Secondary metabolite production

Secondary metabolites are organic compounds that are not directly involved in the normal growth, development, or reproduction of an organism.

1. SECONDARY METABOLITE PRODUCTION

2. INTRODUCTION Higher plants synthesize a wide variety of low molecular weight compounds in addition to the essential primary metabolites. These are called secondary metabolites These secondary metabolites offer protection against pests,they act as attractatnts and as the plant’s own hormones

3. WHY SECONDARY METABOLITES Chemically warfare to protect plants from the attacks by predators, pathogens, or competitors Attract pollinators are seed dispersal agent Important for abiotic stress Medicine Industrial additives

4. CLASSIFICATION OF SECONDARY METABOLITES They are classified on the basis of 1)chemical structure 2)chemical composition 3)solubility in various solvents 4)pathways by which they are synthsized

5. A simple classification of secondary metabolites includes three main groups 1)the terpenes - made from mevalonic acid, composed almost entirely of carbon and hygrogen 2) phenolics – made from simple sugars, containing benzene rings, hydrogen and oxygen 3) nitrogen-containing compounds – extremely diverse, may also contain sulphur

6. ALKALOIDS Most are derived from a few common amino acids (i.e..tyrosine , tryptophan , ornithine or argenine and lysine Compounds have a ring structure and a nitrogen residue Indole alkaloids is the largest group in this family , derived from tryptophan Widely used as medicine

7. CHARACTERISTICS They are bitter in taste. Derived from amino acids.The amino acids that are most often serve as alkaloidal precursors are: phenylalanine, tyrosine, tryptophan, histidine , anthranilic acid, lysine and ornithine . Alkaloids form double salts with compounds of mercury, gold, platinum and other heavy metals. These salts are obtained as precipitate which are microcrystals .

8. Insoluble or sparingly soluble in water, but the salts formed on reaction with acids are usually freely soluble. Most are crystalline solids although a few are amorphous.

9. Free alkaloids are usually soluble in polar solvents like ether, chloroform Some alkaloids are liquid because of lacking of oxygen in their molecules. ( e.g coniine, nicotine, spartenine )

10. Sources and Occurrence of Alkaloids Alkaloids can occur in plant kingdoms; among the angiosperms, Leguminosae , Papaveraceae , Ranunculaceae , Rubiaceae , Solanaceae , Berberidaceae are outstanding alkaloid-yielding plants.

11. Uses of Alkaloids in Plants: Poisonous agents which protect plants against insects and herbivores End products of detoxification reactions representing a metabolic locking-up of compounds otherwise harmful to the plants. For regulatory growth factors Reserve substance capable of supplying nitrogen or other elements necessary to the plant’s economy

12. TERPENES Terpenes are natural products that are structurally related to isoprene Isopropene (2-methyl-1,3-butadiene) H 2 C C CH 3 CH CH 2 OR

13. The number of C atoms is a multiple of 5, C 5 C 10 C 15 C 20 C 25 C 30 C 35 C 40 Each group of 5 C is an isoprene subunit They can be saturated or unsaturated Many contain O atoms as well.

14. PHENOLICS Derived from aromatic amino acids, such as phenylalanine, tyrosin , and trytophan . All contain structures derived from phenol Some examples: Coumarins : antimicrobial agents, feeding deterrents, and germination inhibitors. Lignin: abundant in secondary cell wall, rigid and resistant to extraction or many degradation reagents Anthocyanins Flavones Flavnols Phenols are present in every plant they attract pollinators to the plant and even impact how these plants act with one another. .

15. GLYCOSIDES Compounds that contain a carbonhydrate and a noncarbohydrate Glycosides are present in vacuoles in inactive form Glucosinolates : found primarily in the mustard family to give the pungent taste There are four type of linkages present between glycone and aglycone : C-linkage/ glycosidic bond, O-linkage/ glycosidic bond N-linkage/ glycosidic bond S-linkage/ glycosidic bond

16. Sources and effects of major secondary metabolites carotenoids are organic pigments occuring in plants and are mostly found in red orange and yellow fruits and vegetables . they have antioxidative effects and prevents cancer They boost the immune system and reduce the risk of heart attacks

17. PHYTOSTEROLS Found in- sunflower seeds , sesame,nuts and soy beans Protect against colon cancerand low cholestrol level

18. SAPONINS Flavour additives found in- legumes and spinach Boost the immune system Lower the cholestrol level in blood Reduce the risk of intestinal cancer

19. FLAVONOIDS Organic pigment Gives plant red violet or blue colour Inhibit the growth of bacteria and viruses Protect the cells against the damage of freeradicals Influence blood coagulation

20. SULPHIDES Contain sulphur Belongs to the lily family onions ,leeks, asparagus, garlic Inhibit the growth of bacteria Lower cholestrol levels Preventive efect against cancer

21. CURCUMIN Extracted from turmeric. Coloring and medicinal uses Wound healing Antiulcer Anti inflammatory Antimicrobial & antiviral Hepatoprotective Antioxidant No toxicity.

