ARY acoGNosy : AND PHY TOC, COURSE OUTCOMES hy re ie es (CE “resets ilbeable to: ou M acrical COUT r" ; ng carious chemical [ets Afier com ming. na iosb a schnigues if thedetermination of leaf constany, cul Mere Is opie tet ol — spade detectadulteration. apy of root bal C3 Performithe amicroscops © salty andpurity by pharmacopoeial standards, coral? rugs fortherr ae COS Evaluate th _ aaa [Course our Hee . Se 0) ~ ocrec ‘ periment |_—_*F 0uteg, Expt. Title of EXP cor fay Oye No. tagcat | Cot : | Coz PS ae 7 nical Tests: Tragacanth V . Analisis of wf Crude nude Drugs by Che Ay __and Acacia Tests: aera | — x ests: Agar, | > Analysis of Crude Drugs by Chemical 8 Vv PA ‘Starch. and Gelatin \ aad c : Honey and / 3 “analysis of Crude Drugs by Chemical Tests Honey y ieee Castor Oil aad ae 4 Determination of stomatal number and index V fae 5 Determination of vein islet number and vein termination y Po number | > : = | é etel ade ratio Ela Determination of palisade | V “A Determination of size of starch grains by using eye | ieee ee | piece micrometer V | Determination of size of calcium oxalate crystals a Determination of Fibre length and width | ayo eee sey 10 Determination of number of starch grains by eee Lycopodium spore method y rae | | iE He _ Determination of Ash value 12 ee — Determination of Extractive values of crude drugs 13 termina a } ¥ 2 _ Determination of moisture content of crude drugs | | ri Det re | | __Determmnation of swelling index of erude drug | | | Is _Peteminaton, sain | 1 10n of 0 jo o foaming inde of crude drug | | 1 __1__1—_Experiment No. 01 »\ Analysis of Crude Drugs: Tragacanth and Acacia) 1.Aim To analyse the given samples of Tragacanth and Acacia with the help of chemical tests oanaly 2, Practical significance cee Chemical test 1s one of the methods of analysis of crude drugs for their ma it 7 P are difficult determine their identity, quality and purity. Due to gummy nature of tragacanth and acacta 7 a aan to identify only with visual examination. Hence chemical tests should be carried out to identify an their tical, the students will be able to analyse the g sind Acacia by performing chemical ests. formed to iven samples of Tragacanth ality and purity. In this pra cal outcomes (PrO) Pe at -O | Practical outcomes | Mapped | BIL After completion of this practical, the students will be able to: \ co oy Pro 1 | Explain the significance of chemical tests in identifying the crude col BIL2 drugs. \ | ‘Analyse the given sample of Tragacanth and Acacia by performing | cor | BTL4 | chemical tests. \ \ \ Collaborate and communicate with fellow students. | cor | BTLS Follow cleanliness, safety and ethical practices in the laboratory. | col | BTLS | 4, Theory Chemical tests are required for the confirmation of identity of crude drugs. The phytoconstituents presentin crude drugs are identified with specific chemical tests, For example, in digitalis cardiac glycosides are identified with Keller Killiani test while anthraquinone glycosides are identified with Borntrger’s test. Sometimes chemical tests are used to distinguish different varieties of crude drugs. For example, Gambier fluorescence test is carried out to distinguish between pale and black catechu. For unorganized drugs, chemical test plays important role in evaluation of crude drugs. In this experiment, tragacanth contains water soluble portion, tragacanthin and water insoluble portion bassorin which is composed mainly of sugars and uronic acid units. Acacia consists of arabian, a complex. mixture of calcium, magnesium and potassium salts of arabic acid. The colour change with lead acetate test distinguishes the identification of tragacanth and acacia. A)Tragacanth Synonyms: Gum Tragacanth; Gummi Tragacantha, Gum Dragon. Biological source: The dried gummy exudation obtained from the incision of Astragalus gummifera Labill Family: Leguminosae Physical characters Colour: White to pale yellowish white Odour: Odourless Trinity Publishing House, SataraTT _Laboratory Manual of Pharmacognosy & Phytochemistry | Experiment No. 01 geneous wells rapidly and forms a thick homo} Taste: Mucilaginous Solubility: It is partly soluble in water and w ith water it s adhesive gelatinous mass. Chemical constituents luble which is termed as ‘ragacanthin’ and water insoluble which is known as qed into three types of constituents xylose and galacturonic acid. A ile a third type is believed to 1. Two vital fractions: Water so "bassorin’ 2. Itis composed mainly of sugars and uronic cid un s tragacanthic acid on nydrolys' nd arabinose after its and can be divi sis yields galactose, its hydrolysis W 3, The acidic constituent neutral polysaccharide affords galactose a be steroidal glycoside gto manage diarthoea- as a suspending Uses | Asademulcentin cough and cold preparations an 2. Asanemollientin cosmetics. Isifying agent ‘Along with acacla it is used nthe pills. nding and as an emul gas an excipient i 3, Asa thickening. suspe! agent 4, Mucilage of tragacanth is us sdasa binding agent? the tablets and als 5. Tragacanth powder Is used as an adhesive. 6. Itisalso used 1n lotions for external use andalsoin spermicidal jel er for ice cre 0,3" tio} rted to inhibit the growth of cancer cells in vitro and1 lies. n and also ins in vivo. 2 , auces. asa stabiliz am in 0.2 4 concentra 7 Itisalso used as been repo! umand Gum Arabic. usp. Acacia is the dried gummy exudation obtained from the stems and eacia. In India. 1015 found as dried gummy her African species of es of Acacta arabica wild 8. Tragacanth hi B) Acacia Synonyms Ac Biological source A branches of fcacia Snes exudation obtained from the guminosae acia gum. Indian Gi cording to the al (L.) Wild or ot stems and branch Family: Le toredin colour Characteristics Colour a) Tears These are white, pale-yellow and sometimes creamy-brown b) Powder: These are off white, pale-yellow or light-brown In appearance Odour: Odourless Taste. Bland and mucilagimous Shape and size alorovoid with diameter of about 2.5-3.0cm presence of cracks or fissures produced on the n nature and exposed either due to the ng. The fracture IS usually very brittle 1 Tears: Spheroids Appearance Tears ar during the e invariably opaque outer surface process of ripen surface appears to be glossy Solubility: [tis soluble in ater, insoluble in alcohol 15°o of moisture an Standard: It should contain not more than tannin, starch and dextrin. 5° of ash. Indian gum should not contin Chemical constituents plex mixture of calcium, magnes1um and potas: Acacia consists principally of arabian, which ts a com| Tanity Publishing House Sate fAatory Manual of Ph Laboratory armacognosy & Phytochemistry | = Experiment No. 01 salts of arabic acid. Arabic acid Sedat i sa beebed polysaccharide that yields L-arabinose, D-galactose, D- ydrolysis. 1, 3-Linked D-galactopyranose units form the tpackbone chain of the molecule and the term ; ; inal re: ‘Acacia contains 12-15% of w: sidues of the 1, 6-linked side chains are primarily uronic acids c ater and several occluded enz pectinases. The total ash content should binthelaeesors a Sept schcas|Gxidasss. Perse ea Uses 1. Asademulcent. 2. Asa vital pharmaceutical aid for emulsification and to serve as a thickening agent 3. Asa binding agent for tablets of example, cough lozenge: / HS S. 4, Gum acacia solution has consiste1 . sistency similar to blood and is adi i . : and is administered intravenously in hemodial Pete ei p sly inhemodialysis 5, In the manufacture of adhesives and ink, and asa binding medium for marbling colors. 