Development and Validation of a Simple Isocratic HPLC Method for Simultaneous Estimation of Phytosterols in Cissus quadrangularis

Cissus quadrangularis L. is a promising remedy prescribed in the ancient Ayurvedic literature for bone fracture healing properties. As this activity has been extensively investigated and well established, a range of formulations containing C. quadrangularis has been marketed. This work reports the development and validation of a reliable RP-HPLC method for the analysis of phytosterols in the various extracts of the plant. The proposed method utilizes a Cosmosil C(8) column (250 ΄ 4.6 mm) with a compatible Phenomenex C(8) guard column with isocratic elution of acetonitrile and water (95:5 v/v) at 25°. An effluent flow rate of 2 ml/min and UV detection at 202 nm was used for the analysis of phytosterols. The described method was linear in the range of 1-500 μg/ml, with excellent correlation coefficients. The precision, robustness and ruggedness values were also within the prescribed limits (less than 2%). The recovery values were within the range, which indicates that the accuracy of the analysis was good and that the interference of the matrix with the recovery of phytosterols was low. The phytosterols were found to be stable in a stock solution for 48 h (% RSD was below 2%) and no interfering extra peaks were observed under controlled stress conditions. The proposed method is simple, specific, precise, accurate, and reproducible and thus can be used for routine analysis of C. quadrangularis phytosterols in quality control laboratories.