22. FLAVORS Cocoa seeds Vennila Strawberry Cinnamon Cardamom Cloves Pepper areca

23. PRODUCTION OF SECONDARY METABOLITES IN CULTURES/BIOREACTORS 1)choice of explants any part of the plant can be used dicotelydons are rather amenable for callus tissue induction procured explants are sterilized prior to culture by using ethanol

24. 2)NUTIENT CULTURE MEDIA To induce callus from an explant and to cultivate the callus and cells in suspension

25. 3)Callus induction A callus cell culture is usually sustained on gel medium. Callus induction medium consists of agar and a mixture of macronutrients  and micronutrients  for the given cell type. There are several types of basal salt mixtures used in plant tissue culture, but most notably modified Murashige and Skoog medium  White's medium ,  and woody plant medium .  Vitamins are also provided to enhance growth such as Gamborg B5 vitamins .  For plant cells, enrichment with  nitrogen  phosphorus, and potassium is especially important .

26. SUSPENSION CULTURE The growth rate of suspension cultured cell is generally faster than that of the solid culture. The former is more desirable, particularly in the production of useful metabolites on a large scale

27. FERMENTORS OR BIOREACTORS Carboy system-described by tulecke and nickell in 1959,consists of a rubber- stoppered 20 litre carboy fitted with four tubes. Filtered compressed air is employed for oxygen supply, aeration and agitation of the medium Stirred jar fermetor - used for microbial cultivation Agitation speed of 50 to 100 rpm

28. HAIRY ROOT CULTURE It is the culture produced after the infection of explants and cultures by the gram negative soil bacteria Agrobacterium rhizogens This process take advantage of the naturally occuring hairy root disease in dicotyledons

29. Agrobacterium recognizes some signal molecules exuded by wonded plant cells and becomes attached to it. The bacteria contain the Root inducing plasmid ( Ri -plasmid). The bacteria Genetically trasfer part of the Ri -plasmid called the trasfer DNA (T-DNA) to the plant genome .

30. The plant cell to proliferate by increasing the rate of cell division ( cytokine expression ) and cell elongation ( auxin expression) to produce the hairy roots. Produce the opines which is a type of unusual amino acids ( octopine , agropine , nopaline , mannopine amd cucumopine ) which is used by the bacterium as a carbon , nitrogen and energy source.

31. Induction of hairy root cultures In Vitro Explants are wounded and then inoculated with Agrobacterium rhizogenes . Usually two or three days later, the explant can be transferred into solid media with antibiotics, such as cefotaxime , vancomycine or ampicillin to kill or eliminate redundant bacteria. The hairy roots will be induced within a short period of time, which varies from one week to over a month depending on different plant species.

32. The decontaminated hairy roots can be subcultured on phytohormone –free medium .

33. Commercial production of secondary metabolites higher plants contain a variety of metabolic products which are useful as medicines ,food additives,perfumes,etc . however , non-compromising reduction in plant sources,increases in labour cost and other problems in order to obtain these high value added substances from natural plants ,have forced to develop analternative approachtowards the use of plant cell culture for production of these metabolic products .since ptc is not affected by changes in environmental conditions such as climate or natural depredation, improved production is possible

34. PHARMACEUTICALLY IMPORTANT CHEMICALS 1)Morphine alkaloids Codeine is an anlagesic and cough suppressing drug from Papaver somniferum also morphine can be converted into codeine many workers have tried to produce codeine by undifferentiated cells of these plants

35. BERBERINE it is an isoquinoline alkaloid which is produced and distibuted in roots of Coptis japonica and cortex of Phellonderon amurense Berberine chloride is used in treatment of intestinal disorders

36. TORPANE ALKALOIDS Scopolamine and hyoscyamine areused commercially as anesthetic and antispasmodic drugs. Occur in the leaves of members of the family Solanaceace including Duboisia myoporoides Datura leichhartdtti , Scopolia , Atropa, and Hyoscyamus

37. ANTITUMOUR COMPOUNDS

38. VINCA ALKALOIDS Extracted from Catharanthus roseus Treat leukemias , hodgkin’s disease and antitumor activity

39. GINSENG Roots of Panax ginseng is widely used as tonic Effective for genetic disorders, diabetes and weak circulation Root contains saponins and sapogeninis

40. FOOD COLORING AGENTS

41. SAFFRON Made from the stamens of Crocus sativus and used as flavouring agent and colourant Plant contains crocin (yellow pigment), safranal (fragrance), picocrocin (bitter substance) Baked goods, soaps, cheese Medicinal value for stomach ailments

42. SAFFLOWER YELLOW Yellow pigment obtained from the floret of the Carthamus tinctorius Major pigment is carthamin Carthamin is a quinoid form of isocarthamin Used in baked goods and beverages

43. CHEWING GUMS CHICLE Chicle a raw material is obtained from the latex of Achras sapota Chicle contains 60% resin and 15% of rubber Resin consisits of lupeo , α - amyrin and β - amyrin Rubber consists of cis and trans- 1,4-polyisoprene Synthesized through mevalonic acid pathway

44. TAXOL It is a ditrpene amide Extracted from the bark of wild grown Taxus brevifolia Treats breast and lung cancer