University of Khartoum

Secondary Metabolites Productionin Cultured TissuesofDaturaandSolanum Species

Secondary Metabolites Productionin Cultured TissuesofDaturaandSolanum Species

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Title: Secondary Metabolites Productionin Cultured TissuesofDaturaandSolanum Species
Author: Kanani, Zeinab Mohammed Mekki
Abstract: The study aimed to establish tissue culture protocols for Datura and Solanum species and to investigate the biosynthetic potentialities of cultured tissues. Approaches to increase the production of secondary compounds by application of elicitors were followed. Antioxidant activity of these compounds was tested. Callus tissues of Datura stramonium, D.innoxia, Solanum nigrum and S.dubium were initiated on Murashige and Skoog (MS) medium supplemented with various combinations of auxins {2,4-dichlorophenoxy acetic acid (2,4-D), indole-3-acetic acid (IAA), α-naphthalene acetic acid (NAA)}, and cytokinin {6-benzyl amino purine (BAP), Kinetin} were used. Standard methods of biochemical analysis for alkaloids were followed. The combined method gas chromatography/ mass spectrometry (GC/MS) was used for alkaloids determination. Polyphenol content was determined using modified Folin-Ciocalteu method. Flavonoids content was determined usingNaNO3, AlCl3 and NaOH.1, 1-diphenyl-2-picryl hydrazyl (DPPH) assays were used for the evaluation of free radical-scavenging properties of methanolic extracts of intact organs and callus tissue. Tissue culture was initiated from leaves and sterile seedlings after breaking seed dormancy. Callus culture was established and the most effective growth regulators were Kin, IAA and 2,4-D for D.stramonium leaf explant, S.nigrum leaf and seed and S.dubium leaf explant. The concentration 1mg/l Kin with 0.5mg/l 2,4-D was the best for callus formation, and the calli of S.dubium formed very rapidly and were the biggest among all other species. D.innoxia leaf initiated callus on MS medium supplemented with 2mg/l Kin and 0.5-1mg/l 2,4-D. MS medium supplemented with 4mg/l IAA, 1.5mg/l Kin and 0.5mg/l 2,4-D was the best for seed explant of D.stramonium and D.innoxia as well. 2,4-D alone was better for callus maintenance and growth, also it was the best for S.dubium seed although the callus initiation was very slow. In S.nigrum, the callus formation percentage of leaf explant was low when cultured on MS medium with NAA and BA. UV light increased callus mass; white tissues appeared in D.stramonium seed callus, D.innoxia and S.nigrum leaf calli. Calli from the four species treated with NaCl senescenced and died. There were no remarkable changes on callus treated with GA3. Insertion of platinum wire in callus tissue enhanced callus growth in D. stramonium, D.innoxia and S.nigrum, resulting in a compact calli for seeds and leaves, while S.dubium leaf became more friable.Phytochemical screening of intact plant tissues and callus cultures of the species under investigation indicated that higher value for total alkaloid was obtained from S.dubium leaves, whereas the lowest values were recorded in D.innoxia fruits and in S.nigrum seeds, fruits and leaves. In callus tissue, the highest mean total alkaloid recorded for S.dubium while the lowest values were scored in D.stramonium, D.innoxia and S.nigrum. S.nigrum seeds treated with GA3 recorded the highest value for total alkaloid, whereas S.nigrum leaves under control conditionsand leaves treated with platinum wire scored the lowest values withS.dubiumleaves treated with GA3and D.stramoniumseeds under control conditions. Seeds as explants were better than leaves in alkaloid production while intact plant leaves were higher than seeds and fruits. The quantitative analysis for atropine by GC/MS recorded that D.innoxia seeds treated with NaCl had the highest atropine concentration (18.82ppm). The highest mean total polyphenol/plant recorded for D.stramonium while the lowest value recorded for S.nigrum. In callus tissue, S.dubium scored the highest value for total polyphenol, also the interaction between species, plant part and treatment recorded that S.dubium seeds under UV light had the highest value for total polyphenols. While the lowest total polyphenols value was recorded in S.nigrum and for the interaction, D.stramonium seeds treated with platinum wire showed the lowest value as well. In flavonoids detection S.nigrum leaves scored the highest value for total flavonoids in intact plant. The calli formed from S.dubium scored the highest value for total flavonoid while D.innoxia and S.nigrum recorded as the lowest source for total flavonoids. Leaves of S.dubium treated with NaCl recorded the highest value for flavonoid whereas leaves of S.nigrum treated with GA3 scored the lowest value. In intact plant the highest antioxidant activity was detected in S.dubium while the lowest found in S.nigrum especially the seeds. The results of antioxidant activity in callus tissues showed that D.stramoniumseeds under control conditions scored the highest value although the lowest value was recorded for S.dubium leaves under control conditions. The effect of elicitor differed from one test to another, in alkaloid detection there were no significant differences between them while NaCl scored the highest atropine concentration. For polyphenols UV light recorded as the best elicitor whereas GA3 was the worst. In flavonoids contents NaCl was the best elicitor. Abiotic elicitors had a great effect on increasing secondary metabolite production among the four species, there was a variation in secondary metabolites production between intact plant and callus, also plant parts were described as an important factor in secondary metabolites production. Recommendations for future work: Screening other members of the wild growing Solanaceae for secondary metabolites production, also work on cell, callus and suspension cultures to scale up production of secondary metabolites (alkaloids).
URI: http://khartoumspace.uofk.edu/123456789/25369


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