Successfully reported this slideshow.
We use your LinkedIn profile and activity data to personalize ads and to show you more relevant ads. You can change your ad preferences anytime.

Extraction and identification of phyto-constituents

6,712 views

Published on

method to separate compounds based on their relative solubilities in two different immiscible liquids, usually water and an organic solvent. It is an extraction of a substance from one liquid into another liquid phase.

Published in: Education

Extraction and identification of phyto-constituents

  1. 1. EXTRACTION AND IDENTIFICATION OF PHYTO-CONSTITUENTS K.Sudheer Kumar, Assistant professor. Dept.of Pharmacognosy Chilkur Balaji college of Pharmacy Hyderabad. E-mail:sudheer.y2k8@gmail.com K.Sudheer Kumar, Assistant professor. Dept.of Pharmacognosy Chilkur Balaji college of Pharmacy Hyderabad. E-mail:sudheer.y2k8@gmail.com
  2. 2. INTRODUCTION Extraction is the process of efficiently dissolving and separating the desired constituent from crude drug with the use of solvent The choice of solvent depends on charachteristics of the secondary metabolites like polarity, pH, thermal stability. The solvent should non toxic,inflammable,easy to remove. Factors effecting extraction: a)Quantity and chemical nature b)Size of powder of crude drug c)Moisture content d)Nature and volume of solvent e)Mixing ration of solvent and sample f)Temperature of extraction process g)pH of extraction solvent h)Lipophilicity of solvent mixture and sample INTRODUCTION Extraction is the process of efficiently dissolving and separating the desired constituent from crude drug with the use of solvent The choice of solvent depends on charachteristics of the secondary metabolites like polarity, pH, thermal stability. The solvent should non toxic,inflammable,easy to remove. Factors effecting extraction: a)Quantity and chemical nature b)Size of powder of crude drug c)Moisture content d)Nature and volume of solvent e)Mixing ration of solvent and sample f)Temperature of extraction process g)pH of extraction solvent h)Lipophilicity of solvent mixture and sample
  3. 3. Types of extractions Maceration : Maceration is the extraction process of crude drugs from crushed or pressed plant and animal material, which is soaked in a suitable solvent stirring at room temperature. Place solid material with whole of menustrum in a closed vessel and allow stand for 7 days Shaking occasionally/strain pressing marc Mixing liquids obtained Clarify by filtration Seven days are consider to adequate period of equilibrium between solute and solvent
  4. 4. PERCOLATION Passage of menustrum under gravity column Moisten the solid material with sufficient quantity of menustrum Allow to stand for 4 hrs in well closed vessel Packed in a percolator(24 hrs) Allow percolation to proceed slowly until percolation measures about three quarters of volume required for finished tincture Press marc mix expressed liquid with percolate Clarify by filtration
  5. 5. SOXHLET EXTRACTION It is also known as counter current extraction A piece of laboratory apparatus invented in 1879 by Franz von Soxhlet. Originally designed for the extraction of a lipid from a solid material. However, a Soxhlet extractor is not limited to the extraction of lipids. Typically, a Soxhlet extraction is only required where the desired compound has a limited solubility in a solvent, and the impurity is insoluble in that solvent. SOXHLET EXTRACTION It is also known as counter current extraction A piece of laboratory apparatus invented in 1879 by Franz von Soxhlet. Originally designed for the extraction of a lipid from a solid material. However, a Soxhlet extractor is not limited to the extraction of lipids. Typically, a Soxhlet extraction is only required where the desired compound has a limited solubility in a solvent, and the impurity is insoluble in that solvent.
  6. 6. Drug is formed in to powder and the solvent is mixed in to it in equal quantity place in apparatus and evaporate with heat condensation takes place ,vapors are collected. The thimble is loaded into the main chamber of the Soxhlete xtractor. The extraction solvent to be used is placed in a distillation flask. The flask is placed on the heating element. The Soxhlet extractor is placed atop the flask. A reflux condenser is placed atop the extractor.
