SUKRASNO SITI KUSMARDIYANI

-- explain --

Tekelan, Epatorium reptansKumis kucing, Orthosiphon stamineus

Authenticity ---explain

KUSMARDIYANI

BOTANIC IDENTITY TEST Morphological, Microscopical, Microchemical,

and Genetic Methods

© SK

“SIMPLISIA”

Ver. 22SK005

CRUDE DRUGS

DriedIntactSliced

Morphological Method

THE ORGAN COMPOSING CRUDE DRUGS:– Leaves and tops (herbs)– Barks– Woods– Leaves or leaflets– Fruits– Seeds– Subterranean Organs

– Aerial stem• Dimension,

• shape,

• colour,

• herbaceous or woody,

• up right or creeping,

• smooth or ridged,

• Hairs

• Position and arrangement of leaves– Radical or cauline

– Adnation

– Alternate

– Opposite

– Decussate

– Whorled

Morphological Method

LEAVES AND TOPS

Alternate = Placed singly along the stem or axis, not opposite or whorled.Distichous = Arranged in two vertical ranks.Opposite leaves = With two leaves at a node, one on each side of the stem or axis.Deccusate = In opposite pairs, each pair at right angles to the next.

LEAF ARRANGEMENTS

ALTERNATE ALTERNATE, DISTICHOUS

OPPOSITE OPPOSITE, DECUSSATE

Get more types of leaf arrangements and give examples

Cambridge Glossary of Botanical Terms

Wallis, 573.

TERMS FOR LEAF DESCRIPTION

• Origin of preparation– Trunk, branches, roots.– Whole, inner bark

• Inner surface– Colour, striations, furrows

• Size and shape– Lichens, mosses, lenticels, cracks or furrows,

colour before and after scrapping.• Fracture

– Short, fibrous, splintery, granular• Transverse surface

– Smoothed transverse surface after staining with fluoroglucinol and HCl

Morphological Method

BARKS / CORTEX

© SK

Cinnamomi cortex

Litseae cortex

Cinchonae cortex

Alstoniae cortex

• Size and colour– after and before staining

• Relative density– Guaiacum 1.33 and Poplar 0.38

• Hardness and behavior when split• Transverse surface

– Distribution of wood fibers and wood parenchyma, annual ring (true or false)– Distance between medullary rays and annual rings

• Longitudinal surfaces– Height of medullary rays.

Morphological Method

WOODS

Ligustrinae lignum

Sappan lignum

• A: primary structure• B: development of a

complete cambial ring• C: beginning of secondary

growth• D: stem after a number of

seasons of growth

• Types of vascular bundle• E: collateral• F: bicollateral• G: amphivasal• H: amphicribal

Trease, 517.Stem structure of dicotyledons

STEM

• Duration – Deciduous or evergreen

• Leave base– Stipulate or exstipulate

• Petiole– Petiolate or sessile– Shape, colour, hair

• Lamina– Composition, incision, shape, venation,

margin, apex, base, surface, texture

Morphological MethodLeaves or Leaflets

Morphology of leavesA. Shape

1. acicular 2. elliptical 3. oval 4. oblong 5. round

6. linear 7. lanceolate 8. ovate

9. obovate 10. subulate 11. spatulate 12. diamond shape 13. cuneate

14. cordate 15. auriculate 16. lyrate 17. reniform

Trease, 519.

Morphology of leavesB. Composition and Incision

1. pinnatifid 2.pinnatripartie 3. pinnatisect, 4.palmatifid 5. imparipinnate

Trease, 519.

Morphology of leaves

D. Margin

E. Base

C. Apex

Trease, 519.