5, Requirements Apparatus: Acacia and tragacanth: test tube, test tube holder, matchbox, beaker Chemicals : a) Tragacanth: 90% alcohol, potassium hydroxide, lead acetate, ferric chloride (10%), iodine b) Acacia: Lead sub acetate, lead acetate, Fehling solution A and B, fersie chloride sation, bors ruthenium red, iodine. . " 6. Requirements used Frogs 19, ‘Testtuhe, Testube holden. — fo Ales KOH, clead Aeetoti fehliing Scln # 2 B Le aa 7. Procedurt | 1) Chemical Tests for Tragacanth Sr. No. Test | Observation | 1 Tragacanth + solution of potash’ KOH — Boil | Canary yellow colour 2 | Tragacanth + lead acetate solution \ Deep yellow precipitate 3 Tragacanth + few drops of FeCI3 (10 %) | Reddish brown colour L 4 | Test solution + Iodine | Olive green colour is produced 11) Chemical Tests for Acacia Preparation of test solution 1g of powdered drug + sufficient quantity of water and shake /stir Use this test solution for the following tests: 7 [Se No. ' “Test rt | Observation 1 Fehling Test: Hydrolyze test solution with dil | Red coloured precipitate due to HCl and boil. Then add [mL Fehling solution A \ and B, again heat on boiling water bath. | presence of reducing sugars Lactose 03 Trinity Publishing House, Satara| Ferric Chloride Test: FeCl, s¢ solution. i fe. jorax Tes! | shake well a asmall guanuty “of, powde: 4 slide and observe under microscope 5 | Mount Test Aqueous te: 2ml “Test solution + st solution * lead 1 Aqueous test solution + Ht rin ruthenium _Labe cad | Heavy white borax > oratory Manual of F of Pharmacognosy & Phytoch ey Misty, Observation 7 |< er oreciniin> No blue/black colour precipitate gy ~ due absence of Tannins. precipitate Ta . ate Stiff translucent mass. No red coloured crystal. No crimson colour is produced | red on a slide and ¢ 6 | Test solution ~ Iodine 8. Observations 1) Tragacanth ‘ Test Observation Inference / 1 | Tragacanth + solution of potash yellow Colo Toragorcant h | | KOH = Boil fi 7 present [ { [2 | Tragacanth ~ lead acetate solution 4 ello w pp Ir Fragacanth | - - te [3 | Tragacanth + few drops of FeCl, Redd&b- b Tera ae ( ) Colpy presenk 4 | Test solution + lode Olen Freon Faagaegnth L 7 Colby Present Il) Acacia | SN | Test | Observation [ Inference Febling Test: Hydrolyze test solution a colo + pp BE | pea a cl 1 on boiling water bath | with dil. HCl and boil. Then add [mL | Fehling solution A and B, again heat te to sreducH [fg Sugars — Present ine blue / bl ack | Jeannine 2 | Ferric Chloride Test. 2mL Test | solution + 2 to 3 drops of FeCl solution Coley abse nk { | T Lead acetate Test Aqucous test | ae hr he. pp t | faa cl | me ce & solution + lead acetate. {| Borax Test: Aqueous test solution + borax + shake well. | Supe qanslu cot rae masse Paresen -esult he basis of chemical analysis, it was found that given unknown crude drugs are tragacorfan Cntaraee Laboratory Manual of Pharmacognosy & Phytochemistry | Experiment No. 01 10. Conclusion rom the chemical tests, ican be concluded that ragacanth and acacia were Sdentiti'e e) (identified and Cong tome qd: (confirmed/ notconfirmed) by using chemical test . 11. References | Pharmacognosy lab Manual, Iyengar MA, Nayak SGK, Pharma med Press, Hyderabad >. Practical Pharmacognosy, Wallis TE, 4th Edition, Pharma med Press, Hyderabad 3. Indian Pharmacopoeia 2020. 12. Synopsis questions 1. Give synonym and biological source of tragacanth. 2. Give the Fehling test for acacia. 3. Why chemical analysis of crude drugs are required? 4, Give the chemical constituents of acacia and tragacanth. 5, Whatare the uses of acacia and tragacanth? (Space for answers) de ANSWERS Syne hymn = Teaggacantha, Gury Tragacanth Bio. Source ~ Obtained frrorn deed gurormy exudation Keron the InchsTen of Astygulas Gyuronifiew dabiit: Q. Ay drelyse Soln with Hel 2 boil , then add + mt fehting Solution ,again heat en a twalte bath ,B Red 5 rs fotmed , 5 Chemfceal Analysts eoude ens vs used Eo detberr Delentity ualtty) putty, ahemPrcal 1K ane goqutved t Conforms the Fdenht eh crude drug. > The phyroc™ tb present Shy crude’ dug cue aletierntin | hy vassous 6) test: 0 HY. thers Constituents | Acadia & Arabic Had, calen Ragnacium , pettqssiurm Sat cy feces t Had oe Hao, Speymes (oaidaso, pestieoxro. ase, pee Hyases)\ 5 Trinity Publishing House, Satarais - nual of Pharmacognosy g __ Laboratory Manual of Phare mL E PH Yoh ag, Experiment No. 04 her ty, } Tt ' neblients oF weal Seluble) Bossorin( Tra gocanth Tragucen,| Lo heme Ca 9 ‘ioadar insoluble Prag ae, ¢ Glucoronie 5 Acid which yeilde gakactose, wylase, J Dey A nictvad polysace harvoles aPfords ge acloxre a Pres St! drolucu 7 ovabanese, aftes Pts hy on Me rks: Signature of teacher with date Trinity Publishing House, SataraLATIN Fae Experiment No. 02 aoe ANALYSIS OF CRUDE DRUGS: AGAR, STARCH AND GE 1.Aim Tanalyse AES Starch and Gelatin by performing chemical tests ‘panalYs {t is performed t0 heir evaluation. : difficult to identify only with visual confirm their quality and purity. In identification ofthe genuine species al significance one of the mi ity and purity. Agar, starch and ould be carried out to identify and ests In | ethods of analysis of crude drugs for 1 2. Practi¢ gelatin are Chemical test is determine their identity, qual examination. Hence chemical this practical, the students wil of Agate Starch and Gelatin I tests shi | lear the importance of chemical t 3. practical outcomes (PrO) i EEE CEE EEE EEE Eee 7 Practical outcomes ‘After completion of this practical, the sts in identifying the crude students will be able to: Explain the significance of chemical te drugs ‘Analyse the given sample of Agar, Starch and Gelatin by performing col BTL4 chemical tests. pro 3 | Collaborate and communicate with fellow students. col BTLS p14 | Follow cleanliness, safety and ethical practices in the laboratory. col | BTLS | 4, Theory Chemical tests are characters. Therefore, solubility, physical characteristics and study of unorganized drugs. Agar and starch contains carb teins. Due to presence of reducing sugars, \ds to those tests. Starch gives blue colot ives all tests for proteins (Million’s reagen| ry do not have microscopic important parameters for the hydrates particularly polysaccharides while agar responds to Molisch’s test and Fehling’s ur with iodine solution due to 1, Biuret required for identification of unorganized drugs as the’ chemical test are gelatin contains pro test while starch does not respon presence of complex polysaccharide, amylose. Gelatin g test, Xanthoproteic test and Ninhydrin test) positive. Al Agar Synonyms: Agar-agat, Vegetable Gelatin, Biological source: It is the dried aqueous Gelidium amansii (Lamouroux) or various species of red algae like Gracilari Japanese or Chinese Gelatin, Japanese Isinglass. extract or gelatinous substance obtained by extraction from ia and ferocladia. Family: Gelidiaceae Physical Characters Colour: Colourless Odour: Not distinct Taste: Mucilaginous Solubility: It is soluble in hot water but insoluble in cold water andalcohol Trinity Publishing House, SataraLaboratory Manual of Pharmacognosy & Phy, - che, Mis th Experiment No 02 Jerent polysaccharides known a two dif tis responsible for the gel property ofay saccharide and contans ble for the viscosity of agar solution olymer and respons! Cc hemical constituents Jex heteropel se isa neutral galactose P arose | : Pe si 4gar is a comp! je units. Agarapectin ist ropectin. Aga lactose and L-galacte Uses | Agar is used to treat chronie consupanen 2 Asa laxanve hing agent for suppositories. nt. an emulsifie 3. Asa suspendi tion and ointment and » AS dey, a) 4. As surgical lubricant ant. in production of medicinal encapsula| 5 As a tablet excipient, disintegra impression mould base