  7. 7. 1. Stirrer bar 2. Still pot 3. Distillation path 4. Thimble 5. Solid 6. Siphon top 7. Siphon exit 8. Expansion adapter 9. Condenser 10. Cooling water in 11. Cooling water out 1. Stirrer bar 2. Still pot 3. Distillation path 4. Thimble 5. Solid 6. Siphon top 7. Siphon exit 8. Expansion adapter 9. Condenser 10. Cooling water in 11. Cooling water out Parts of Soxhlet extractorParts of Soxhlet extractor
  8. 8. Continuous extraction of a component from a solid mixture. Boiling solvent vapors rise up through the larger side-arm Condensed drops of solvent fall into the porous cup, dissolving out the desired component from a solid mixture When the smaller side-arm fills to overflowing, it initiates a siphoning action. Residual solvent then drains out of the porous cup, as fresh solvent drops continue to fall into the porous cup. The solvent, containing the dissolved component, is siphoned into the boiler below Continuous extraction of a component from a solid mixture. Boiling solvent vapors rise up through the larger side-arm Condensed drops of solvent fall into the porous cup, dissolving out the desired component from a solid mixture When the smaller side-arm fills to overflowing, it initiates a siphoning action. Residual solvent then drains out of the porous cup, as fresh solvent drops continue to fall into the porous cup. The solvent, containing the dissolved component, is siphoned into the boiler below STEPS
  9. 9. INFUSION Infusion is the process of extracting chemical compounds or flavors from plant material in a solvent such as water, oil or alcohol, by allowing the material to remain suspended in the solvent over time (a process often called steeping). An infusion is also the name for the resultant liquid. The process of infusion is distinct from decoction, which involves boiling the plant material, or percolation, in which the water passes through the material This is very imp and simple method for extraction used for vitamins, volatile ingredients and soft ingredients in which the powdered drug is extracted with hot or cold water. INFUSION Infusion is the process of extracting chemical compounds or flavors from plant material in a solvent such as water, oil or alcohol, by allowing the material to remain suspended in the solvent over time (a process often called steeping). An infusion is also the name for the resultant liquid. The process of infusion is distinct from decoction, which involves boiling the plant material, or percolation, in which the water passes through the material This is very imp and simple method for extraction used for vitamins, volatile ingredients and soft ingredients in which the powdered drug is extracted with hot or cold water.
  10. 10. Most people are familiar with an infusion which is what people do when they put a tea bag or tea ball in a cup of hot water and allow it to steep a couple minutes before drinking. Infusions are used to extract vitamins and volatile ingredients from soft ingredients like leaves, flowers, citrus peelings, etc. Green tea, black tea, peppermint tea and chamomile tea are examples of an infusion. The short brewing time helps to retain the volatile ingredients while drinking. Most people are familiar with an infusion which is what people do when they put a tea bag or tea ball in a cup of hot water and allow it to steep a couple minutes before drinking. Infusions are used to extract vitamins and volatile ingredients from soft ingredients like leaves, flowers, citrus peelings, etc. Green tea, black tea, peppermint tea and chamomile tea are examples of an infusion. The short brewing time helps to retain the volatile ingredients while drinking.
  11. 11. METHOD Powder drug is soaked in hot or Luke warm water Filtered otter specified time Press the marc and extract again with fresh water or Luke warm water METHOD Powder drug is soaked in hot or Luke warm water Filtered otter specified time Press the marc and extract again with fresh water or Luke warm water
  12. 12. DECOCTION Decoction is a method of extraction by boiling of dissolved chemicals from herbal or plant material, which may include stems, roots, bark and rhizomes. Decoction involves first mashing and then boiling in water to extract oils, volatile organic compounds and other chemical substances. Decoction can be used to make herbal teas, teas, coffees, tinctures and similar solutions. Decoction means concentrate by boiling .this method is useful for the wood,seed,type of drugs. Powdered drug is boiled with water Filtered
  13. 13. A decoction is used to extract primarily the mineral salts and bitter principles of plants from hard materials such as roots, bark, seeds and wood. These hard materials generally require boiling for at least 10 minutes and then are allowed to steep for a number of hours. The tea is boiled down and concentrated so that water needs to be added before drinking. The word "decoct" means to concentrate by boiling. Essiac tea and taheebo bark tea are examples of a decoction.
  14. 14. WATER DISTILLATION This method is useful for the extraction of essential oils Distilled water is water that has had many of its impurities removed through distillation. Distillation involves boiling the water and then condensing the steam into a clean container. Applications In chemical and biological laboratories, as well as in industry, cheaper alternatives such as deionized water are preferred to distilled water. But if these alternatives are not pure enough, distilled water is used. If exceptionally high purity water is required, double distilled water is used.
