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Файл:Ординатура / Хирургия / Библиотека им академика М.И. Перельмана / Книга_5857_Библиотеки_им_академика_М_И_Перельмана.pdf
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- •Foreword
- •Contributors
- •Preface
- •Contents
- •1. General Pharmacology
- •2. Pharmacology of Peripheral Nervous System
- •3. Pharmacology of Cardiovascular System
- •4. Drugs Acting on Urinary System
- •5. Drugs Acting on Respiratory System
- •6. Pharmacology of Central Nervous System
- •7. Chemotherapy
- •8. Autacoids and their Antagonists
- •9. Pharmacology of Drug Acting on theGastrointestinal Tract
- •10. Immunopharmacology
- •11. Vitamin and Minerals
- •12. Hormones
- •1. Introduction to Pharmacognosy
- •2. Sources and Classification of Crude Drugs
- •3. Factors Influencing Quality of Crude Drugs
- •4. Techniques in Microscopy
- •5. Introduction of Phytoconstituents
- •6. Glycosides
- •7. Alkaloids
- •8. Terpenoids, Volatile Oils and Resins
- •9. Principles of Plant Classification
- •10. Pharmaceutical Aids
- •11. Plant Products
- •12. Toxic Drugs
- •13. Poisonous Plants
- •14. Enzymes
- •15. Quantitative Microscopy
- •16. Biogenetic Pathways
- •17. Herbarium
- •18. Herbal Formulation
- •19. Plant Tissue Culture
- •20. Herbal Cosmetics
- •21. Herbal Formulation
- •1. Cellular Components
- •2. Carbohydrates
- •3. Proteins
- •4. Lipids
- •5. Vitamins
- •6. Biological Oxidation and Reduction
- •7. Enzymes
- •8. Nucleic Acids
- •9. Hereditary Diseases
- •1. Plant Cell
- •3. Fermentation
- •4. Recombinant DNA Technology
- •5. Proteomics
- •1. Introduction to Microbiology
- •2. Microscopy
- •3. Staining Methods
- •4. Biology of Microorganisms
- •5. Fungi and Viruses

18. Herbal Formulation
Herbaldrugformulationdeneasdosageformconsisting
ofone or more herbs orprocessed herb(s), herbal
preparationsquantiedtoprovidespecicnourishingand
beautifying benet todiagnose, treat, mitigate diseases
ofhumanbeingsoranimalsand/ortoalterthestructure
orphysiology ofhumanbeingsoranimals.Herbaldrug
formulations are themeans by which plant drugor
preparationorisolatearedeliveredtositesofactionwithin
the body through topical, parental, nasal, and ophthalmic
routeofadministration.
AYURVEDIC FORMULATIONS
Ayurveda,the “Science of life,” or longevity, is more
than5,000 years old holistic alternative science from
India.AyurvedaoftencalledMother of all Healing and
isaimedatthephysical,mentalandspiritualwell-being
ofhumanbeings.
Ayurvedais based on Panchamahabhutas or the five
elements: Vayu (air),Teja (re), Aap (water), Prithvi
(earth) and Akasha(aether) which build-upthe living
microcosm (human beings) and the macrocosm (external
universe).When pooled collectively in pairs,the
Panchamahabhutasform Tridosha or the three humours,
namely Vata (responsiblefor body movement), Pitta
(responsiblefor bodily chemical reactions such as
metabolism and temperature) and Kapha(responsiblefor
growth, protection, lubrication and sustenance).
Types of Ayurvedic Formulations
1. Classical Ayurvedic medicines: TraditionalAyurvedic
textbooks as Charaka Samhita and Sushruta Samhita
contain the procedural information and formula to
formulatethesepreparations,e.g.Ark, Asava, Arishta,
Ghrita, Bhasma etc.