sed asa gel in nutrient media for bacterial 6 Itisextensively u B) Starch Synonym Amy lu Biological soure cultures. e. It 1s the polysaccharide granules isolated from fully grown grains of Maize (Zeq Zea mq, Linn), Rice (Oriaza sativa Linn); and Wheat (Triticum aestivum Linn) belonging to Family Gramin eae any oI from the tubers of potato (Solanum tuberosum Linn) belonging to family Solanaceae. Characteristics Sr. No. Characters | Maize | Rice Wheat Potat. | otato : Col ur White | White Faint grey Yellowish tint | 2 Odour : Odourless | Odourless Odourless Odourl : a ou i s less | : Taste Mucilagenous | Mucilagenous Mucilagenous Mucilagenous Sha, Ss ins. i pe ealetanie Simple or Mostly Simple Flattened ovoid | ang hilum compound (large and small) | or subspherical, | | central, rarely grains (2-150 grains, faint well-marked ete grains | components) striations, striations, hilum ; imple or polyhedral Hilum appears eccentric. | compound with sharp as line. L grains. angles 5 | Size 5_3 i T : | Size 5-30um | 2-10 um Small: 10-100 um | 2-10 um Large: | | - | 10-45 um 6 HH al Pi / Neutral | Alkaline | Acidic Acidi Moisture | 13 (%v w) TB vw) 13% _ leomeat | [130% | 13 (“v/w) 20 (Yv/w) 8 Ash content | 0. vw) 3 (%ww) 0.6 (Ywiw ; | (Ywiw) | 0.3 (Yow/w) 0.3 (Yow/w) ch JS present in differe Fi ae eis nt parts of the plant in the form of granules of varying size. It is found abundant) . seed, root. me and as smaller grains in chl ig P. edtdieceite. : 's in chlorophyll containing tissue of the pl. as leat ° s ant such as leaf es of different origins can be identified by studying their size shape and structure, as well osition : s as well as, positio = 111—_ °° °° °° © acognosy & Phytochernistry | Expenment No. 02 ratory Manual of Pharm jopectin and B- Labo! of the hilum and striations Chemically, starches are polysaccharide containing amylopectin 38 amylose Starch turns blue to violet when treated with iodine solution. Starches of pharmaceut Se ae obtained from maize, TCE, wheat and potato. These starches can be differentiated from ca jnicroscopical examination — ‘A comparative account of their macroscoptcal, microscopical and phy sical characteristics 15 B1¥ env * mounted in Smiths starch reagent table 4.2. For purpose of microscopical studies, the powder should be containing equal parts of glycerin, water and 50% acetic acid Chemical constituents Chemically, starches are polysaccharides containing two Co} amylose. Amylopectin 1s insoluble in water; it has highly branc! hundred short chains of about 20-25 D-glucose units each. Amylose 1s wats yields disaccharide (+) maltose and monosaccharide D- (+) glucose. mylopectin and B- smposed of several hich on hydrolysis mplex polysaccharides. 3 shed structure which 1s cc er soluble w Uses 1. Inthe formulations of tab} 2.1In dusting powder as an absorbent and protective. 3, As antidote in iodine poisoning + Asa nutritional food in cereal based weaning foods e. 5. Topically and externally to alloy itching. 6. Asastarting material for the production of glucose, glucose syrup, dextrose and dext Jets and pills as a diluent as wellas disintegrating agent g. Farex and Cerelac. rin, 7, Insuppository as a base. c) Gelatin Synonyms: Gelatin, Gelatinum Biological source: Gelatin is a protein derivative obtained by hy evaporating aqueous extract of bones, tendons, ligaments, connec animals, like ox and sheep. Family: Bovidae Physical characters Colour: Pale yellow Odour: Characteristic Taste: Slight mucilaginous or gelatinous Solubility: It is soluble in hot water but insoluble in cold water and alcohol Chemical constituents , drolysis of collagen, and after purifying or tive tissues of mammals and skins of r ‘ cietdum tan which on hydrolysis gives a mixture of amino acids. The approximate ami wi : a lycine (25.5 %)s cas (8.7%), valine (2.5%), leucine (3.2%), isoleucine (1.4 ysteine and cysteine (0.1%), methionine (1.0%), tyrosine (0.5%), aspartic acid (6.6%), glutami (11.4%), arginine (8.1%), lysine (4.1%), and histidine (0.8%). ‘ “ve at enter, gelatin isan incomplete protein lacking tryptophan. The gelatinizing compound is k tee rin and the adhesive nature of gelatin is due to the presence of glutin. LIti . Itis used to prepare pastilles, pastes, suppositories, capsules, pill-coatings. 2. It is also used a: i Sa suspendin: a i i ili i 7 ig agent, tablet binder, coating agent, stabilizer, thickener and text.Laboratory Manual of Pharmacognosy g Phytoch, y _— othe, _—_—_—— prepare cultures and asa nutrient fi seriolog’ yon in bactero! i f : : = higused as abase for the manufacture of suppositorieg hich ts used eso 7 stin which is employed asa topical protectant, food products and bacteriologic Culture Meg Experiment No. 02 oe c a so used to inhibit cry stalliz h glycerin W c.it forms zine gel ascommercial 3 Itisa Glycerinated gelatin w ath + When combined with zin dasanutrient, gelatin isused, i 6 Itisalso use 5. Requirements et Apparatus Test tube. test ube holder, matchbox, beaker hemicals ‘| red, KOH, Molise al NaOH, barium chloride, ruthenium re olisch, Teagy, a) Agar Fehling soltuion and B. HCl, Cone. H.SO, N 2010dine e. c. i Jution Aand B, HCI. b) Starch: Potassium iodide. iodine, Fehling so : . ¢) Gelatin CuSO., HNO... NaOH, Millon's reagent, NaNO. , ninhydrin alcoholic solution, soda lime, Dion, acid. Tannic acid (10%), 6. Requirements used fect Tbe, Tost Tube Holder, naqoit, Batle, ton 7 u Np! Peek Reagent eae AEG SSE eee 7. Procedure 1) Chemical Tests for Agar Preparation of Stock solution: g of powdered drug + sufficient quantity of water, Use this test solution for the following tests, | Sr.No. Test Observation || Fehling Test: SmL Test solution + 0.5mL HCI > | A Red coloured precipitate due to bol ~ ImL Fehling A and B + ImL of NaOH + _| presence of reducing sugar. Again heat it on boiling water bath > | Barium Chlonide Test: Take 2mL Test solution + | White precipitate /turbidity of barium. 2 10 3mL of BaCl. solution, 3 A small quantity of powder + ruthenium red Particles acquire red or pink colour, | solution. Examine under microscope. | 4 | Testsolution = KOH > Warm | Canary yellow colour. i | 5 | Test solution -» Boil Jelly is formed on cooling. 6 | Test solution ~ \ 20 iodine Deep crimson to brown colour, | > ct 7 | Molisch Reagent Test: Test solution + Molisch | Purple red or violet Ting at interface. | Teagent ~ |-2 drops of Conc. H.SO, from the | |__| Side of test tube. | |..._y Manual of Pharmacognosy & Phytochemsty | borat mm Chemical ‘Tests for Starch gn.No| aa / Starch grains in water + A drop of Jodine Observ under the microscope Hfodine-potassium iodide reagent Test: Starch *, Todine-potassium iodide reagent. 0.5g of starch + 20mL water > boil it gently for 2-3 minutes and cool. f | Observation oa e 11g presence of Deep blue colour due to presence 0) amylose Intense blue black colour. ilage. | Formation of Starch mut ation as nO reduction takes ‘ ae aqueous solution of picric acid. | 2-3 minutes and co +g _ | Starch mucilage + Fehling’s reagent — Heat No red color: place. a EL [5 _| Sm starch mucilage + 0.5mL HCI ~ beat Red colour. (30min). After 30min. + NaOH + Febling's reagent > heat until the mixture is alkaline to litmus and warm for few min. perform the test. peer [ees up Chemical Tests for ‘Gelatin ‘ : HH sr.No.|"' Test Observation il Biuret test: 2 mL alkaline solution of gelatin + Red or violet colour. CuSO, + 3ml. NaOH. - 2 Xanthopoteic Test: Gelatin + HNO, +40% Yellow colour and gives orange colou NaOH drop to drop. when made alkaline. 