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AN AGE OLD TREASURE OF PLANTS FOR

THE NEW WORLDS MEDICINE : BIOTECHNOLOGICAL PERSPECTIVE

PROF. (DR.) G. B. DIXIT.


DEPARTMENT OF BOTANY

SHIVAJI UNIVERSITY KOLHAPUR

Dedicated to

Late Dr. V. N. NAIK


A devoted Plant taxonomist of our country

MEDICINAL
PLANTS
Plants have been used for medicinal purpose not only by humans since prehistoric timesbut are also used to History of medicinal plants in treat our livestock. India
(4500 B.C.-1600 B.C.) - 67 medicinal plants have been mentioned.
(2500 B.C. -1500 B.C.) - mention has

Rig-Veda AtharvaVeda

been made of about 290 plants used as remedies for curing diseases.

Ayurveda is UPVEDA of Atharvaveda whose authority was Lord Dhanvantri, who established the real During this period Greek and Romans function of earliest Took substantial medical science. material from India to enrich their medicinal knowledge (Pal and Jain, 1998). Higher plants have been the traditional source for raw materials and medicinal formulations, ever since the dawn of civilization.

MEDICINE
Art of preserving and restoring health especially by other means than surgery, drugs, potions etc.
Plants Sources of medicines Animals Microbe s Minerals

Plant Medicines
For many centuries in all great literature, cultures like Hindu, Buddhist, Confucian, Islamic, Roman, Christian the practice of medicine was associated with certain moral behaviours, qualities and values.

The moral physician was competent, dedicated caped confidences and cared for the sick poor without charge. As healers gradually formed themselves into practitioners question of relationships between practitioners and of economic rivalry also appeared as the agenda of medical ethic. At mid twentieth century, the traditional medical morality encountered unprecedented problems. At the same time science had brought much more effective treatments than had ever existed. Physicians started earning more money and thereby social prestige than the profession had previously enjoyed. Old ideas about medical morality were challenged by these conditions.

In general twentieth century had many more inventions which has changed the face of this world. To name few, Mendelian
genetics, Hybrid breeding, mutation breeding, molecular biology, r-DNA technology etc. along with tremendous development in the field of The big challenge was of ever increasing energy, electronics and human population and various ailments/ instrumentation. diseases like cancer, aids and other diseases of viral origins, epidemic in nature and to date

Resistance to antibiotics, antimalarials and other drugs occurs with increasing frequency. Crisis like these have created a new demand for Phytochemical screening programmes to detect antiviral plant products. In this context, zoopharmacognosy
can also play an important role (e.g. Rubia chimpanzee interaction, Rubiatriol, anticancer agent. Since these primates have coexisted and co evolved with the plants of the Thus the increasing demand and decreasing plant tropical forests for millions of years). resources indicate the need of multifaceted program develop which includes conventional and nonconventional ways to conserve the knowledge and

What is Biotechnolog
Biotechnology defines any technological application that uses a biological system, living organisms or derivates thereof to make or modify products or processes for specific use (Conservation of Biodiversity). Plant tissue culture was originally developed as a research tool to study the physiology and biochemistry of plants, without the complication of It the whole-plant structure. has made significant contributions in:

The production of plant material Plant breeding Gene banks The production of chemical compounds

SEED CULTURE EMBRYO CULTURE


MERISTEM CULTURE

CELL CULTURE

PLANT TISSUE CULTURE

PROTOPLAST CULTURE

CALLUS CULTURE BUD CULTURE

ORGAN CULTURE

ADVANCED TECHNIQUES USED IN PLANT TISSUE CULTURE


Agrobacteri Somatic

Extraction
and analysis of active principles from cell cultures

um
mediated genetic

Micropropa gation

embryogen

esis

transformat
ion

The first commercial process to be developed was micropropagation, which is now a worldwide industry producing millions of plants per annum. Micropropagation is the practice of rapidly multiplying stock plant material to produce a large number of progeny plants, using modern plant tissue culture Micropropagation is used to methods. multiply novel plants, such as those that have been genetically modified or bred through

