Quantitative Estimation of Saffron Components by Using HPLC an Approach for The Determination of Saffron Quality

Saffron is composed of four active major ingredients name Crocin-I, Crocin-II Picrocrocin and Safranal. Crocin’s are related to produce golden red color whereas picrocrocin and safranal are responsible for the taste and aroma. Spectrophotometric analysis of saffron is available but methodology of HPLC analysis is not readily available for quality checking of saffron. In this study each component of saffron was identified and analyzed by HPLC for better quality assessment. Methods: 200 ppm of saffron sample was prepared in 50% methanol and filtrated through 0.22 µm syringe filter. Crocin I and II were determined at 440 nm where methanol:0.1% phosphoric acid (55:45) was used as mobile phase. Picrocrocin was determined at 257 nm and mixture of acetonitrile:water (13:87) was used as mobile phase. Determination of safranal was done at 330 nm where gradient mixture of acetonitrile, phosphoric acid and water was used as mobile phase. Separations were done by using C18 250×4.6, 5 µm HPLC columns and injection volume was 20 uL. Column temperature was set at 25°C. Result: According to ISO 3632 protocols crocin concentration of saffron samples was found 246.00 picrocrocin was found 98.48 and safranal was found 41.965 which belongs to grade I, whereas in HPLC analysis crocin I was found 24.256 mg/g, crocin II was found 9.825 mg/g, picrocrocin was found 32.295 mg/g and safranal was found 0.512 mg/g. Conclusion: HPLC is the most suitable analytical techniques used for qualitative and quantitative determination of different chemical constituents, mixtures of organic components and biological extracts etc. HPLC will be the most suitable and alternative analytical technique for qualitative and quantitative estimation of saffron constituents.