3 Millon’s Test: Millon’s reagent + Gelatin White precipitate and changes to red solution +NaNO,. wa S heating. i 4 Ninhydrin Test: An alcoholic solution of ninhydrin| Red to violet colour. + aqueous solution of gelatin. Boil for 2 min. 5 Soda Lime Test: 1g of powdered drug in test Evolution of ammonia. tube + few crystals of soda lime. 6 ‘Asolution of gelatin (0.5 g) in water (10 mL) + White buff coloured precipitate P few drops of tannic acid (10%). i Picric Acid Test; Take ImL test solution + 2-3 mL | Yellow coloured precipitate. 8.Observations | ' 1) Agar SN Test Observation Inference 1 | Fehling Test: SmE. Test solution + 0.5mL HCI - boil + [mL Fehling A ‘and B+ 1mL of NaOH — Again heat it on boiling water bath Rud ppt Vresenes ob uctng Sugary Agay lal Trinity Publishing House, SataraManual of Pharmacogno: _ Laboratory Manua’ Oe POPESY & Phyto, a Test i Test aa ine > | Barium Chlonde Test: 2 mL Test | solution + 2 t0 3 mL of BaCl} tion 7 7 small quantity of powder + __ | 7 | Reol Oo” pink Nig, NY a Inference 7 (eos Agqay Preven lagen Foresen ts | A | ruthenium red solution. Examine par rele ‘ under microscope 4 Test solution ~ KOH — Warm Canauy{etlow - : ll eee Il) Starch : Observation Inference Deep blue Colov lodine > Observe under the Parcients > Demy Akarch present , | microscope lodine-potassium iodide reagent Test: | Starch - Iodine-potassium iodide Pluc- Black Colo Starch present reagent | 0.5g of starch + 20 mL water > boil it gently for 2-3 min, and cool. Mucihage Porm ory Storch present | "4 | Starch mucilage + Febling’s reagent | NO Reol Coloy as nw reduction Starch preesent | [5 | SmL starch mucilage + 0.5mL HCI — Heat WA Red Color | heat (30min). After 30min. + - NaOH - Fehling's reagent > Heat | | unt! the mixture is alkaline to litmus | and warm for few min. perform the | test | III) Gelatin | SN Test Observation Inference | eee - 1 Biuret test’ 2 mL alkaline solution ot] d C [ qelaton olOv | gelatin = CuSO, -3mL NaOH, | Re ° iJ Prosent anthopotcc Test Gelatin “HNO. |Yellow~ Orange | Jela bh | + 40°. NaOH drop to drop. | Coley | Pores ent [3 / Mills Test Millon’ reagent [0 hit ppt charges! gel ann Gelatin Solution + NaNO. bo Rect en harry | Preen t 4 | Ninhydrin Test: : : | qzlabn | Ninhydrin Test: An alcoholic solution] Red +0 violet | g fb of ninhydrin + aqueous solution of | | Y, + mesent * gelatin — Boil for 2 min. Color | eo SatSS -—-—=>3 Experiment No. 02 Manual of Pharmacognosy & Phytochemistry | - —— | "Inference | ervation om ret Observation =} Soda Lime Test: 1g of powdered | | | drug in test tube + few crystals of | a | soda lime. _ | A : ' adn ‘A solution of gelatin (0.5 g) in water | WPL up | ae . | (10 mL) + few drops of tannic acid ep ‘ Present | (10%). | were | Picric Acid Test: Take ImL test 4 elle vo per | Ger a | solution + 2-3 mL aqueous solution | © ent \ of picric acid. | | red 9. Result are ay On the basis ne analysis it was found that given unknown crude drugs are Ay 10. Conclusion ° fe Atidentified! From the chemical tests, it can be concluded that Agar, Starch and Gelatin were Fdenh e he _notidentified) and Condivmedd (confirmed/not confirmed) by using chemical tests. 11. References 1.Pharmacognosy lab Manual, M.A. lyengar & S.G.K . Nayak, Pharma med Press, Hyderabad. 2. Practical Pharmacognosy, T. E. Wallis, Fourth Edition, Pharma med Press, Hyderabad. 3. Indian Pharmacopoeia 2020. 12. Synopsis questions 1. Give the biological source of agar, starch and gelatin. 2. Write the chemical constituents ofagar, starch and gelatin. 3. What are the uses of gelatin and agar? 4. Enlist the chemical tests for agar and starch. 5. Explain Millon’s and Biuret tests. (Space for answers) oo, oO Ny de rb o7s the dased "AQ > exlevark Or extrack 7 Lyeladtum Prmanesye Oy vaniou elatan Obtatnod’ bu ext Species y real Algor Wee Gracia wa 2» } Starch 8 Tt 4s polysaccharide granules totale | 2 ak Yo. gains roarre, C22A mw wast, Rrea, ( ny Sathio Yo vsheert ( Tertius Beabyum oh etm forrity rami nea e@ Q foro rn tubors vi porato (Svlonuy lS,ual of Pharmacogni _ Laboratory Manual OEM CPRMSY & Phytogy, 7 Experiment No. 02 “emi, é o e pfs the proldin abtatined | fron, . 4 1 eS cjetahn e! evqporating ° vi fee, tla an) ot the hydevly sts of Col I] 5] 4 extrgck 7 bone: 2 tend, extracting toate soluble © _- tr 7 ‘ : # "| el Fyaments Conmectitie TFsg ut J vohrmrndl & ov OATR 7 primals fobs Oy 2 Sheep U C oN Ah A 7 © x7 Chemreal Cons Btuin ts ° r co a Agar— & “om plex lertetopelysace harvelt Contains tig ©. alate ; / hin. Agarose 1S oO ys haide: A Jaros, Agave) ec . / nuetyal giance polymes 2 PE t responarts te fev ge! Poeperty “cf Pgeseit Consiste oly D= galactose l= ga loc bs A gqarepec eh ¥° xesponstble for achvity oy eae Selube, tarech - Proytopectsn, B- Aruloce, Amylopectn rs tnselyuble im wate «Dn glose Pe evottue Solub te qelat nn It Contains pootetn glut fr which en hyolerolys: yeF lols QQ mex tee PDmitho Aciol « tyePre SS 5/2 Tanne C8726) YattneCals) duceine (B- 20/0) Aspartate f\, “6<(0),.glutomie Addd Cri ee) lysthrel4ere/e) , hfabidlf ng C+ Be/.\ > 3 & % used +o Prprue Ss ' pouttics, pastes, Suppostta n'a SiN Coa thn pes 7 ( used as binder, fuspenclity Agent, Coahing Agents Stabalvoeun, thichenstxy £ tex fe y— usec] 40 treqt Chronie As q darar tie y 2 b a asus endin ot 2 Hr fes A ellen Beaqen | Supportbetes 7" Agent emular pres, 9 q 9 Marks: Lo topes ID food! » Conshpgken Le (yecreee Signature of teacher with date:—_ Expe ent No. 03 | ») OR OIL ANALYSIS OF CRUDE DRUGS: HONEY AND CASTO) 1.Aim . ad fo analyse Honey and Castor oil by performing, chemical tests. Toa 2, practical significance - . aire 2 {tis performed to Chemical test is one of the methods of analysis of crude drugs for their ev ajuation f +o only with visual determine their identity, quality and purity. Honey and castor oil are difficult to. se va purity in amination. Hence chemical tests should be carried out to identify and confirm their q) > © fice fthe genuine species this practical, the students will learn the importance of chemical tests In identification © g ofhoney and castor oil. 3. Practical outcomes (PrO) tapped) BTL. PrO | Practical outcomes \ ar \ | ‘After completion of this practical, the students will be able to: \ \ = { A 2 TL . pro 1 | Analyse the given sample of Honey and Castor oil by performing \ col \ B \ chemical tests. | \ \ PprO 2 | Evaluate the given samples for quality and purity as per \ col \ BILS \ pharmacopoeial standards. \ \ 4 . r col \ BTLs | PrO 3 | Collaborate and communicate with fellow students. | | Pro 4 | Follow cleanliness, safety and ethical practices in the laboratory. \ col \ BTLS | 4,Theory Chemical tests are required for identification of unorganized drugs as they do not have microscopic characters. Therefore, solubility, physical characteristics and chemical test are important parameters f for the study of unorganized drugs. Honey contains reducing sugars and therefore responds to F iehe’s test and Molisch test positive. Fiehe’s test is used to distinguish pure and adulterated honey. Castor oil contains glyceride esters of higher fatty acids which gives a clear solution at a specific volume of organic solvent such as light petroleum ether. If the volume of light petroleum ether is increased thee fold, then the solution becomes turbid. A) Honey Synonyms: Madhu, Madh, Mel, Purified Honey Biological source: Honey is a viscid and sweet secretion stored in the honey comb by various species ot bees, suchas Apis mellifera, Apis dorsata, Apis florea, Apis indica and other species of Apis. Family: Apidae Physical characters: Colour: Pale yellow to reddish brown viscid Odour: Pleasant and characteristic Taste: Sweet, slightly acrid However, the odour and taste of honey depends upon the availability of flowers and storage, condition a , apeIS, | of Pharmacognosy & PHYtOchen, Manual r ~ ry Manu Laborato! lO Experiment rose — nof desire one rystallizat No - Jardue t0¢ ose) 30-47%, sucrose 0.4. ~6%, de nd gr cohol ypague 4! 