Optimization of in vitro conditions for stigma-like-structure production from half-ovary explants of Crocus sativus L

Summary Studies were conducted to optimize the in vitro production of stigma-like-structures (SLS) that yielded the important biochemical constituents responsible for the color, taste, and aroma naturally found in the stigmas of autumn crocus. Immature half-ovary explants were evaluated for the frequency of proliferation of SLS by culture on five basal media supplemented with different combinations of plant growth regulators and vitamins. The optimum proliferation of SLS was observed on B5 basal medium containing α-naphthalene acetic acid (NAA) (5.4 μM), N⁶-benzyladenine (BA) (44.4 μM), MS organics, casein hydrolysate (0.05%), and l-alanine (11.2 μM) 50–60 d after inoculation. Some explants formed other structures (roots, corms, petals, leaves), the growth and development of which substantially reduced the development of SLS. Removal of brown tissues and other tissues during subculture (3-wk intervals) allowed continuous culture of halfovary explants for 9–10 mo. SLS that had deep red pigment and reached more than 0.5 cm length were removed from the explants and with concomitant reculturing of the ovaries, enabled SLS to be harvested three times. Activated charcoal (1%) added to B5 basal containing NAA (5.4 μM), BA (44.4 μM), and sucrose (3%) was found to be a helpful addendum to prevent browning of explants and accelerate the initiation, growth, and development of SLS. The amounts of crocin, crocetin glucosyl esters, picrocrocin, and safranal in SLS, as determined by high performance liquid chromatography analysis, were found similar to those in natural saffron.     

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

Stigma-like structures were produced in tissue cultures (TC stigmas) from the ovary explants of C. sativus on MS medium supplemented with NAA and BA. The size of these structures was 2 to 3 cm in length. At higher concentrations of both NAA (54 µM) and BA (44 µM) white tubular abnormal structures were observed from ovary explants in addition to the TC stigmas. Crocin and picrocrocin, responsible for colour and bitter taste respectively, were found to be 6 and 11 times lower in TC stigmas than in the natural stigmas. The saffron obtained from tissue cultures was subjected to sensory analysis and compared with the data obtained from chemical analysis. The sensory data indicated that the saffron pigments produced in tissue cultures were one tenth that of natural stigmas. Sensory profile test showed that the tissue culture saffron was low in floral, spicy and fatty characteristics as compared to saffron obtained from flowers. This is the first report on the sensory analysis of a spice produced in tissue cultures.Abbreviations BA benzyladenine - NAA naphthaleneacetic acid