2.Proprietary medicines: These types ofmedicines are
alsoknownasmodernayurvedicmedicinesorpatent
medicines. The ingredients used to prepare such
medicinesarenotavailableinAyurvedictextbooks.The
manufacturingcompanydecidesitsformula,modeof
preparationanddosageform,e.g.syrupsandcapsules
TheAyurvedicformulationsaredescribedbelow:
Ark: Theseareobtainedbydistillationofherbs.Itislight
innatureandveryeasyassimilatedinsystem.Distillation
hasnotmuchtodowithsolubilitybutrathervaporisation
points.Itallowsavolatilesubstancetobeseparatedfrom
anon-volatilesubstance.
Asava and Arishtas: Asavaare Ayurvedic preparations
preparedwithnaturalfermentationusingsugar,waterand
herb whereas Arishta prepared by herbal decoctions. These
Section 2 Pharmacognosy
309

preparationshavelonger shelf-life. The main difference
betweenthetwoisinvolvementofheat.
Asava and Arishtaare unique liquid dosage forms that
containself-generated alcohol. Former is made with
decoctionsofherbsinboilingwaterwhilelaterisprepared
bydirectly using fresh herbal juices or decoction to
undergofermentations.ArishtasareclassicalAyurvedic
preparationstypicallyusedasdigestiveandcardiotonic.
TheArishta(fermenteddecoction)andAsava(fermented
infusion)are considered as a unique and valuable
therapeuticsinAyurveda,duetotheirmedicinalvalue,
sweettasteandeasyavailability.
Asava–Arishta formulations are better known ashydroalcoholicformulations,self-generatedalcoholduringthis
biomedicalfermentationprocesscanreachinbetween4
and12%v/v,andduetoitsimprovedaromaticproperties
andpalatably these formulations have better patient
acceptability.Thesetraditionallyfermentedformulations
haveseveral merits over other Ayurvedic medicines,
likeduetoself-generatedalcoholhasbetterpreservation
quality.
Preparationof Asava is carried out by Hima (Cold
decoction)/Jala, Swarasa (expressed juice) process. In
thepreparationofAsava the drug is coarsely powdered
and added to water, to which the prescribed quantities
ofhoney, jaggery/sugar are added. It contains dilute
solutionsof the readily soluble constituents of crude
drugs. Arishta is prepared by soaking the drugs in water
fora denite period of time (8 hrs)before decoction,
whichfacilitatesthebetterextractionofactiveprinciples
into Kasaya (decoction) and thereby increase in potency.
Itinvolvesboilingofcrudedrugsinaspeciedvolumeof
waterforadenedtimeperiod.Then,itisallowedtocool
andlter.Thisdecoctioniscalled“Quath” or “Kwatha”.
Forkwathapreparation,thepercentageofwaterdepends
uponthenature(hardness)andquantityofthedrug.The
ratioof crude drug towater is generally1 : 4 or1 : 16.
Duringboiling,thevolumeisreducedtoone-fourthofits
originalvolume.Then,theconcentratedextractisltered
andusedassuchorprocessedfurther.Kasayaistransferred
to Sandhana patra and sugar, jaggery or honey are added
toitfollowedbyfermentativeagentandPrakshepa Dravya.
Benefits of Asava–Arishta formulations
1. Self-generated alcoholand hydrolysing enzymes
releasedby yeasts aids extraction of phytochemicals
fromrawherbals.
2.Self-generated alcohol gives pleasing organoleptic
properties and dramatically improves palatability of
the preparation.
3. Palatabilityimprovement ensures patient compliance
and hence clinical outcome
4. Alcohol is considered as “Yogvahi”meaningfacilitate
fasterandevendistributionofdruginbody.
5. Self-generatedalcoholimprovesbioavailabilityofdrug
molecules.
Section 2 Pharmacognosy
6. Self-generated alcohol actsas a preservative, Asava–
310
Arishtapreparationlastforseveralyears,withoutspecial
storage conditions.
7. Preservationbenetsavailabilityofherbsindependent
ofseasonalavailabilityofthebotanicaldrugs.
