Quantification of Alendronate Sodium Hydrate and Its Related Compound According to JP Method (DS-413)

Alendronic acid is widely used for the treatment of osteoporosis. According to the JP method (17^th edition), quantification of alendronate sodium hydrate and its related compound should be done using HPLC with a column packed with styrene divinyl benzene copolymer gel. The method also mentions to pretreat the alendronic acid by adding 9-fluorenylmethyl chloroformate to treat the amino groups to form 9-fluorenylmethyloxycarbony (FMOC)-alendronic acid and FMOC-4-aminobutyric acid. The system suitability requires to satisfy resolution ≥ 6 and relative standard deviation (RSD) of peak area ≤ 2.0% (repeated six times) for FMOC-alendronic acid and FMOC-4-aminobutyric acid. RSpak DS-413 was confirmed to satisfy all required conditions for the analysis of alendronic acid.

Sample : 20 μL
prepared according to JP
1. FMOC-Alendronic acid
2. FMOC-4-Aminobutyric acid

Column    : Shodex RSpak DS-413 (4.6 mm I.D. x 150 mm)
Eluent    : (A) 10 mM Trisodium citrate + 10 mM Na2HPO4 aq. (adjusted to pH 8.0 with H3PO4)
                /CH3CN=85/15
            (B) 10 mM Trisodium citrate + 10 mM Na2HPO4 aq. (adjusted to pH 8.0 with H3PO4)
                /CH3CN=30/70
             Linear gradient ; (B%) 0 to 50 % (0 to 20 min), 50 to 100 % (20 to 35 min),
                                    100 to 0 % (35 to 36 min), 0 % (36 to 45 min)
Flow rate : 0.8 mL/min
Detector  : UV (266 nm)
Column temp. : 45 °C