  15. 15. Heating of plant material with water Vapor of volatile oil component is passed to wards condenser along with steam Oil and water layers are separated on cooling Heating of plant material with water Vapor of volatile oil component is passed to wards condenser along with steam Oil and water layers are separated on cooling Water distillation unitWater distillation unit
  16. 16. STEAM DISTILLATION Steam distillation is a special type of distillation (a separation process) for temperature sensitive materials like natural aromatic compounds. It once was a popular laboratory method for purification of organic compounds, but has become obsolete by vacuum distillation. STEAM DISTILLATION Steam distillation is a special type of distillation (a separation process) for temperature sensitive materials like natural aromatic compounds. It once was a popular laboratory method for purification of organic compounds, but has become obsolete by vacuum distillation.
  17. 17. Steam distillation method Steam passed through plant material suspended in water Steam which can collected and cool it subsequently In direct steam distillation meshes are used to separate steam from material and vapor collected Steam distillation method Steam passed through plant material suspended in water Steam which can collected and cool it subsequently In direct steam distillation meshes are used to separate steam from material and vapor collected
  18. 18. EXPRESSION OF OILS When oils are taken from the rind of fruits they are called "expressed oils".  There are many different ways to extract essential oils and they are listed below: a.Enfleurage b.Expressed Oils c.Steam Distillation d.Solvent Extraction e.Fractional Distillation and Percolation f.Carbon Dioxide Extraction g.Phytonic Process EXPRESSION OF OILS When oils are taken from the rind of fruits they are called "expressed oils".  There are many different ways to extract essential oils and they are listed below: a.Enfleurage b.Expressed Oils c.Steam Distillation d.Solvent Extraction e.Fractional Distillation and Percolation f.Carbon Dioxide Extraction g.Phytonic Process
  19. 19. a)Enfleurage is one of the oldest methods of extracting essential oils and is rarely used these days because of its high cost. It involves placing the flower petals on a layer of glass that is first spread with a thin layer of fat called "chassis". The volatile oil diffuses into the fat, then the fat is collected and the oil is extracted from the fat using alcohol. b)When oils are taken from the rind of fruits they are called "expressed oils". This method is cold and does not involve any solvents or heat of any kind. Most of the citrus oils are extracted in this manner including grapefruit, lime, orange and lemon essential oils.
  20. 20. C)Steam distillation has been used for hundreds of years and today remains one of the most favorably methods of extracting essential oils. There are actually three different steam processes that are described further in the distilling section. But in all of the methods, steam is used to rupture the oil membranes in the plant and release the essential oil. The steam carries the essential oil to a condenser and then as it re- liquefies the lighter essential oil floats on top. The water and oil is then separated and the water portion is referred to as the hydrosol, hydrolyte, or flower or floral water. C)Steam distillation has been used for hundreds of years and today remains one of the most favorably methods of extracting essential oils. There are actually three different steam processes that are described further in the distilling section. But in all of the methods, steam is used to rupture the oil membranes in the plant and release the essential oil. The steam carries the essential oil to a condenser and then as it re- liquefies the lighter essential oil floats on top. The water and oil is then separated and the water portion is referred to as the hydrosol, hydrolyte, or flower or floral water.
  21. 21. D)Solvent extraction is a method of extracting essential oils that is dominated by the perfume industry. The oil particles are released when the concrete substance is then mixed with alcohol. Once it is separated out, the chemicals remain in the oil and the oil is used by the perfume industry. These are perfume or aromatherapy grade oils. E) Fractional distillation separates the volatile oil in different fractions or portions at various boiling points. This is used in oil refineries for distillation of petroleum products and is not suited for therapeutic grade essential oils. D)Solvent extraction is a method of extracting essential oils that is dominated by the perfume industry. The oil particles are released when the concrete substance is then mixed with alcohol. Once it is separated out, the chemicals remain in the oil and the oil is used by the perfume industry. These are perfume or aromatherapy grade oils. E) Fractional distillation separates the volatile oil in different fractions or portions at various boiling points. This is used in oil refineries for distillation of petroleum products and is not suited for therapeutic grade essential oils.