8. Microbialtransformationmayreducetoxicityofphyto
ingredients.
9. Microbialtransformationincreaseschemicaldiversity
intheformulation,whichiscrucialforitsclinicalaction
overtherawdrug.
10.Biochemicaltransformationmakesthedrugavailable
initsmetabolisedor activated form,whichtranslates
into quick action.
Examples: Dasamoolaarishtam, Amruthaarishtam,
Kutajaarishtam, Draakshaarishtam, Abhayaarishtam,
Balarishtam, Devadaarvaarishtam, Asokaarishtam,
Jeerakaarishtam, Khadirashtam, Karpoorasavam, Pippalyaaadi
Asavam, Punarnavaasavam, Usiraashtam Aravindaarishtam,
and Patrangaasavam.
Avaleha or Lehya: Itisasemi-solidpreparationofdrugs
preparedbythe additionofjaggeryorsugarcandyand
boiled with prescribed drug juices decoction, Honey,
ifrequired, is added when the preparation iscold and
mixed well. Ghee or oil is added when preparation is hot.
TheLehyashouldneitherbehardnorathickuid.When
pulpofthedrugsisaddedandgheeoroilispresentinthe
preparation,thiscanberolledbetweenthengers.
Thesepreparationsgenerallyhave:
1. Kasaya or other liquids.
2. Madhur Dravya (sweetening agents) like jaggery, sugar,
sugar candy and honey.
3. Prakshepa Dravyas (additives)theseareherbalsadded
to the Avaleha to increase its potency, palatability or
toimprove organoleptic properties. There aretwo
typesofPrakshepa Dravya—Kasthoushadhis (herbal) and
Rasoushadhis (metallic).
3.Powdersorpulpsofcertaindrugs.
4. Sneh Dravyas (fats)likeghee,oil.
Method of Preparation
1.Jaggery,sugarorsugarcandyisdissolvedintheliquid
andstrainedtoremovetheforeignparticlesandboiled
overamoderatere.
2.Itshouldberemovedfromre,whenpressedbetween
ngersitbecomesthready(Tantuvat), or when it sinks
inwaterwithoutgettingeasilydissolved.
3. Powdered drugs are then added in small quantities and
stirredcontinuouslytoformahomogenousmixture.
4. Ghee or oil is added and mixed well.
5. Finally, honey is added when the preparation becomes
cool and mixed well.
Storage: The Lehya should be kept in glass or porcelain
jars. It can also be kept in a metal container which does not
react with it. Normally, the Lehya should be used within
one year.
Examples: Dashamulaharitaki, Bilvadileha, Vasavaleha, Citraka
Haritaki, Chyavanaprasa, Kusmandaka Rasayana, Vyaghri
Haritaki, Kalyanaka Guda, Ashvagandhadi Lehya, etc.

Tailas: Thesearealsoknownasmedicatedoilsforminga
groupofdrugsinayurvedicsystemofmedicinewiththe
principle is to extract the therapeutic compounds into oil.
Preparation:Themethodofpreparationrequiresheating
ofoil with prescribed Kashayas (decoction)and Kalkas
(powdereddrugs)accordingtoformula.Theyaregenerally
usedforAbhyanga (external application).
Itconsistsof:
1. Drava (any liquid medium as prescribed in the
composition)
2. Kalka(Finepasteofthespecieddrug)
3. Sneha Dravya (Tailas)
4. Gandha Dravya(Perfumingagents).
The medicated Tailaswillhavetheodour,colourandtaste
ofthedrugsusedintheprocess.Ifaconsiderableamountof
milk is used in the preparation, the Tailas will become thick
andmaysolidifyincoldseasons.Tailasarepreservedin
goodqualityofglass,steelorpolythenecontainers.These
medicatedpreparationsretainthetherapeuticefcacyfor
16th months.
