Induction of Callus from Pistils of Crocus sativus L. and Production of Color Compounds in the Callus

Abstract

The calli were induced from the pistils of saffron. MS, 1/2 MS, and NN medium with BA and NAA at 1 mg/l each were the most effective for the induction and growth of the calli. The calli produced crocin, crocetin-di (monoglucosyl-diglucosyl) ester and crocetin-di (monoglucosyl) ester. The calli producing those pigments have been maintained for more than two years under dark conditions. Saffron, Crocus sativus L., was originated from south Europe and has been recognized for its medicinal properties for a long time. The serum cholesterol levels of rabbits were reduced by half after intra muscular injection of crocetin. 1) Its principle value, however, now is as a food spice and colorant.2) The main pigments of saffron are mono-and diglycosyl esters of crocetin. Besides these crocetin derivatives, small amounts of a-carotene, B-carotene, lycopene and zeaxanthin have been isolated.3) The cultivation of saffron and harvesting of pistils require intensive labor.We have tried to produce color and medicinal compounds from callus derived from the pistil of saffron in vitro. Here we show the induction of callus from saffron, Crocus sativus L. and the pigmentation. Materials and MethodsBulb of saffron. The bulb of saffron was purchased from a local market and sprouted at room temperature under the natural sun beam. The sprouts sized 3-5 cm were cut with a blade and sterilized by dipping into 70% ethanol 2-3 times. The sheaths of the sprouts were removed and the pistils collected from the sprouts. The pistils were cut about 5 mm long from the top of the stigma and used as explants. Six explants were placed onto the 10 ml of agar medium in a disposable plastic petridish.Culture of explants. The explants were cultured for three months under the continuous illumination of fluorescent lamp at 3,000lx at 20C. The callus induction was checked with a microscope at one month after the inoculation. The growth of the callus were checked with the naked eyes at three months after the inoculation.The explants were also cultured in a flask containing 5 ml of liquid media listed in Table 2 and gyrated at 100 rpm.Maintenance of the callus. Three months after the initiation of culture, half of each callus was maintained by changing the agar medium every three months under light conditions. The rest of the half was kept in the dark. When the callus was placed onto the new medium a part with no color was removed by cutting with a blade. Since no difference in color was observed between the calli maintained under the dark and that in light conditions for one year, all calli have cur-

Chemical and sensory analysis of saffron produced through tissue cultures of Crocus sativus

“…The concentration of crocin and picrocrocin in TC stigmas was lower by 6 and 11 times respectively than the natural stigmas. Such a trend in the pigment composition was also observed in earlier studies (Hori et al 1988;.…”

“…It grows under specific climatic conditions and the harvesting of flowers needs skilled labour, which leads to the high cost of this spice. Attempts have been made to induce saffron in tissue cultures from stigmas (Hori et al 1988;Sarma et al 1990) and ovaries . Recently Fakhrai & Evans (1990) induced stigma-like structures successfully by using different floral explants of Crocus sativus.…”

Ultrastructure of chromoplasts and other plastids inCrocus sativusL. (Iridaceae)

“…The use and demand of saffron are increasing, but its world production is low compared to the demand, due to the high cost of production. Many in vitro cultures and biotechnological experiments have been performed to produce crocetin glycosilated derivatives and other saffron components Hori et al, 1988;Sarma et al, 1991) and new techniques have been applied to analyse saffron (Amelotti & Mannino, 1977;Basker & Neghbi, 1983. Recently, corms and leaves of Crocus sativus have also been investigated as a source of new important compounds of medicinal and industrial use (Escribano et al, 2000;Bergoin et al, 2003).…”

Cryopreservation of Spices Genetic Resources

“…Johnson et al (1996) reported biotransformation of ferulic acid vanillamine to capsacin and vanillin in immobilised cell cultures of Capsicum frutescens. Reports on the in vitro synthesis of crocin, picrocrocin and safranel from saffron stigma (Himeno and Sano, 1995) and colour components from cells derived from pistils (Hori et al, 1988) are available for further scaling up. Callus and cell cultures were established in nutmeg, clove, camphor, ginger, lavender, mint, thyme, celery etc.…”