1d alCO turns pag waterand fon tr otuble > (fruct . . Wy Solubility Sol’ nt 23-36%, levulose ( ts of essential oil, beeswax, pollen Brain, ituents jextrose 2 ants s , d . constitu: : dextre Lamou nins and an admi) Chemical 4 Iso contains smal Jouring pigments, Vitam Xture ,, vontaitt , soc colouring pig Honey ce mt altose, dextrin, ce id, malto nic ac inulase nvertase sedatv eptic action O i 5 ¢. antiseptic act tongue and duodenal ulcers, liver disord., t, tong i i > eve and ine pulmonary tuberculosis, marasmus, Tickey sorders. + unnary diso . tes hi ws ation: ac icide. arbu es. chaps. sealds. whitlows and skin inflammation; as e ‘ : he treatment of aphthae and other infection of the oral mu, OU the treatmel hi id fi nt of preoperative cancer. B) Castor ail Synonyms Cie Biological source |: cons communi Family Physical characters Color Coloriess ; pale \ Odor Characteristic nausea’ Taste Bland Nauseating Solubility J). insoluble in water but soluble in alcohol, light petroleum, chloroform, benzene and ether. Chemical constituents Castor o11 contains about 50 10 60%, fixed oil, which contains glyceride ester of higher fatty acids such as Ca neinoleic acid 80% isonicinile Ca ic act, s O1C acid, Stearic acid. j Co i icaci : : as nicinine Sosteric acid, linoleic acid. Italso Contains alkaloids such Uses !Asamild Purgative Or cathartic 2. Asa lubricant Requirements Paratusatory Manual of Pharmacognosy & Phytochemistry | Labor emicals in ey Ether. 1% resorcinol, HC| alpha napthol. ¢ a) Hones solvent ether one Castor oil: alcohol, light petroleum ether b) r 6, Requirements used “Teet Aube, “Tort tube , a 7. Procedure 1) Chemical Tests for Honey —— | Sr. No. s) Jl 5 | Take about 3 mL of honey shake thoroughly: allow 2 | evaporate to dryness. - Separate the upper ethe: dish, éVaporate, Residu *2mL of ether> layers to Separate aity ial layer put ina china © * 1% resorcino| and HC} 2 | Molisch’s Test, | Honey + alpha napthol + Cone. SO, | 3 Reducing Sugar Test Heat honey + 4, drop Of mixture of Fehling’ shake for 5-10 min. | Separate the upper ethereal layer and 1% Wc astor oil SN Test Observation Inference | 1 | 10mL of castor oil + 5 mL of light | petroleum ether (40 -60° C) ee Increase the amount of petroleum | ether about 1SmL | 4 = 2. | Oil ~ equal volume of alcohol and ___| ¢001 t0 0°C for 3h 9. Result On the ba che the basis of chemical analysis. it was found that given unknow1 n crude drugs are Hone ; and 0. Conclusion tcan be cone! e concluded that Honey and Castor oil were identifi ed and confirmed by using chemical tests. 1. References Pharmacognosy lab M, ngarand S ‘ayak, Pharma med Pri s ‘anual, M.A. lyengara ind S.G.K.N, yi Hyderabad. Trinity Publishing House, Sata’Laboratory Manual of Pharmacognosy & ppp yochemtstry | Ex 03 periment No. 0 2, Practical Pharmacounosy, 11. Wallis, pourth dition, Pt a 2 + indian Pharmacopoeia 2020 Mion, Pharma med Press, Hyderabad 12. Synopsis questions 1. Give synonym and biological source of honey 2. Write the synonym and biological source of castor oil 4 Give the chemical constituents ofhoney. 4. Give the chemical constituents of castor oi) 5, Whatare the uses of honey? 6. Give the uses of castor oil, 7 (Space for answers) ge Money te a Vad g Sweet ° ih Jone | Co Y role oy of beet Such as 4s s ° Apt mela dedar, Apts dorvsate, Apts jiwerea, Pypie Snaltca + : Machu, Jel, 3 - Oleurn Rearns , {rand or | Bro. Goures 27b Consists of fren onl a ope, Seed © 9 0 of wWMeeints Communttt o , Zl f Secerroahon Stovee! 6 NValerous Specrer 3+ chem: Conshtunte vf alae e dentro , Sucre: (33 Seruicko se. UY. 60- 6oe/> dined oil oii He atten pow peshies se Huw eeten Nild laxative, Bactesioetdal , Seah Ge daxahus | purgai hur 2 chathanc Bt a dubsecant . o an peep cual ory poink, enemas Area, polrs h Trinity Publishing House, Satara_— periment No. 03 Laborato | Observations }) Honey eee ee a ——] : Seer Observation Inference > —T 5 Peer ee oO Ee rmLof| [cern antl Adultevate, 1 3:mb of honey Take abou! hly, allow a > shake thoroug ethe | Jayers to separate and evaporate © | dryness yer put Separate the upper ethenal la | machina dish. & aporale nd HCI | jo resorcinol an h’s Te y ~ alpha napthol ~ SO. Molt Hone! ar Test "3. | Reducing S ~ A drop of mixture of | | | Heat honey Residue + | Red Honey Honey Pores ent | Honey Preerl wed Colby us Odio Beielé 4 Cupeo | Febling’s soluuon Aand B, 10 mL Honey ~ 5 mL petroleum oF po 4 | solvent ether — shake for 5-10 min. | Separate the upper ethereal layer and +1% | | evaporate na china dish. mL HCl. polu ttevate of atone : Color peustst fom some Hme | solution of resorcinol in | II) Castor oil Inference Observation | Test 1 | 10mL of | pewroleum ethel | Increase the am ether about 1SmL castor oil + 5 mL of light (40. -60° C) ount of petroleum [giaeagisasIoee ee _} —___—__ (Oil ~ equal volume of alcohol and _ cool 10 0°C for 3h pee er ! Result in the basis of che! ), Conclusion san be concluded that Honey and Castor o1 References >harmacognosy lab Manual, M.A. Iyengar and ical analysts, 1 Was found that given unknown crude drugs are ee and | were identified and confirmed by using chemical tests. S.G.K. Nayak, Pharma med Press, Hyderabad. gata? 4 HousLaboratory Manual of Pharmacognosy & Phytochemistry | Experiment No. 03 2, Practical Pharmacognosy, T. EF. Wallis, Fourth Edition Pharma med P. i ess, Hyderabad 3, Indian Pharmacopoeia 2020, ress, Hyderaba 12. Synopsis questions |. Give synonym and biological source of honey, 2. Write the synonym and biological source of castor oil. 3. Give the chemical constituents ofhoney. 4. Give the chemical constituents of castor oil 5, Whatare the uses ofhoney? 6. Give the uses of castor oil. (Space for answers) © 6 cle Money oa vVidd 2 Sweet Seereahon Stoveel tH Dloney Comb by wvaetous Speder ees Such as Apis roela tedau, Apts dovsate, Apis Piesea, Apts Gnatca: Mache, els Oleurm, Retr, fvand orl Bio. Sourus 29 Consiels of fens ant of ope. Seed: of erveeinrs Communttt » 3. Chem: ConeHtumnte “hf Honey & dleatrove , Sucrose Serucho se (| te ee BO- GOe/s jive oil 5° Me altn pr pea hes TV ochre Hee eaten Nfld hacatieer, Bactesioetdal , Sedat te. B= dacatus; purgatue 2 chathamc At a dubm cant ' g olte he ual? OY pounks,enemas Aire Uk, P On peep Trinity Publishing House, SataraExperiment No. 03 Marks 20 Laboratory Manual of Pharmacognosy &P hyt Meco x _— Signature of teacher with dateExperiment No. 04 DETERMINATION OF STOMATAL NUMBER AND INDEX 1,Aim : atura leat he stomatal number and stomatal index in the given sample of Datura leat To determine U ecies of crude nificance rent varieties OF SP 2. practical sig! Linear measurement of leaf constituents is useful tool to identify diffe s. These are also helpful in identification of the adulterants in crude drugs. Stomata are teats Is. Determination of stomatal number and stomatal index |, the students will learn the leaf surfaces, herbaceous stem, petioles and tendril «, useful to estimate the rate of water Joss through stomata. In this practical ; in f stomatal number and stomatal index in f constituents in order t t climate leaves. present on the linear measurement of Leal importance © compensate the differences between sun and shade leaves, dry climate leaves and wet 3. practical outcomes (PrOs) - apped| BTL pro Practical outcomes After completion of this practical co 1, the students will be able to: a pro 1 | Explain the significance of stomatal number and stomatal index. co2 BIL2 pro 2 | Identify the given leaf sample with the help of linear measurement. co2 \ BTL4 pro 3 Examine average number of stomata present in Isq. mm. area of leaf . 