Churnas
ItisdenedasanepowderofdrugordrugsinAyurvedic
systemofmedicine.DrugsmentionedinPatha, are cleaned
properly,driedthoroughly, pulverisedandthensieved.
The Churna isfree owing and retains itspotency for
oneyear,if preservedinairtightcontainers.TheChurna
consistingofnepowderofherbsinappropriateratiowas
subjectedtostandardisationbymeansofvariousphysical,
chemical and microbiological methods.
Storage: The packed materials should be stored in cool,
dry and dark conditions.
Standardisation of Churnas
Itgenerallyinvolvesthefollowingparameters:
1.Determinationofsievesize
2.Lossondrying/moisturecontent
3.TLC
4. Total ash
5.Acid-insolubleash
6.Water-solubleash
7.Extractivevalueinwater,alcoholandothersolvents
8. Phytoconstituents
9. Microbial contaminations
10.Heavymetallimittestformercury,arsenic,cadmium,
and lead
11. Microscopic analysis
Lepas
Lepas are semi-solid preparations intended for external
applicationtotheskinorcertainmucousmembranesfor
emollient,protective,therapeuticorprophylacticpurposes
whereadegreeofocclusionisdesired.Theyusuallyconsist
ofsolutionsordispersions ofoneormoremedicaments
in suitable bases. The base should not produce irritation
orsensitisation of theskin, nor shouldit retard wound
healing; it should be smooth, inert, odourless, physically
and chemically stable and compatible with the skin and
withincorporatedmedicaments.Theproportionsof the
base ingredients should be such that the ointment is not
toosoftortoohard for convenient use.Theconsistency
shouldbesuchthattheointmentspreadsandsoftenswhen
stress is applied.
Standardisation of Lehyas
•Lossofdrying
•Ashvalues
•Extractivevalues
• pH
• Thin layer chromatography.
Ghrita
These are preparations in which ghee is boiled with
prescribed Kasayas (decoction) and kalkas of drugs
accordingtoformulationasperAyurvedicformulary.
Goggulu
Ayurvedicmedicines prepared by the exudates, and
obtainedfromtheplantCommiphora mukul, are known as
Goggulu.TherearevedifferentvarietiesofGogguluin
AyurvedicShastrabutusuallytwovarieties,Mahiskasa and
Kanakaarepreferredformedicinalpreparation.Exudates
insmallpiecesaretakeninapieceofclothandboiledin
Gomutara or Dugdha or Triphala Kasayua until the exudates
passinto the uid through thecloth to the maximum.
Theuidafterlteringisboiledtillitformsamass.After
drying,themassisformedintoapastebyaddinggheetill
it becomes waxy.
Rasa Rasayan
Ayurvedicmedicinescontainingmineraldrugsas main
ingredients are called Rasa Rasayan or Ras-Yoga. They are
inpill formorinpowderform/forest,minerals,suchas
Anrala, Swarna, Rajata, Tamra, etc.andsulphurimpuried
stateareusedtoconvertBhasmaform,calledKajuali then
other drugs are added in small quantities, mixed well and
groundedtoformnepowder.
Vati or Gutika
Medicinesprepared in the form of tablets orpills are
known as Vati or Gutika,thesearemadeofoneor more
drugsofplant,animalormineralorigin.
Bhasmas
The Bhasmas are the powder of substances obtainedby
calcinations.ThepreparationofBhasmasincludefollowing
stages:
1. Purva Karma: Sodhana(purication)
2. Pardhana Karma: Marana(incineration/calcination)
3. Paschat Karma: Amritikarna, Lohitikarna
Sodhana (Purification)
Itispreparedfrompuriedminerals,metals,marineand
animalproducts. The following changes are observed:
Afterpurification, the materials become free from
impuritiesandbecomeneaswell.