  22. 22. F) Carbon Dioxide Extraction or Super critical fluid extraction or carbon dioxide extraction is also a fairly new solvent extraction process that uses carbon dioxide at very high pressure. The carbon dioxide is injected into the tank where the plant material is contained and acts as a solvent to extract the oil. G)The Phytonic process is a one of the newest methods of extracting essential oils using non-CFCs (non-chlorofluorocarbons). It is also called Florasol Extraction and the oils are referred to as phytols. F) Carbon Dioxide Extraction or Super critical fluid extraction or carbon dioxide extraction is also a fairly new solvent extraction process that uses carbon dioxide at very high pressure. The carbon dioxide is injected into the tank where the plant material is contained and acts as a solvent to extract the oil. G)The Phytonic process is a one of the newest methods of extracting essential oils using non-CFCs (non-chlorofluorocarbons). It is also called Florasol Extraction and the oils are referred to as phytols.
  23. 23. MICRO WAVE ASSISTED EXTRACTION Micro wave works on high temperature produced by micro-waves evaporate moisture in plant powder which dehydrates cellulos,ruptures cell wall, and thus solubilizes phytoconstituents. Microwave-assisted extraction (MAE) or simply microwave extraction is a relatively new extraction technique that combines microwave and traditional solvent extraction. Application of microwaves for heating the solvents and plant tissues in extraction process, which increases the kinetic of extraction, is called microwave-assisted extraction. MAE has a number of advantages, e.g., shorter extraction time, less solvent, higher extraction rate and lower cost, MICRO WAVE ASSISTED EXTRACTION Micro wave works on high temperature produced by micro-waves evaporate moisture in plant powder which dehydrates cellulos,ruptures cell wall, and thus solubilizes phytoconstituents. Microwave-assisted extraction (MAE) or simply microwave extraction is a relatively new extraction technique that combines microwave and traditional solvent extraction. Application of microwaves for heating the solvents and plant tissues in extraction process, which increases the kinetic of extraction, is called microwave-assisted extraction. MAE has a number of advantages, e.g., shorter extraction time, less solvent, higher extraction rate and lower cost,
  24. 24. Traditional method of extraction of compounds from various matrices, especially natural products. The use of MAE in natural products extraction started in the late 1980s, and through the technological developments, it has now become one of the popular and cost-effective extraction methods available today. Several advanced MAE instrumentations and methodologies have become available, e.g., pressurized microwave-assisted extraction (PMAE) and solvent-free microwave-assisted extraction (SFMAE). Traditional method of extraction of compounds from various matrices, especially natural products. The use of MAE in natural products extraction started in the late 1980s, and through the technological developments, it has now become one of the popular and cost-effective extraction methods available today. Several advanced MAE instrumentations and methodologies have become available, e.g., pressurized microwave-assisted extraction (PMAE) and solvent-free microwave-assisted extraction (SFMAE).
  25. 25. SONICATION Sound waves of high frequency pulses of 20khz are generated in an ultrasonic bath Samples are kept in suitable vessel with solvent Waves are generated from transmitters. Thus increase cell wall permeability and cause solubilization of maximum amount of active constituents The frequency of a sound wave is a measure of how often the particles of a substance vibrate Sonicators either produce sound waves into a water bath, where samples are placed, or can be probes that are put directly into the sample to be sonicated. SONICATION Sound waves of high frequency pulses of 20khz are generated in an ultrasonic bath Samples are kept in suitable vessel with solvent Waves are generated from transmitters. Thus increase cell wall permeability and cause solubilization of maximum amount of active constituents The frequency of a sound wave is a measure of how often the particles of a substance vibrate Sonicators either produce sound waves into a water bath, where samples are placed, or can be probes that are put directly into the sample to be sonicated.
  26. 26. SUBLIMATION Some substances on heating/cooling changes from solid to gas or vice versa Plant material is warmed and vapour allowed come in contact with cold surface The sublimating substance vaporize from plant material and condensed as solid on cold surface . SUBLIMATION Some substances on heating/cooling changes from solid to gas or vice versa Plant material is warmed and vapour allowed come in contact with cold surface The sublimating substance vaporize from plant material and condensed as solid on cold surface .
  27. 27. COUNTER CURRENT EXTRACTION Liquid-liquid extraction-partition The apparatus consists of series of tubes with one lower stationary phase and upper immiscible liquids in moving mobile phase. Mobile phase moves from one tube to other Extract is placed in first tube and apparatus agitated, and layers are allowed to separate The components of mixture will distilled between the two layers according to their partition coefficients The upper moving phase is moved along to the second tube contains lower phase. COUNTER CURRENT EXTRACTION Liquid-liquid extraction-partition The apparatus consists of series of tubes with one lower stationary phase and upper immiscible liquids in moving mobile phase. Mobile phase moves from one tube to other Extract is placed in first tube and apparatus agitated, and layers are allowed to separate The components of mixture will distilled between the two layers according to their partition coefficients The upper moving phase is moved along to the second tube contains lower phase.