3 epidermis using camera lucida. co2 | BTL3 | pro 4 | Calculate the stomatal number and stomatal index of the given leaf Cco2 BTL3 sample. PprO 5 | Collaborate and communicate with fellow students. \ Cco2 \ BTLS 4. Theory For the measurement of dimensions of plant constituents, micrometers required bb trace the magnified image of the microscopic object. With the proper adjustment of camera lucida, it is possible to trace the required outlines on the black drawing sheets with the help of pencils. Itis accurate and fast method of tracing the object. Microscopic objects are measured with micrometers such as stage micrometer and eyepiece micrometer. Stage micrometer (SM) is a calibrated scale having 100 divisions of | mm and therefore | division if equal to 0.01 mm or 10 pm or micro meter. are required. Camera lucida is Figure 4.1: Diagram of Camera lucidaLaboratory Manual of Pharmacognog —_——— SY & Ph Experment No. 04 _ Hoth, ~ Nig, ‘, = ) yo 2 30 40 50 60 7 wiih Ul inkl . Figure 4.3; Micrometer slide rd cell and surrounding subsidiary cells. [t, fu co. Cty and carbon dioxide exchange. Stomata are Prese Tons * : idney shaped guai Thon leaf surfaces. herbaceous stem, petioles and tendrils. There are two Ken ye Nee en mee suy oun woth subsidiary cells. Based on the arrangement of epidermal a ‘e i semana ° Stomata si diacvte (subsidiary cells are arranged with long axis at right angles 10 oe 8), Paraeyi, th arrangement of subsidiary cells with long axis 1s parallel to the stomatal pore), anisocytic (sto u i hich i: surrounded by three to four subsidiary cells one of w is surrounded by indefinite number of subsidiary cells without any specific ‘ype of. arrangement), Stomatal number is defined as the average number of stomata per sq. mm. of epidermis ofthe leat. The ay number of stomata per sq. mm may vary for the leaves of the same plant grown in different environme,y, : under different climatic conditions. It is, however shown that the ratio of the number of stomata to thera, number of epidermal cells in a given area of epidermis is fairly constant for any age of the plant ang Unig different climatic conditions. Figure 4.2: Ocular micrometer . eni! a at Stomata 1s a minute epidermal opening has gu: transpiration and re gulation of the water, oxygen s smaller than other two) and anomocysj,, A i Stop, N Stomatal index is the percentage which the number of stomata forms to the total number of epiderma| Cell each stoma being counted as one cell. Stomatal index can be calculated by using the following equation: — (S x 100) (E+S) S= Number of stomata per unit area Stomatal index = E= Number of epidermal cells in the same unit area (a) Anisocytic stomata (b) Diacytic stomata (c) Paracytic stomata (d) Anomocytic stomata Figure 4.4: Types of stomata Whilst stomatal number varies considerably with the age of the leaf and due to changes in environment! conditions, stomatal index is relatively constant and therefore, it has diagnostic significance for a given species of the plant. It is employed for the differentiation of allied or closely related to species of same genus mair dried, as wellas fresh conditions Trinity Publishing House, Sata? iexperiment No = Manual of Pharmacognosy & Phytochemistry | Labora pied. 1: Stomatal Index of Various Crude Drugs gable 4-1: SOM | | Stomatal Index | Lowe rpalisade | 114-124-133 V71-190-207 | tel [127-174-194 Tatura siramonivn | teai81-204 7 | Digitalis tanata "139-144-147 Fs | Digi purpurea 16 40 5. Requirements s; Stage micrometer, camera lucida, drawing boa lass, blade, cello tape, black drawing sheet, dark i ce] jam} 1d, miero slides, coverslips, forceps, Spin lamP aratus Ai watchg colored pencil with sharp lead s Chemicals: Chioral hydrate solution, glycerin Instrument: Compound microscope 6. Requirements used 7, Precautions a) Usea mature leaf for determination of leaf constituents. b) Keep the microscope in vertical position while using camera lucida. c) Adjust the microscope lamp in such a way that the illumination on drawing paper place at the side of microscope which should be equal to that on the mounted object. d) Use drawing board to provide support to drawing sheet, ifrequired tiltit at proper angle. e) Ensure equal magnification on all parts of the board. 8. Procedure A, Preparation of lamina a) Take a mature leaf, if it is too small to identify take the whole leaf and if the leaf is large, cut it into a number of pieces of 5 mm square from the middle portion between the lamina and midrib b) Place the leaf in test tube containing chloral hydrate and boil the tube in water bath. c) Separate out the epidermis and place it on a slide with the help of a brush along with 1-2 drops of chloral hydrate. d) Then cool it and place a coverslip. Fresh leaf a, Take a given leaf sample and separate epidermis. If the leaf sample is thick, peel off the epidermis. by ing House, Sataramacognosy & Phytoch, en r hy n. The epidermis separates ou expenment No. O8 serit peaking timo Pe py sheer Mg with 1-2 drobsioF ch ro thchtoral Isat jong loral pb. Bol the lea jemi aa sinde with hve, vy place tne ePIde a coversiiP for 30min. na water bath o€ cool and then ple t whether the stomata ar © Pres pry leaf e i wjnatest tube o sat the leat chloral hydrate in? ; are 2 tin o at ere miessenPe b Cur he leat ito 880 PIES ath surfaces ornot oe eins facing dow” (Toobserve the upper epidermis) 4 " othe lower epidermis). caf with the ¥ veins facing up coverslip. \d trace th jidermis then use ut t eared | rds. (To obser ifwith jycerinand place sjower™ an ¢ Place thech Place the other ha e Add nvo drops ofgl f Label the slides as “U} Jeafis too thick and dai Jeaf with chloral hydrate a b Place one half with the upper surface « Carefully scrape ofthe upper tissue, 4 Clean it with a brush dipped in chloral hydrate .pidermis. Turn the layer upside to trace the cells. cedure with the second half, this time placi he epidermal cellsand stomata, following method- pper” and ne leaf into two halves. rk to separate the eP s given ins! facing downwards. with the edge ofa bla solution. The layer of Ifthe a Clear the tepno.| and urbing upper epidermis, de, without dist gon the surface is the cells remainin, upper ¢} ¢ Repeat the proce ing the lower surface facing downwards, proce, asgiven instep no. 3and4- “ f Usually herbs and small shrubs have stomata on both surfaces. In tree species, stomata are present on th, j ventral leaf, almost the same numby, More stomata are present on the lower surface in dors: Jower surface. inisobila B) Calibration ofucular micrometer a) Place the SM on the stage of the microscopeand focus I b) Place the ocular micrometer (OM) in one of the eye pit evepreces. align the scale of OM with that of SM. ©) Count the divisions of OM coinciding with divisions of SM. sions of stage micrometer itatagiven magnification e.g. 40x. eces of microscope. While observing through th, le If 4 divisions eyepiece micrometer = 3 divi: The value of one divis r 1 sion of eyepiece micromete: f¢ i er is calcul: ! division of SM =0.01mm = 10 ee 3 divisions of SM =30 When 4 div e) Sek isions eyepiece micrometer =3 divisions of stage micromet 30 a ; s of s = 7 ot ! division of eyepiece micrometer = 1/4 X 30= 7.5, 7 , en place the microscopic deciola pic object on the stage of 1 ge of microscope and i ig s observe, if it meas y easures 6 divi Stons ws- then the size of object will be- 