Second Stage (Marana)
Thepurieddrugis mixed withKasayaofdrugs,andit
isgroundwith motar and pestle forspeciedperiodof
time. The cakes are prepared and their size and thickness
dependonheavinessofthedrug.Thecakesaredriedin
the sunlight and placed in Sarava (shallow earthen plate)
Section 2 Pharmacognosy
311

and closed with other plate and sealed with clay smeared
cloth and dried. The pit is dug in open space and sealed
platesandcowdungarelledinthepit.Fireisputonall
thesidesandwhenburningisover,allowedtocool.The
earthenpotisremovedandsealis openedandcontents
aretakenout.Themedicineis ground into ne powder
in a Khalva(motarandpestle).Theprocessoftriturating
isrepeatedasmanytimesforpropernessandquality.
EVALUATION CRITERIA OF PLANT EXTRACT
1. Description
2. Colour
3. Odour
4. Taste(ifnecessary)
5. Lossondryingat105°C/Moisturecontent
6. Total ash
7. Acid-insolubleash
8. Water-solubleextractives
9. Alcohol-solubleextractives
10. pH
11. Bulk density
12. Tap density
13. Volatileoilcontent(ifpresent)
14. Residualsolvent
15. Chemicaltestforsecondarymetabolites,e.g.alkaloids,
terpenoids,avonoids,glycosides,soponins,etc.
16. TLC/HPTLC/HPLCwithmarker(s).
17. Testforheavy/toxicmetals(lead,cadmium,mercury,
arsenic)(limitsasperASUPharmacopoeia)
18. Pesticide residue organochlorine pesticides,
organophosphorus pesticides, pyrethroids (limits as
perASUPharmacopoeia)
19. Microbial contamination
Totalviableaerobiccount
Enterobacteriaceae
Totalfungalcount
(LimitsasperASUPharmacopoeia)
20. Testforspecicpathogen
Escherichia coli, Salmonella spp., Staphyloccocus aureus,
Pseudomonas
Aeruginosa
(LimitsasperASUPharmacopoeia)
21. Aatoxins(B
1 B2 G1 G2
)
(LimitsasperASUPharmacopoeia)
22. Shelf-life.
19. Plant Tissue Culture
INTRODUCTION
Planttissueculture(PTC)isappliedforasepticcultureof
cells, tissues, organs and their components under suitable
in vitro physical and chemical conditions using nutrient
media(liquid/semi-solid)forproductionofprimaryand
secondarymetabolitesorforplantregeneration.
Basic tissue culture production
Plant cell culture is based on two concepts plasticity and
totipotency. The plasticity allows plants to alter their
metabolism,growth,anddevelopmenttobestsuittheir
environment.
Section 2 Pharmacognosy
312

HISTORYOFPLANTTISSUECULTURE
1902 The German botanist Gottlieb Haberlandt regarded as the father of plant tissue proposed concept of in vitro
cell culture.