• Type of inflorescences– Racemose, cymose or mixed

• Axis or receptacle of influorescence• Type of flower• Receptacle of the flower• Calyx• Corolla (number of petals, venation, oil glands• Androecium• Gynaecium

Morphology of Flowers

Inflorescences

Actinomorphic Radially symmetrical, a line draw through the middle of the structure along any plane will produce a mirror image on either side

ZygomorphicBilaterally symmetrical, a line draw through the middle of the structure along only one plane will produce a mirror image on either side

Irregular Bilaterally symmetrical, a flower in which all parts are not similar in size and arragemnent on the receptacle

FLOWER TYPES

INFLORESCENCE

Trease, 521.

PERIANTH FORMS

Plant Identification Terminology, 170-171.

PERIANTH FORMS

Plant Identification Terminology, 170-171.

• Classification:– Simple, Aggregate, Collective

• Simple, dry, indehiscent fruits– Achene, nut, caryopis

• Simple, dry, dehiscent fruits– Legume, follicle, capsules

• Schizocarpic or splitting fruits

• Succulent fruits (druppe, berry)

Morphological MethodFRUITS

• Shape and dimensions– Adhesion

– Dehiscence

– Pericarp

– Placentation

– Seeds

Morphological Method

FRUITS

Foeniculi fructus

Coriandri fructus

Capsici fructusCardamomi fructus

© SKPiperis nigri fructus

Trease, 522.

FRUITSFruits of the Solanaceae:

A – B capsulesA ripe fruit of Datura stramoniumB pyxidia of Hyoscyamus niger

with upper fruit showing calyx partly removed

C berries of Atropa belladonaD berries of Capsicum sp

• Size, shape and colour

• Funicle

• Hilum and micropyle

• Seed coats

• Perisperm

• Endosperm

• Embryo

Morphological Method

SEEDS

Coffeae arabicae semen

Coffeae robustae semen

© SK

Myristicae semen

Piperis albi semen

• Morphological nature– Root, rhizome

• Condition – Fresh, dry, whole or sliced, peeled or un peeled

• Subaerial stems • Part of the crude drug or adulteration

• Subterranean stems• Size and shape• Direction of growth and branching• Surface characters• Fracture and texture• Transverse section

• Roots• Kind• Size and shape• Surface characters• Fracture and texture• Transverse section

Morphological MethodSUBTERRANEAN ORGANS

Languatis rhizoma

© SK

Calami rhizoma

Zingiberis rhizoma

Rauwolfiae radix

Curcumaedomesticae rhizoma

Fixed oils, fats, waxes, volatile oils, resins, oleoresins, oleo-gum-resins, balsam

Dried juices

Latices

Extracts

Physical state– Solid– Liquid

Odour and taste

Chemical tests

Unorganized Drugs

© SK

MicroscopicalAnalysis

Trease, 520.

Trease, 528.

Anomocytic Anisocytic Diacytic Paracytic Actinocytic Bidiacytic

Ver. 22SK005

Give description for each type of stomata.

BENTUK DAN KONFIGURASI STOMATA yang berlainan dapatdigunakan sebagai patokan analisis mikroskopik serbuk simplisia.

Stahl, 54

Tipe stomata khusus:

1. Tipe Ranunculaceae= tipe anomositik

2. Tipe Cruciferae= tipe aninositik

3. Tipe Caryophyllaceae= tipe diasitik

4. Tipe Rubiaceae= tipe parasitik

Ver. 22SK005

Trease, 529.

EPIDERMISA. Lower epidermis of Digitalis purpureaB. Lower epidermis of Hyoscyamus nigerC. Upper epidermis of Atropa belladonnaD. Lower epidermis of Cassia angustifoliaE. Lower epidermis of Rosmarinus officinalisF. Lower epidermis of Mentha piperitaG. Lower epidermis of Pilocarpus jaborandiH. Upper epidermis of Lobelia inflataI. Lower epidermis of Digitalis lanataJ. Lower epidermis of Erythroxylum coca

A

B and C

D

E and F

G

Type of stomata: anomocyticanisocytic

paracytic

actinocyticdiacytic

© SK

Microscopical Analysis

• Epidermal trichomes (hairs)Leaves, stems, flowers, fruits and seeds may have trichomes.

