HPLC determination of flavonoids in hairy-root cultures of Scutellaria baicalensis Georgi

A simple and sensitive liquid chromatographic method has been established for determination of the main flavonoid components of Scutellaria boicolensis Georgi (Lamiaceae) roots and hairy-root cultures. Wogon, the dried root of the plant, is used in traditional Chinese medicine for treatment of bronchitis, hepatitis, tumors, and inflammatory diseases. Lyophilized hairy roots were extracted with methanol. The crude extracts were purified by SPE on Supelco LC-8 cartridges. HPLC separations were performed on a Eurospher 100-C-8 reversed-phase column. The mobile phase was a gradient prepared from mixtures of acetonitrile and 0.1% trifluoroacetic acid. Peaks were identified by addition of standards and/or by diode-array detection. Baicalein 7-O-glucuronide (baicalin), wogonin 7-O-glucuronide (wogonoside), baicalein, wogonin, and acteoside were determined by the external standard method at 280 nm. We found that the aglycon (baicalein and wogonin) content of the transformed roots was consistently higher than that of the intact root from Siberia

Analysis of polyacetylenes by HPLC in hairy root cultures of Lobelia inflata cultivated in bioreactor

Lobelia inflato L. contains pharmacologically important secondary metabolites. The main alkaloid is lobeline which has a stimulatory effect on the respiratory centre. Recently high quantity of polyacetylenes was determined in the plant and hairy root cultures. With the aim of increasing of the secondary metabolite production hairy root cultures were produced (by direct infection of the sterile plants with Agrobacterium rhizogenes strain R1601). A new HPLC method was developed for the determination of lobetyolin and lobetyolinin, the main polyacetylene compounds of L. inflata hairy roots. The analysis was performed on Hypersil MOS column (250 x 4 mm I.D.). The mobile phase was 15:85 (v/v) mixture of acetonitrile and water. Peaks were identified by addition of standards and by DAD. The external standardized method allowed the determination of lobetyolin and lobetyolinin with good sensitivity and reliability. This method was used to measure the polyacetylene contents of hairy roots grew intensively in bioreactor. High contents of lobetyolin (3.6%) and lobetyolinin (0.8-1.6%) were detected in the hairy roots because of their great biosynthetic capacity

HPLC analysis of alizarin and purpurin produced by Rubia tinctorum L. hairy root cultures

We have determined the quantities of the anthraquinones alizarin and purpurin, with especial regard to their effective antigenotoxic activity, in genetically transformed hairy root cultures of Rubia tinctorum L. Hairy roots were cultured on solid and in liquid Gamborg B5 and 1/2 NMS media in a shaking cabinet and in a bioreactor. Methanolic extracts of lyophilized hairy roots were hydrolysed, and then purified by solid phase extraction with good recovery. A new HPLC method was developed for the determination of alizarin and purpurin. The analysis was performed on a Luna C8 RP column using a 45:55 (v/v) mixture of acetonitrile:20 mM ammonium formate-formic acid buffer (pH 3.00) as eluent. Peaks were identified by addition of standards and by diode-array detection. External standardization allowed the determination of alizarin and purpurin with good sensitivity and reliability. The maximum purpurin content was observed in cultures cultivated on solid B5 medium (5.94 mg g(-1)). The highest alizarin content was measured in cultures cultivated on solid 1/2 NMS medium (2.14 mg g(-1))