6X75=45u d) By follow e steps, rate Ir 5 By ing the above three ste] er magn Ms as wel Ox coe lor ee steps, calibrate the ocular scale for oth er magnifications as is Well i.e. 10X ¢) Remove SM 'M once the ocular scale is calibrated for diffe ifferent having sample material. ° magnification(s) and place the s| sli 7 : rinity Publishing House, Satara4 M mi ment No. 0 poratory Ma’ jal of Pharmacognosy & Phytochemistry | Expenm= abe a a pacing of Stomata in tnisexpermen’ the number of stomata has to be determined per square millimelre adjust the drawing board if swift camera-lucida is used, It is not nece opera lucida sary to adjust angle with Abbe’s cam anthe hep ofstage micrometer, draw Tine of Imm using 1010 magnification on a drawing sheet and ue" i }Ocm square on that ine (1 mm {Oem) if magnification is correct oe place thestagermicrometer with apreparedsid ofthe leaf even for stomatal index 3g. Mark the number of stomata in the square with the help of camera lucida Count the stomata that gives the number of stomata per sq. mm. e ¢Take 95 reading and calculate the average. Calculate the stomatal index by counting number of stomata and epidermal cells £ 9, observations a) Total no- ofepidermal cells (E) in 1sq. mm area in upper surface (A) = ___ p) Total no- ofepidermal cells (E) in 1sq. mm area in lower surface (B) c) Total no. of stomata per sq. mm (S) in upper surface (A) = _ d) Total no. of stomata per sq. mm (S) in lower surface (B)= _ 10. Observation table ——t SN No. of epidermal cells) No. of Stomata in 1sq. mm area (E) per sq. mm (S) ——y -| Upper Lower Upper Lower | Upper Lower Upper | Lower | surface (A) | surface (B) |surface (A) | surface (B) | surface (A) |surface @) | surface (A) | surface (B)) | | we CI | | |Stomatal number | ‘Stomatal index Trinity Publishing House, Satara> Expenment No 04 _ SV No. of epidermal cells jin fsq.mm area (F) Upper surface (A) surface Laboratory Manual of Pharmacognosy & pp, ab ®Y 8 Phviog, o Soc of Stomata [Stomatal number | Stomatatigg nde x per sq.m oy] +> $$ 2 Lower) Upper Lower Upper | ; ; | ce (A) | surface (B) | surfacy Lo (surface (A) surface (B) surface “pe ee (A) I suntan | [ & Lower Upper tal (Space for affixing black sheet) ——_ Trinity Publishing House, SataraExpenment NO 04 tory Manual of Pharmacognosy & Phytocherni al Labor! “caleutatlons 1 of Eygpiece micrometer (for one division ration ) t alibi ye divisions of stage micrometercoincide with __ divisions on eyepiece MC 1 5 division “ divisions of stage micrometer: mm mn 7 af nicromete - 3 divisions of eyepiece micrometer divisions of stage n im gwen fSM=0.01mm~ 104. therefore, | division of eyepiece micrometer “ stomatal number | nufhber (Average) in upper epidermis - | number (Average) in lower epidermis B) 1, Stomatal 2. stomatal 5 © stomatal index (Ave stomatal index Average) in upper epidermis = Le . 4s | under a microse ahigher der i jant species or subspecies. vein islets in a leat type significance and val have taxonomic -Vein islet number a seers islet number — te 49.5 -22.5 25.6 -32.8 “95-25 | | fig wtotolia 32.7 - 40.2 ceva acuofolia | « purpurea 20-55 24-42 | 31 Enthronion coca [- 8.0- 12.0 16.1 - 21.0 | 4 _ |] lides, cover glasses, forceps. Spiri, 5. Requirements cro S| er, camera lucida, drawing board mi colored pencil with sharp lead. Apparatus Stage micromet lo tape, black drawing sheet, dark lamp small watch glass. blade, cel Chemicals: Chloral hydrate solution, glycerin. Instrument Compound microscope. 6. Requirements used 7. Precautions Take precautions as per prev 10us experiment. 8. Procedure A) Preparation of lamina Procedure as per previous experiment B) Tracing of Veinislet number and Vein termination number a) Boil the leaf pieces forabout 30 min ina chloral hydrate solution to remove the debris. b) Place the preparation ina test tube of glycerin water. ¢ ) Set up the camera lucida and use the stage micrometer to divide the paper into I sq. mm squares. SS Trinity Publishing House, Sataratory Manual of Pharmacognosy & Labor Phytochemistry | Experment No. 05 Replace the stage micrometer with the ele Cette ale XA Wa Geog ‘wed leaf preparation and trace the veins tn four contunueds aque O10 pe Hi Mares or T mm X 4 mm rectangles When a superimposed image of the leat grtion and Paper IS VISIDIE, USE the microscope to trace the vein-islet and vein termination number jnutancously 1d 25 observations as 7 arange f ¢) Reco! geand indicate the mean y alue 0 Count the number of vein-islets and vein te nd vein termination number in the square or rectangle, ay well as the ber of incomplete vein-islets nul and vein termination number and any two adjacent sides of the square oF rectangle ory calculate the value fc © square ; y) To calculate | for one square mm, divide the total number of yei-tset numbers and ve termination number in four adjoining squares by four h) Record the observations as a range and indicate the mean value 9. Observations 1. The vein islet number in Isq.mmareain | rectangle 2. The vein islet number in 4 sq. mmarea in4 rectangles = 3 ‘The vein termination numberin Isq. mm area in | rectangle = 4. ‘The vein termination number in Isq. mm area in4 rectangles = ; 10. Observation table 1] Vein islet number and vein termination number [SN | Vein islet number in Mean value 1sq. mm area in J rectangle Vein termination Mean value number | sq. mm. area in 4 rectangles 10 Trinity Publishing House, Satara ieee aes eee eee ee et—T Vein termination Mean y, Mean vay ~ aly ec | Vein is © qq. mm area in | number 1 sq. mm ee _— pee ee | _ . (Space for affixing black drawing sheet) al Eee ee eee eee Trinity Publishing House, Sata’éManual of Laboratory M anual o| Pharmacognosy & Phytoch = Wochemistry | 11. Results ~ ____ Experiment No. 05 le Die Was found to be in the range of ‘he vein termination number in g: f oT ' given leaf Sample was found und to bein the Tange of 12. Conclusion From the vein islet number and vein termination ny b beoe | | TTT mber, itcan be concluded that the given leaf sample may 13. References ay hutps:/Pharmacyinfoline.com/determination-ot-yein | » am a i in Pharmacognosy” 4th edition., a ed 7. c) Khande! ~ Practical Pharmacognosy Techniques & Bisdinedacaaiabdah Ket Prakashan 14. Synopsis questions a) Define vein-islet number, b) What is the significance of vein islet number? c) Give the veinlet termination number for Digitalis Purpurea. d) What is vein termination number? . e) Why vein termination number should be determined? sma] | n vein Pele number vefers 40 the Court 0 veine ascociateal Struature en Peanes aldin Photosynth ests 2 tenf S berud tue Support . b ~ vetn Fslet numben Fndroater that leap Structuyal Complen? fegeeis plaot photos yothests : physfeat nutment dretabutathon , (Space for answers) a* 5 | count vi Vin seamithnabon number aefers +0 the vein endfrq fp the Cea. th undlewsta ndrr ( team? naho n numb ex_ helps . ly vets Vaceutar Systm Pnt moa ales -the teat es Vern 4 Maton number helps fh uh denstanding earring ° the leag vascular Systm Pntereates , aiding ip Studuma nuterent dfstrbubon ,wates transport ' © orice transport 7 Overall ead gunetion arty 7 Trinity Pub ing House, Satara ey eo, _ Signature of Teacher with daté rinity Publishing House, $22”Experiment No. 06 DETERMINATION OF PALISADE RATIO 7 aio, sine te palisaderatio ofthe given sample ot Vi qode practical significance 2 de ratio studies are required for identification of leafy constituents of crude drugs. It is an important sal chara Pleaf, if C1 poet for determination and characterization of leafy drugs. These are the constant for a parucular eras of plant and thus they ae