The process of initiation and development of embryos or embryo-like structures from somatic cells

AGROBACTERIUM MEDIATED GENETIC TRANSFORMATION Agrobacterium tumefaciens - or Agrobacterium rhizogenes-mediated transformation is to date the most commonly used method for obtaining transgenic plants. The tumorigenic host plant species for range A. tumefaciens include Large number of dicots and some monocots Gymnosperms. Agrobacteria areand naturally occurring, ubiquitous soil

borne pathogens. A. tumefaciens causes crown gall disease (tumors). A. rhizogenes causes root hair disease (hairy root) Other bacterial groups also contain species capable of interkingdom genetic exchange (Gelvin 2005).

Renewed interest in the therapeutic potential of medicinal plants means that researchers are concerned not only ethnopharmacological usage of with validating plant, but also with identifying, isolating and characterizing the active The reason that certain plants components. are effective against particular diseases is because they contain specific chemical compounds (the active principles)

EXTRACTION AND ANALYSIS OF ACTIVE PRINCIPLES

BIOTECHNOLOGICAL PRODUCTION OF SECONDARY METABOLITES Medicinal plants are the most


exclusive source of life-saving drugs for majority of the population. Theworld's secondary metabolites are known to play a major role in the adaptation of plants to their environment and also represent an important source of pharmaceuticals. Scientists have developed a form of root culture
using Agrobacterium rhizogenes, the cause of hairy root disease. Cells transformed with some of the bacterias DNA, causes the cells to be more sensitive to the hormones they produce. The cells form into roots. These roots grow very fast and produce the secondary

critical step at the beginning of an extensive quality assurance process. Identification of plants at the species level is traditionally achieved by careful examination of the specimens macroscopic and microscopic morphology. Plant breeding, pharmacology, pharmacognosy and Physiological plant anatomy other quality control disciplines (Haberlandt,1884-1945), have also contributed lot in the in-vitro field. culture of plant cells (White, 1939,Gauthret, 1939), Phytochemistry (Mothes,1900-1983), the design of bioreactors (Tulecke

PLANT IDENTIFICATIO Unequivocal identification and authentication of the plants used N for production is an elementary and

DNA FINGERPRINTING

Medicinal Plants: DNA-Based Identification


Identification of plants at the species level is utmost essential while making the medicines from plants. Identifying Species (DNA-Barcoding) Any type of organism can be identified by examination This genomic of DNA sequences which is unique to that species. fingerprinting can differentiate between individuals, species and populations and has proven useful for the characterization of sample homogeneity and detection

Molecular biological methods used for the Name Acronym authentication of medicinal plants
Polymerase chain reaction Allele-specific diagnostic PCR Amplification refractory mutation system Amplified fragmented length polymorphism Arbitrarily primed PCR Direct amplification of length polymorphism Multiplex PCR PCR-selective restriction Randomly amplified polymorphic DNA Sequence characterized amplified region Restriction length polymorphism DNA microarray DNA sequencing PCR-SR RAPD SCAR RFLP ARMS AFLP AP-PCR DALP PCR

Inter simple sequence repeat-anchored PCR


Multiplex amplification refractory mutation system Simple sequence repeat polymorphism

ISSR-PCR
MARMS SSR

Nuclear and chloroplast genes used for authentication of medicinal plants Gene Genome
18S rRNA Internal transcribed spacers (ITS) of 18S, 5.8S and 26S rRNA Nuclear Nuclear

Intergenic spacer of the 5S rRNA (5S gene spacer)


26S rRNA atpA, atpB, atpF, atpH chlB matK psbA, psbK, psbI rbcL rp14, rpl16 rpoB, rpoC1 rps16 trnC, trnD, trnF, trnK, trnL

Nuclear
Nuclear Chloroplast Chloroplast Chloroplast Chloroplast Chloroplast Chloroplast Chloroplast Chloroplast Chloroplast