1904 Hannig cultured embryos from several cruciferous species.
1922 Root and stem tips were successfully cultured by Kolte and Robbins.
1926 Went discovered first plant growth promoting hormone—Indole acetic acid (IAA)
1934 White suggested vitamin B as growth supplement in tissue culture media for tomato root tip.
1939 Gautheret, White and Nobecourt established endless proliferation of callus cultures
1941 Overbeek was first to add coconut milk for cell division in Datura.
1946 Ball raised whole plants of Lupinus by shoot tip culture.
1954 Muir was first to break callus tissues into single cells.
1955 Skoog and Miller discovered kinetin as cell division hormone.
1957 Skoog and Miller gave concept of hormonal control (auxin and cytokinin) of organ.
1959 Reinert and Steward regenerated embryos from callus clumps and cell suspension of Daucus carota.
1960 Cocking was first to isolate protoplast by enzymatic degradation of cell wall.
1962 Murashige and Skoog developed MS medium with higher salt concentration.
1962 Kanta and Maheshwari developed test tube fertilisation technique.
1966 Steward demonstrated totipotency by regenerating carrot plants from single cells.
1970 Smith and Nathans discovered first restriction enzyme from Haemophilus influenza (Hind III)
1970 Baltimore isolated reverse transcriptase from RNA tumour virus
1972 Carlson produced first interspecific hybrid of Nicotiana by protoplast fusion.
1972 Berg produced first recombinant DNA, combining SV/40 virus and λ virus
1974 Reinhard introduced biotransformation in plant tissue cultures.
1977 Chilton, et al. successfully integrated Ti plasmid DNA from Agrobacterium tumefaciens in plants.
1978 Melchers et al. carried out somatic hybridization of tomato and potato resulting in pomato.
1981 Larkin and Scowcroft introduced the term somaclonal variation.
1983 Pelletier, et al. conducted intergeneric cytoplasmic hybridisation in radish and grape.
1984 Horsh, et al. developed transgenic tobacco by transformation with Agrobacterium.
1987 Klien et al. developed biolistic gene transfer method for plant transformation.
2005 Rice genome sequenced under International Rice Genome Sequencing Project.
Basic Requirements for Tissue Culture Laboratories
A laboratory, in which tissue culture techniques are
performedusuallyincludesthefollowingfacilities:
1. Equipment and apparatus
2. Washing and storage area
3. Media preparation area
4. Sterilisation area
S. No. Equipment Description
1. Spirit burner For flame sterilisation of instruments
2. Refrigerator/ freezer To store temperature sensitive chemicals as well as stock solutions
3. Balances To weight various chemicals needed for preparation of chemicals
4. Hot plate cum magnetic stirrer To make media and for dissolution of nutrients
5. pH meter To adjust pH of the medium
6. Autoclave Moist heat sterilisation is needed for sterilise media, water and glasswares.
7. Hot-air oven For sterilisation of the equipment
8. Shaker To maintain cell suspension culture
5. Environmentallycontrolledincubatorsorculturerooms
fullyequippedwithcontroldevices.
6. Observation/data collection areaequipped with
computers.
1. Equipment and Apparatus
Thevariety of equipment essential fortissue culture
laboratories are:
Contd...
Section 2 Pharmacognosy
313

S. No. Equipment Description
9. Laminar air flow hood For aseptic transfer of explants to media and for subculturing
10. BOD incubators To maintain constant temperature to facilitate callus culture and maintenance.
11. Centrifuge To sediment cells and clean protoplasts
12. Glasswares and plastic wares Erlenmeyer’s volumetric flask, petridish, measuring cylinder, pipettes,
micropipettes and funnels
General Procedures involved in Plant Tissue Culture
In vitroculturing of plant tissue involves the following
steps:
1. Sterilisationofglasswaresandequipment
2. Preparationandsterilisationofexplants
3. Productionofcallusfromtheexplants
4. Proliferationofculturedcallus
5. Subculturingofcallus
6. Suspension culture.
Sterilization of Glasswares and Equipment
Heat sterilisation is the most widely used and reliable
methodofsterilisation,involvingdestructionofenzymes
and other essential cell constituents.
Preparation of explants
Explants:Cell,tissueororganofaplantthatisusedto
start in vitrocultures.Thedifferentexplantscanbeused
fortissueculture,commonlyusedareauxiliarybudsand
meristems.
Production of callus from the explants
Thecallus,growingmassofcellswhichisdedifferentiation
ofexplants:
1. The unorganised sterilized explants are transferred
asepticallyontodenedmedium.
2. TheaskistransferredtoBODincubatorformaintenance
oftheculture.
3. Temperature(25±20°C)andlightisneeded(necessary
forcallus).
4. Sufcientamountofcallusisproducedwithin3–8days
ofincubation.
Proliferation of Callus
Theculturedevelops,cutintosmallpiecesandintroduced
intothe fresh medium consisting ofthe hormones and
nutrientswhich support growthand proliferation of
explants.Proliferationmediumisthemediuminwhich
moreamountofcallusisproduced.
