Subash Chandra Verma

gIsolation and purification of acetylshikonin and β-acetoxyisovalerylshikonin from cell suspension cultures of Arnebia euchroma (Royle) Johnston using rapid preparative HPLC*

Shikonin and its derivatives are important red colored naphthoquinone pigments found in a large number of Arnebia species, including A. euchroma, that are responsible for the various pharmacological activities exhibited by the plant. The precise separation of each naphthoquinone is essential for total quality evaluation and bioactivity analysis of herbal formulations of A. euchroma. Furthermore, the overexploitation of this useful plant has resulted in species becoming endangered. With this in mind, a simple and rapid preparative scale HPLC method with single compound recovery for the isolation and purification of two shikonin derivatives (i. e. acetylshikonin, beta-acetoxyisovalerylshikonin) from cell suspension cultures of A. euchroma is presented. The compounds were separated on a C(18) column within 10 min using acetonitrile/methanol (95:5) as mobile phase in isocratic mode. The isolated compounds were found to be more than 98% pure. The LOD for acetylshikonin and beta-acetoxyisovalerylshikonin was estimated at 0.063 and 0.146 mug/mL, respectively, while the LOQ was found to be 0.209 and 0.487 mug/mL, respectively. The recoveries accomplished for both the shikonin derivatives were in the range of 94.7-96.8%. The repeatability, expressed as %RSD, of acetylshikonin and beta-acetoxyisovalerylshikonin was found to be 1.74 and 1.27, respectively.

Microwave-assisted extraction and rapid isolation of ursolic acid from the leaves of Eucalyptus × hybrida Maiden and its quantification using HPLC-diode array technique.

Ursolic acid (UA) is the most important bioactive phytoconstituent of Eucalyptus × hybrida Maiden leaves and exhibits anticancer, antimutagenic, anti-inflammatory, antioxidative, and antiprotozoal activities. In this study, microwave-assisted extraction technique was employed for rapid isolation of UA from the leaves of Eucalyptus × hybrida and simultaneously HPLC-diode array method was developed for the quantification of UA. Effects of several experimental parameters on the extraction efficiencies of UA, such as type and volume of extraction solvents, microwave power and extraction time, were evaluated. The optimal extraction conditions were found to be 20 mL of a mixture of chloroform/methanol, 60:40; liquid-to-material ratio, 4:1; preleaching time, 10 min; microwave power, 600 W; temperature, 50°C; and microwave irradiation time, 5 min. Under the optimum conditions, the yield of UA was found to be 1.95 ± 0.08% in the dry leaves of Eucalyptus × hybrida. The results showed that microwave-assisted extraction is a more rapid extraction method with higher yield and lower solvent consumptions than the conventional method. It is a faster, convenient, and appropriate method and it may be used for rapid isolation and quantification of UA and other important phytoconstituents present in the leaves of Eucalyptus × hybrida.

Microwave extraction and rapid isolation of arjunic acid from Terminalia arjuna (Roxb. ex DC.) stem bark and quantification of arjunic acid and arjunolic acid using HPLC‐PDA technique

Arjunic acid and arjunolic acid are main bioactive components of Terminalia arjuna stem bark and reported for various biological activities. In this study, microwave-assisted extraction (MAE) of arjunic and arjunolic acid from stem bark of T. arjuna was investigated with developed and validated HPLC-PDA method, which resulted in the isolation of a novel anticancer molecule i.e. arjunic acid. Effects of several experimental parameters, such as type and volume of extraction solvents, microwave power, microwave extraction time, on the extraction efficiencies of arjunic, and arjunolic acid from stem bark of T. arjuna were evaluated. The optimal extraction conditions identified were 5.0 g quantity of stem bark powder, 20 mL of ethyl acetate, preleaching time 10 min, microwave power 600 W, temperature 65°C, and microwave irradiation time 5 min. The results showed that MAE is a more rapid extraction method with higher yield and lower solvent consumptions than reported methods. The HPLC-PDA analysis method was developed and validated to have good linearity, precision, sensitivity, and accuracy. MAE-HPLC-PDA is a faster, convenient, and appropriate method for isolation and determination of arjunic acid and arjunolic acid in the stem bark of T. arjuna.