Current development in tissue culture technology indicate that transcription factors are efficient new molecular tools, for plant metabolic engineering to increase the production of valuable compounds. Such as taxol, morphine, codeine, L Dopa, Berberin, Diosgenin, Vinblastin and Vincristine etc. Taxol : (plaxitaxol), a complex diterpene alkaloid found in the bark of Taxus brevifolia, is one the most promising anticancer agents known to its unique In 1989, Christen et al., reported for mode of action on the microthe first time the production of taxol by tubular cell system (Jordan Taxus cell cultures various factors and Wilson, 1995 influencing stability and recovery of paclitaxol from suspension cultures by Taxol Nguyea et al., 2001, Wu et al., 2001

RESEARCH ACTIVITIES OF OUR LABORATORY


Protocols for micropropagation of different valued plants have been established. Biochemical assay of some medicinally important plants are also being carried out to assure the quality of the regenerants.

Micropropagation of Hemidesmus indicus and Rubia cordifolia has been established by using different phytohormones. Hemidesmus indicus Rubia cordifolia

Progressive Stages of tissue culture in Hemidesmus indicus


Callus

Hardened

Multiple shoot induction in Hemidesmus indicus.

Rooting

Somatic embryogenesis in Hemidesmus indicus

Rubia cordifolia

Progressive stages of tissue culture in Rubia cordifolia

Ghatge S. R., Kudale S and Dixit G. B. An Improved Plant Regeneration System for High Frequency Multiplication of Rubia cordifolia L.: A Rare Medicinal Plant. Asian Journal of Biotechnology,3(4) 397-405, 2011.

Tissue culture in Drimia spp.

Proscillaridin A

HPLC profiles of standard Pro. A

Protocol for in vitro multiplication of Chlorophytum sp., a valued medicinal plant has been established.
MS + BAP (1mg/l)

MS + 2,4-D (2mg/l) + BAP (1mg/l)

MS + IBA (1.5mg/l)

Plantlet Regeneration in Chlorophytum sp.

MS + 2,4-D (2mg/l) + BAP (1mg/l) MS + 2,4-D (2mg/l) + BAP (1mg/l)

Saw dust + MS basal (liquid

Soil

Tissue culture in Grape

Fruits of Grape cv

Kedage V. V., Jai C. Tilak, G. B. Dixit, TPA Devasagayam and MinaI Mhatre Antioxidant Properties of Different Varieties of Grapes (Vitis Vinifera L.) in India. Critical Reviews in Food Science and

Suitable protocol for in vitro regeneration of Nothapodytes nimmoniana, which is a valued medicinal plant, has been established.

Camptothecin

Nothapodytes nimmoniana (Grah.) Mabb. Pai S. R., Nimbalkar M. S., Pawar N. V., Patil R. P., and Dixit G. B. Seasonal
discrepancy in phenolic content and antioxidant properties from bark of Nothapodytes

Protocol for in vitro regeneration of Ancistrocladus heynenanus has been established

Ancistrocladus heynenanus Wall .ex Grah.

120 DAYS MS + BAP 3.0

120 DAYS WPM + BAP

Zingiber officinale Rosc.


An effective protocol for Ginger in vitro regeneration was established and Hardening of the tissue culture raised Pawar N. V., Pai S. R., Nimbalkar M. S and Dixit G. B. RPplantlets was done successfully.

HPLC analysis of Phenolic Antioxidant compound 6gingerol from different ginger cultivars. Food Chemistry. 126: 1330-1336, 2011.

[6] Gingerol
O CH3O HO

H
O

Germplasm collection

Tissue cultural stud

Conservation of Ceropegia species through Biotechnological tools

Flower

Multiple shoots

Micropropagation of Ceropegia

In vitro tuber formation

Micropropagation of Ceropegia panchganiensis Blatt & McCann

Agrobacterium mediated genetic transformation induced hairy roots in Rubia cordifolia and quantification of Alizarin

ALIZAR IN

DNA barcoding for identification of the enigmatic plant Ramkand

Agave sisalana
Nimbalkar et al. (2011) DNA barcoding for identification of the enigmatic plant

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