Simultaneous Estimation of Luteolin and Apigenin in Methanolic Leaf Extract of Cardiospermum Halicacabum by HPLC

Abstract

Cardiospermum halicacabum (Sapindaceae) widely spread in Asia, tropical and sub-tropical Africa is commonly consumed as leafy vegetable for the treatment of rheumatoid arthritis in India. The present study was aimed to develop and validate an RP-HPLC separation for the simultaneous estimation of luteolin and apigenin in the leaves of Cardiospermum halicacabum. HPLC separation was accomplished on an oyster ODS 3 column (5 µm, 200 mm × 4.6 mm, i.d.) with gradient elution of potassium dihydrogen phosphate buffer (20 mM, pH 3.5 adjusted with ortho phosphoric acid, v/v) (A) and acetonitrile (B). The elution program as follows; 0-10 min 15-60% B, 10-15 min 60-75%B, 15-20 min 75-100% B, 20-25 min B hold at 100%. The method was validated as per ICH guidelines for system suitability, linearity, specificity, precision, accuracy, LOD and LOQ. The method was linear in the range of 1-70 µg/ml. The relative standard deviation of the method was less than 2% for intra-day and inter-day precision assay and the average recoveries were between 98 and 102%. LOD, LOQ of luteolin and apigenin was found to be 0.03, 0.91 and 0.041, 0.13 µg/ml respectively. The proposed method was found to be linear, specific, accurate, precise and robust