Trichomes– Non glandular (clothing)

• Unicellular, multicellular

– Glandular• Unicellular or multicellular stalk• Unicellular or multicellular head

EPIDERMAL TRICHOMES

Trease, 530.

A Papillae of lower epidermis of Coca leafB – G Unicellular hairsB Papillae epidermal cell with cystolith from leaf

of CannabisC Cystolith clothing hair from floral bract of

CannabisD Lobelia inflata leafE Senna leafF Lignified hair of AilanthusG ComfreyH Group of unicellular hairs from Hamamelis

leafI T-shaped hair of Artemisia absinthium

Trease, 531.

EPIDERMAL TRICHOMES

A – H Uniseriate clothing hairsI Multicellular branched hairJ Biseriate hair

A Datura metelB Datura stramoniumC Mentha piperitaD Thymus vulgarisE Plantago lanceolataF Hyoscyamus nigerG Digitalis purpureaH Xanthium strumariumI Verbascum thapsusJ Calendula officinalis

GLANDULAR HAIRS

Trease, 532.

A – B Atropa belladonnaC Datura stramoniumD Digitalis purpureaE Multicellular labiatae

glandular hairF Hyoscyamus nigerG – H Primula vulgarisI Digitalis luteaJ Cannabis sativaK Artemisia maritima

Phloem

A.sieve tubes with companion cells,

B and C longitudinal and transversal views of sieve tube,

D. sieve plate in winter condition,

E. radial longitudinal view of laticifers

A. tracheidB. fibre tracheidC. xylem fibreD. septate fibreE. annular F. spiral vesselG. scalariform vesselH. reticulate vesselI. vessel with round bordered

pits and simple perforation pits

J. hexagonal pitsK. vessel segmentL.M.N. bordered pitO.half bordered pitP. transverse section vessel

Xylem

Trease, 532.

Microscopical Analysis

• Secretory cells– Oil cells– Secretary cavities or sacs

• Schizogenous• Lysigenous• Schizolysigenous

– Vittae• Schizogenous oleoresin canals

– Latex• Cells or tissue containing fluid with milky appearance

SECRETORY CELLS AND DUCTS

Trease, 535.

Ergastic Cell Contents

• Microscopically, physically or chemically identified non-living particles.

– Carbohydrate, protein, fixed oils, fat, glycoside, volatile oil, gum, silica, etc.

– Crystal • Calcium oxalate• Calcium carbonate

ERGASTIC CELL CONTENT

Trease, 537.

Calcium oxalate crystalsA-D tetragonal systemE-I monoclinic systemA1-A3 rosette crystals of

tetragonal systemD a tetragonal prismE a monoclinic prismG raphidesH a single needle crystalI a sphaerocrystal

MICROCHEMICAL ANALYSISDistribution of tissue

– Phloroglucinol and HCl• Lignified cells: red

– Chlor-zinc-iodine solution • Cellulose walls: blue or violet• Lignified or suberized walls: yellow or brown• Starch grains: blue

Granules of wheat starch, stained with iodine, photographed through a light microscope - Wikipedia -

Cinchona bark, moistured with phloroglucinol followed by adding hydrochloric acid - Kusmardiyani-

Clearing, defatting and bleaching• Chloral hydrate

– Dissolve chlorophyll, protein, starch, resin, volatile oil, but not sodium oxalate

• Solution of potash– Dissolve aleurone, starch, protein, swelling cell walls.

• Ether-ethanol– Defatting

• NaOCl– Bleaching dark colour sections.