Subculturing of callus
Theperiodictransferofcellsto thefreshmediumatthe
intervalof4–6weeksshouldbedonetomaintainviability
ofthecells.
Suspension culture
Cellsuspensionculturetechniquesareveryimportantfor
plantbiotransformation andplant genetic engineering.
Whenfriablecallusisplacedintotheappropriate liquid
mediumandagitated,singlecellsand/orsmallclumpsof
cells are released into the medium and continue to grow
anddivide, producing a cell-suspension culture.The
inoculum used to initiate cell suspension culture should
neitherbetoosmallto affect cellnumbersnortoolarge
toallowthebuild-upoftoxicproductsorstressedcellsto
lethallevels.
Culture Media
Themedium consists of macro-c andmicro-c elements,
sugar,vitamins, amino acids, growth regulatorsand
miscellaneous supplements likeyeast extract and fruit
juices.
S. No. Agents used in surface
sterilisation
1 Sodium hypochlorite Laundry bleach is 5.25% sodium hypochlorite.
2 Ethanol (or isopropyl
alcohol)
3 Calcium hypochlorite Calcium hypochlorite may be less injurious to plant tissues than sodium hypochlorite.
4 Mercuric chloride It is extremely toxic to both plants and humans. It is critical that many rinses be used
5 Hydrogen peroxide The concentration of hydrogen peroxide used for surface sterilisation of plant material
Used in percentage (%)
70% ethanol is used prior to treatment with other compounds
The concentration that is generally used is 3.25%.
to remove all traces of the mineral from the plant material.
is 30%
Section 2 Pharmacognosy
314

Composition of media used in plant tissue culture
Media Composition
S. No. Nutrients Composition Functions
1 Macronutrients
Nitrogen (N),
Potassium (K),
Phosphorus (P),
Calcium (Ca),
25–60 mM
20–30 mM
1-3–mM
Required for plant cell or tissue growth
Magnesium (Mg),and
Sulphur (S)
2 Micronutrients
Iron (Fe)
Manganese (Mn)
Zinc (Zn)
Boron (B)
Copper (Cu)
Molybdenum (Mo)
1 µM
20–90 µM
5–30 µM
25–100 µM
0.1 µM
1 µM
Micronutrients have role in the functioning of the genetic
apparatus and are involved with the activity of growth
substances
3 Carbon and Energy
Source
Sucrose
2–3% For supplying carbohydrate needed for CO
during photosynthesis
Glucose
Fructose
Lactose,
Galactose,
Rafinose,
Maltose,
Starch
assimilation
2
Section 2 Pharmacognosy
Contd...
315

S. No. Nutrients Composition Functions
4 Vitamins
Thiamin (B
pyridoxine (B
inositol
), nicotinic acid,
1
), and myo-
6
0.1–10 mg/Liter
0.1–5.0 mg/Liter
0.1–10 mg/Liter
50-5000 mg/Liter
Vitamins are added in small amounts for growth and survival.
5 Amino acids and
Nitrogen
L-glutamine
L-asparagine
L-tyrosine
6 Solidifying and Gelling
Agents
Agar 0.5 and 1.0%
7 Support System
Perforated cellophane
Filter paper bridges
Filter paper wicks
Polyurethane foam
Polyester fleece
8 Growth Regulators
Auxins,
Cytokinins,
Gibberellins
Abscisic acid
Type of Plant Tissue Culture:
Amino acids are added in small amounts for growth
up to 8 mM
100 mg/liter
100 mg/liter
Plant tissue and organs are most suitably retained above
the surface of culture medium by increasing its viscosity with
some kind of gelling agent
Their selection and the concentrations depend on the plant
species and the purpose of the culture
CALLUS CULTURE
Step1:Tissuesamplefromanyregionofplantiscultured
Step2:Undenedcallusculture
Section 2 Pharmacognosy
Step3:Undenedcallusculturegrows
Step 4: Single cell culture
316
Step 5: Culturing results in new plant
SUSPENSION CULTURE
Thesuspensioncultureisthemethodofculturingisolated
cellinliquidmediumviatwomethods—mechanicaland
enzymatic digestions. Suspension culture is maintained
undercontrolled condition of light temperature and
aerationtofollowthepredicablepatternorgrowthcurves.