  28. 28. SUPERCRITICAL FLUID EXTRACTION SCF A supercritical fluid SCF is any substance at a temperature and pressure above its critical point, where distinct liquid and gas phases do not exist. It can effuse through solids like a gas, and dissolve materials like a liquid. In addition, close to the critical point, small changes in pressure or temperature result in large changes in density, Supercritical extraction and fractionation of natural matter is one of the early and most studied applications in the field of supercritical fluids. supercritical Fluid Extraction (SFE) is the process of separating one component (the extractant) from another (the matrix) using supercritical fluids as the extracting solvent. A supercritical fluid SCF is any substance at a temperature and pressure above its critical point, where distinct liquid and gas phases do not exist. It can effuse through solids like a gas, and dissolve materials like a liquid. In addition, close to the critical point, small changes in pressure or temperature result in large changes in density, Supercritical extraction and fractionation of natural matter is one of the early and most studied applications in the field of supercritical fluids. supercritical Fluid Extraction (SFE) is the process of separating one component (the extractant) from another (the matrix) using supercritical fluids as the extracting solvent.
  29. 29. Extraction is usually from a solid matrix, but can also be from liquids. SFE can be used as a sample preparation step for analytical purposes, Supercritical fluid chromatography (SFC) is a hybrid of gas and liquid chromatography that combines some of the best features of each. Supercritical fluid chromatography is of importance because it permits the separation and determination of a group of compounds that are not conveniently handled by either gas liquid or liquid chromatography. SFC has been applied to a wide variety of including natural products, drugs, foods, pesticides and herbicides, surfactants, polymers and polymer additives. Extraction is usually from a solid matrix, but can also be from liquids. SFE can be used as a sample preparation step for analytical purposes, Supercritical fluid chromatography (SFC) is a hybrid of gas and liquid chromatography that combines some of the best features of each. Supercritical fluid chromatography is of importance because it permits the separation and determination of a group of compounds that are not conveniently handled by either gas liquid or liquid chromatography. SFC has been applied to a wide variety of including natural products, drugs, foods, pesticides and herbicides, surfactants, polymers and polymer additives.
  30. 30. METHODS OF PURIFICATIONMETHODS OF PURIFICATION 1.CHROMATOGRAPHY 2.DISTILLATION 3.EVAPARATION 4.CRYSTALISATION 5.SUBLIMATION 6.PRECEPTATION 1.CHROMATOGRAPHY 2.DISTILLATION 3.EVAPARATION 4.CRYSTALISATION 5.SUBLIMATION 6.PRECEPTATION PHYSICALPHYSICAL CHEMICALCHEMICAL 1.HYDROLISIS , 2.SALT FORMATION 3.ACETYLATION 1.HYDROLISIS , 2.SALT FORMATION 3.ACETYLATION
  31. 31. SENNOSIDES FROM SENNA Sennosides are obtained from dried leaflets of cassia angustifolia belonging to the family Leguminosae Sennosides are of A,B,C,D.Senna shows laxative property. Extraction: Senna leaves are dried and powdered to coarse powdered senna is extracted with benzene in electric shaker 2 hrs Filter the extract and dried with 70% methanol for 5hrs and again extract marc with fresh methanol for 2 hrs Filter and combine with extracts and concentrate up to 1/4 th of its
  32. 32. Add sufficient quantity of HCL to make pH 3.2 and keep a side for 2 hrs at 5°C Filter and add sufficient quantity of alcoholic anhydrous calcium chloride with continuous stirring Add ammonia to bring pH 8 and kept a side for 2hrs filter and collect ppt of calcium Sennosides Dry the ppt in desiccators and calculate percentage yield
  33. 33. Chemical tests: 1.Borntragers test: Take a little quantity of a solution of sample +water +CCL4 or ether Separate organic layer and shake with dilute ammonia Rose pink color of ammonia layer appears . 2.TLC/HPTLC: a)Stationary phase silica gel G b)Mobile phase :n-proponol: ethyl acetate :water: glacial acetic acid (3:3:2:0:1 v/v) c)Detection:366 nm,RF -0.35,0.25,0.61
  34. 34. 3.HPLC: a.Method : isocratic b.Stationary phase:C18 c.Mobile phase :ethanol:water:acetic acid:tetrahydrofuron. (60:38:2:2). d. Detection:25μ nm.
