Analyzing of Urine 3-Hidroxy Propil Mercapturic Acid on Cyclophosphamide Induced Rat to Determine Ameliorating Effect of Leucaena leucocephala (Lam.) de Wit Seed Extract

Background: A 70% ethanolic extract of Leucaena leucocephala (Lam.) de Wit seeds contain a certain amount of sulfhydryl active compounds and potential for ameliorated cyclophosphamide side effects. Objective: The aim of this study was to analyze of urine 3-hidroxy propil mercapturic acid (3-HPMA) in a rat model for cyclophosphamide-induced hemorrhagic cystitis to determine the effect of Leucaena leucocephala (Lam.) de Wit seeds extract. Materials and methods: The levels of urine 3-hidroxy propil mercapturic acid was measured using LCMS/MS. Urine preparation was carried out by dilution (1:5) with mobile phase and contained a deuteurated internal standard. The LC separation was performed using a C18 column (1.7 μm; 2.1 mm × 100 mm). The mobile phase consisted of formic acid (0.1%) and formic acid (0.1%) in acetonitrile (90:10 v/v), as a starting gradient at flow rate of 0.2 ml/min with total run 7 min. Results: The lower limit of quantification was 40 ng/ml and the linear extended to 20,000 ng/ml. The method has been successfully to be applied in determined of rat urine 3-HPMA. The results showed that the 70% ethanolic extract of seeds can increased the amount of 3-HPMA in rat urine. Conclusion: The 70% ethanolic extract of seeds displayed a potential ameliorating effect against cyclophosphamide side effects.