Scheme of the procedure of initiation and maintenance of plant cell suspension cultures
TYPES OF SUSPENSION CULTURE
Section 2 Pharmacognosy
317

Batch Culture
They are systems in which cells grow
in a fixed volume of culture medium
Incubation: There is a gradual
increase in cellular mass. Growth
ceases due to depletion of nutrients
from the medium
Growth Curve: Sigmoid pattern of
growth over a short period of time
Continuous culture:
Cell suspension culture can be grown
in continuous culture. It involves a
continuous influx of fresh medium and
the withdrawal of the used medium
without any cell loss
Open continuous culture system:
The cell mass as well as the culture
medium are changed periodically.
Replacement is automated, i.e.
addition of fresh medium occurs at
a rate equaling the rate of new cell
formation
Closed continuous culture
system: This is a process in which
cell mass remains unchanged. Only
the composition of the medium is
continuously changed
Chemostat: It is the constant supply
of chemical components/nutrients to
get desired rate of growth
Turbidostat: It is the constant
environment maintenance by input
of medium, which is checked by
measurement of turbidity of cell mass/
density
Applications of Suspension culture (SC)
1. Todevelopspeciccelllineorsinglecellclone.
2. Tostudyeffectofdifferentchemicals.
3. ForsomaticembryogenesisfromSC.
segments maintaining characteristicof organ in aseptic
culture media.
Organcultureisusedprincipallyfor:
1. Themaintenanceofstructuralorganizationintissues
4. Forimmobilizedcellculturesforsecondarymetabolites
orforbiotransformationofchemicalcompounds.
5. Forcommercialproductionofcellmass.
Organ Culture
2. The study of morphogenesis, differentiation, and
3. Forcomparisonofthegrowthandbehaviourofexplanted
Organcultureisthegrowthofplantorganssuchasroots
and shoots, beginning with the organ primordial or
S. No Type of organ
culture
1 Meristem
Culture (apical
tip culture/
shoot tip
culture)
2. Excised root
culture
3. Leaf culture This technique includes in vivo culturing of
4. Flower and
floral meristem
culture
Section 2 Pharmacognosy
Description Applications
Meristem culture is a method in which shoot
apices with a few primordial leaves are
grown in vitro
Root cultures can be established in
vitro from explants of the root tip of both
primary and lateral roots. Root culture
requires exogenous auxins in appropriate
concentration as supplements to the
endogenous levels for their growth
immature/young leaves to obtain leaf discs
Dicot plants: Leaf discs, fragments, leaf
bases and even mesophyll protoplasts
easily show regeneration of roots, shoots or
somatic embryogenesis via callus
Monocots plants: No such potential found
Reproductive parts like flower are in vitro
cultured by this method. This is generally
concerned with two aspects: (a) Induction
of flowering in sterile culture of vegetative
parts and (b) culture of floral buds and floral
organs
whichare to be subjected toexperimentally-varied
environments(e.g.,tohormones,drugs,orradiation).
functioninexcisedorgansorpresumptiveorgans.and
organswiththegrowthandbehaviourofsimilarorgansin
situ.
For maintaining virus-free plants, because
the apical tips of shoots and roots are
generally free from viruses and other
micro-organisms, e.g. sugarcane, potatoes,
banana, garlic, rhubarb, dahlias, strawberry,
orchids, pineapple, and gooseberry.
Cell division
Cell enlargement
Cellular differentiation
Shoot regeneration
Flower and floral meristem cultures are
found in cauliflower, date palm, onion, sugar
beet
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