beneficial in ideutification ofadulteramts In ths practical, the students 8 ill speci the importance of palisade ratio in linear measurement of leafy constituents in order to do critical Jean nation of certain related species of same drugs. getermil 4, practical outcomes (POs) 7 Practical outcomes = | (| After completion of this practical, the students will be able to: co | | (p01 Explain the significance of palisade ratio cor | itd | [p02 [Identify the given leaf sample with the help of palisade ratio cor | BTL | Observe palisade ratio present in Isq. mm. area of leaf epidermis cor PBT Calculate the palisade ratio of the given leaf sample CO2 aris prO 5| Follow cleanliness, safety and ethical practices in the laboratory CO2 BILS | 4, Theory The average number of palisade cells beneath each epidermal cell is defined as the palisade ratio. In pharmacognosy, the palisade ratio is a quantitative metric used to evaluate the quality of plant leaves. Palisade parenchyma is the proportionate to the spongy parenchyma. It is responsible for photosynthesis within the leaf. Unlike vein-islet number, which requires an unbroken portion of the leaf to be determined, palisade ratio can be determined with the powdered drug. Zorning and Weiss introduced the palisade ratio determination technique in their studies on the Composite family in 1925. Figure 6.1 Palisade ratio Ina leaf section, thickness of the palisade parenchyma and the spongy parenchyma is used to calculate the Palisade ratio. The ratio is calculated by dividing the thickness of palisade parenchyma by the thickness of spongy parenchyma. Trinity Publishing House, SataraN Laboratory Manual of Pharmacognosy & Pp — toe h Experiment No. 06 . Sy Je ratio ts calculated by measuring the thickness of the palisade Pare Mis Ina leaf section. the patie rota dvi the thickness of palisade pare ny, ¥) vn ‘yma jy hyma the spongy parenc thickness ofspongy parenchyma §. Requirements sna slides. . ‘ + camera huctda, drawing board micro slides, cover glasses, f,,, amen 1s e od z, ce black drawing sheet, dark colored pencil with sharp leag sy Dip " a i Apparatus: Stage micron mp small wateh glass, blade cello tape, Chemicals: Chloral hydrate solution, glycerin Instrument Compound microscope 6. Requirements used *. Precautions utions as per previous experiment. Take pr 8. Procedure 1) Preparation of lamina Procedure as per previous experiment Il) Tracing of palisade ratio a) Boil the chloral hydrate solution to clear the pieces of leaf (about 2 sq.mm) or powder. b) Immerse the sample in glycerin water and focus with a high magnification lens. | ©) Attach the camera lucida and draw the outlines of four continuous epidermal cells. d) Focus the palisade cells by lowering the draw tube slightly and tracing the palisade cells that are direc, beneath the same four epidermal cells. 9. Observations 4. Palisade ratio in Isq. mm area in I rectangle in upper surface = b Palisade ratio in Isq. mmarea in 4 rectangles in upper surface © Palisade ratio in 1sq. mm area in | rectangle in lower surface = Palisade ratio in Isq. mm area in4 rectangles in lower surface Trinity Publishing House, Sata’\ pi iy yy yy —— T surface (A) n Isq. mm area in I rectangle Lower Surface (B) Experiment No. 06 Palisade ratio 1 sq. mm area in 4rectangles Lower Surfa [Upper surface (A) | Trinity Publishing House, Satara UeoN” snarmacognosy & Phytoc yy Manwalol! a drawins sheet sor apis Meat spor 1) Results dtobe Palisade rane 1 upper » lowersurface in g1¥ surfacein given leaf sample was foun mnleafsample was foundtobe ___~ 12 Conclusion Ze ratio. itcan be concluded that th given leaf sample may be of. 13, References pharmacyinfoline.com determination-of-vein-islet-number/ 2)CK Kekate Practical Pharmacognosy’ 4thedition., Vallabh Prakashan. Page no. 117-119. )\sal. Practical Pharmacognosy Techniques & Experiments’ 22nd edition, Nirali Prakashay, KR Kb omtance of palisade ratio? ©)Give the palisade 10 for Digitalis purpurea. d) How palisade rano 1s determined? ©) Who introduced palisade ratio? (Space for answers) : ation Sefert ty the vat palisad 1 er THIS” meas ! e faho Cell +o 4+herw w rat remenl Of tent usect %n bot Py J nb ayn oO pone -TL. A+ ~The patroa de length JU Thnity Publishing House, Sata'amanual of Pharmacognosy & Ph /lochemsty | Enpermment No. 08 |, Cea A 7 sods abot? shape va plont Celle tm f all?caol eax? 7 ~ U Ane c+ : a 1 . al’sade wand Ps imprrebant becasue + he ) pueen -the eicienct » ang ° lant 7 pe pte synthecu eee J 9 n?q hev paltisade Urako meaning £0” mowers Calls the at ey increas es efger abscor phon 0 hos ’ Sus sWoe aaa aaa nthe 9 enhances pbotos 4? Ineo JU Cc) Ahe pace aoke Can be deteamined PY. mf nowseopic i 4aning Tmages ; reeasuntng the ye 2 wtdth ob patuzade teu» “Then she Length by ahete width +6 ate ) was 2 bewer ° nterooltedn a pall?saole age ol Marks: Signature of teacher with date Trinity Publishing House, Satara *h plant £ beag ® Sechor the ceus tS divided t the pattistale mahon~ Y Paperiment Now07 ; Ne DETERMINATION OF SIZE OF STARCH GRAINS Son sainples of tirch grants by using optical microscopy. 2, Practical significance mierostope plays an important role in determination of s Sow er of sev eral hundreds, Starch is an important source of carbohydrates i dietang ° Ms excipients in industry. The size of starch grains depends on its botanical soy,g. ts Condition daring its growth. Therefore, to Wentify the source of starch, it is esseryicy id of starch grains. In this practical, the students will learn the importance of Meastirginen, to . of ize and shape of starch graing due Ct sizeof starch entifis and will differentiate smallest and largest grains. ‘ 4 Fee 3. Practical outcomes (PrOs) Se : so PrO Practical outcomes ree so. 7s. y° |Mapped) Br After completion of this practical, the students will be able to: . co PrO | Explain the significance of determination of diameter of starch grains. | _CO3__| BTLp 'PrO,2 Deteranne the diameter of starch grains by using eyepiece and stage cos | BT micrometer ~~ x . aativfes ‘|, Analyse the given sample and identify the source of starch grains co3 | BTLg Collaborate and communicate with fellow students. Cco3 BTLs Follow cleanliness, safety and ethical practices in the laboratory co3. | BTLs~ 4. Theory Starch is present in different parts of the plant in the form of granules of varying size. Starch is foung abundantly in fruit. seed. root, rhizome and as smaller grains in chlorophyll containing tissue of the plant such as leaf Starches of different origins can be identified by studying their size, shape and structure, as wel as. position of the hilum and striations. Chemically, starches are polysaccharides containing amylopectin and fi-amy lose Starch tums blue to violet when treated with iodine solution. Starches of pharmaceutical interest are obtained from maize, rice, wheat and potato. These starches can be differentiated from each other by microscopical examination. The shape, size and structure of starch grains are Varying within definite limit in different sources of plant. The diameter of starch grains present in crude drug plays an important role in identification of particular drug and determination of type of adulteration. By comparing the sizes of starch grains in different species, the adulterants and/or substitutes in crude drugs can be distinguished using optical microscopy. The size of the starch grains 1s determined by measuring the size of 20 starch grains to determine the largest and smallest diameters and the difference between diameters of botanical species of drug. Trinity Publishing House, Satara rr ——————————