mg/mL in mobile phase.
Filter portion through 0.45 µm filter for LC analysis.
E. System Suitability
Set mobile phase at flow rate ca 1.5 mL/min. Retention time for
chlorpropamide should not be <4.0 min. Adjust flow rate and/or
solvent ratio (do not exceed 50% CH3CN) for desired retention time.
Column should conform to following performance parameters:
theoretical plates (n) not <1500; tailing factor (T) not >1.5; resolution
(R) between chlorpropamide and PCBS not < 2.0. Relative standard
deviation for 4 consecutive standard injections should be <2.0%.
F. Determination
Make 20 µL injections of standard and test solutions. Determine
peak responses (area or height) obtained and calculate amount of
chlorpropamide:
Chlorpropamide, mg/tablet = (r/r′) × (C/W) × DF × ATW
where r and r′ = responses for test and standard solutions,
respectively; C = concentration of chlorpropamide standard
solution, mg/mL; W = test portion weight, g; DF = dilution factor for
test solution, mL; ATW = average tablet weight, g/tablet.
Reference: JAOAC 69, 519(1986).
CAS-94-20-2 (chlorpropamide)
 2005 AOAC INTERNATIONAL