Disintegration and isolation of tissue• Potassium chlorate and nitric acid• Chromic acid and nitric acid or sulphuric acid• Solution of potash or soda

• Ethanol• Alkanna tinctures• Chloral hydrate and glycerin• Chloral hydrate with iodine• Clove oil (clearing agent for oily powder)• Chlor-zinc-iodine solution (Schultze’s solution)• Copper oxide, ammoniacal solution (swelling cellulose)• Corallin, alkaline solution (stain callose, gums and mucilages)• Ferric chloride (tannin)• Glycerin (mountant)• Iodine

Reagents

• Lactophenol (clearing)• Mercury-nitric acid (Millon’s Reagent) for protein containing material• Nitric acid (crude fibre)• Picric acid (aleurone and animal fibres)• Potassium cupri-tartrate (Fehling’s solution) for reducing sugars• Potassium iodobismuthate (precipitate alkaloid)• Potassium tetraiodomercurate (Mayer’s reagent) precipitate alkaloid• Ruthenium red (gums and mucilages)• Sodium carbonate (disintegrating fibres)• Sodium hypochlorite (clearing, defatting)• Sudan III (oils or suberized walls)• Sulphuric acid (charring, dissolve all but little action on suberin)

Reagents

• AFLP: Amplified Fragment Length Polymorphism

• SSR : Simple Sequence Repeat (microsatellite)

• Sequencing rDNA, ITS: Internal Transcribed Spacer and chloroplast genes

• RAPD: Random Amplified DNA Polymorphism

• RFLP : Restrictive Fragment Length Polymorphism

DNA Profiling

Method of Choice • Level of polymorphism

• Reliability

• Robustness

• RAPD– Easy to start but difficult to get reliable results

• RFLP– Obsolete, low resolution and high cost

• AFLP– Highly polymorphic, easier to start but difficult to data base

• SSR– Highly polymorphic, expensive to develop, used for intra-specific

analysis or closely related species.• SCAR

– Suitable for differentiation of targeted herbs.• Sequencing

– Good for differentiating plants from different families and genera, not polymorphic enough for intervening sequences.

Comparison among Methods

-- get more detail information --

Plant Nomenclature Taxonomy, In: Trease and Evans Pharmacognosy, 15th ed., p.10.

--read and discuss--

addendum

LABORATORY ACTIVITIES

PREPARATION OF STARCH- ISOLATION OF AMYLUM -

Read again and discuss related topics from course material of FA2205 General Pharmacognosy

A procedure for starch production was given in some detail in a Roman treatise by Cato in 184 BCE. Grain was steeped in water for ten days and then pressed. Fresh water was added. Mixing and filtration through linen cloth gave a slurry from which the starch was allowed to settle. It was washed with water and finally dried in the sun.

Raven et al., 2009, Starch – Chemistry and Technology, p.2.

STARCHHandbook of Pharmaceutical Excipients, 5th ed., 2006.

STARCHHandbook of Pharmaceutical Excipients, 5th ed., 2006.

Read and discuss:

Handbook of Pharmaceutical

Excipients, 5th ed., 2006,

p.725-726.

Handbook of Pharmaceutical Excipients, 5th ed., 2006, p. 730.

A procedure for starch production was given in some detail in a Roman treatise by Cato in 184 BCE. Grain was steeped in water for ten days and then pressed. Fresh water was added. Mixing and filtration through linen cloth gave a slurry from which the starch was allowed to settle. It was washed with water and finally dried in the sun.

Raven et al., 2009, Starch – Chemistry and Technology, p.2.

STARCH, PREGELATINIZEDHandbook of Pharmaceutical Excipients, 5th ed., 2006.

Handbook of Pharmaceutical Excipients, 5th ed., 2006.

Read and discuss:

TAPIOCA/CASSAVA STARCH PRODUCTION

Raven et al., 2009, Starch – Chemistry and Technology, p.548.

TAPIOCA/CASSAVA STARCH PRODUCTION

Raven et al., 2009, Starch – Chemistry and Technology, p.548.

TAPIOCA/CASSAVA STARCH PRODUCTION

rice starch

corn starch

Handbook of Pharmaceutical Excipients, 5th ed., 2006.

Compare:

Pregelatinized Starch

Handbook of Pharmaceutical Excipients, 5th ed., 2006.