  35. 35. EXTRACTION OF QUININE AND QUINIDINE Quinine,quinidine,cichonine and cinchonoidine are the popular quinoline alkaloids obtained from dried bark of cinchona officinalis belonging to the family Rubiaceae. All these have anti-malarial properties. Quinine also shows anti- arrhythmatic agent. Extraction: Moist the weighed cinchona bark powder with alcoholic calcium hydroxide or calcium oxide or potassium hydroxide (20%) And sodium hydroxide (5%)keep the mixture as it is for few hours so that alkali can convert cinchona alkaloids to free bases
  36. 36. Being non-polar in nature these free bases can be extracted with benzene for 6 hrs in Soxhlet extractor. After complete extraction filter the benzene extract and add 5% sulphuric acid. Shake the above contents mixture in separating funnel, separate aq.layer from benzene layer and discharge benzene layer adjust pH 6.5 with sodium hydroxide. Precipitate of quinine sulphate are formed filter and recrystalize with hot water Weigh and determine percentage yield and melting point Dissolve this quinine sulphate dilute suphuric acid, add ammonia to crystallize quinine as sharp needles separate these crystals from mother
  37. 37. Identification tests: To the little quantity of sample and 0.25 ml of bromine water ,mix well and add 1 m.l of chloroform, allow to stand few min,add 10%potassium ferrocyanide,shake well and add 3 m.l of 5N NaoH . Quinidine :chloroform layer becomes red. Quinine:chlorofom layer remains color less. TLC/HPLC: Stationary phase:silicagel-G Mobile phase :diethylamine:ether:toulene(10:24:40) Detection :spray with iodoplatinate reagent.
  38. 38. HPLC: Method:Isocretic Stationary phase:C18 Mobile phase: methanol 1 acetonitrile-0.1mol/L:ammonia acetate(45:15:40:v/v) at a flow rate 1.0 ml/min Detection:flurscence at Excitation-325 nm Emission-375 nm
  39. 39. GLYCYRRHIZIN FROM GLYCYRRHIZA Glycyrrhizine is a pentacyclic tri- terpenoid a-glycone obtained from the roots and stolons of glycyrrhiza glabra belonging to the family Leguminosae Liquorice used as anti-ulcer. Extraction : Take accurately weighed quantity of coarse powder of glycyrrhiza roots Extract the powder with chloroform Filter and discharge the filtrate Extract the marc with 0.5 M H2SO4 for few hours
  40. 40. Filter and extract the filtrate with three portions of chloroform Separate and combine the chloroform layers Distill of the chloroform extract to yield a dry residue of glycerrhetinic acid Identification: 1.Libermann test:mix 2 ml of test solution with 2 ml acetic anhydride boil and add 0.5 ml H2SO4.Blue color appears. 2.TLC: a)Stationary phase: silica gel G b)Mobile phase: ethyl acetate c)Formic acid: glacial acetic acid (15:1:2) butanol-glacial acetic acid- water (7:1:2)
  41. 41. LYCOPENE FROM TOMATO Lycopene is a pigment principally responsible for the characteristic deep red color of ripe tomato fruits. it is a natural anti- oxidant,lyccopene is a carotonoid which also shows anti-cancer activity. Lycopene fruits are the ripen fruits of lycopersicum esculentum belonging to the family solanaceae. Extraction : 50 gm of tomato paste was dehydrated by adding 65 ml methanol After 24 hrs thick suspension was filtered The dark red cake was shaken for another 15 min with 75 ml mixture of equal volume of methanol and carbon tetrachloride and separated by filtration
  42. 42. The ccl4 phase transferred to a reparatory funnel add volume of water and shaked well after phase separation ccl4 phase evaporates And residue was diluted with 2 ml of benzene using a dropper,1 ml of boiling methanol was added in portion, then crystals as crude Lycopene were appeared immediately and crystallization was completed by keeping the liquid at room temperature and ice bath The crystals were washed 10 times using benzene and boiling methanol Long red Lycopene prisms were observed under microscope with some colorless impurities
  43. 43. For more purification column chromatography on active acidic alumina using toluene as eluent was done The deep red zone was collected after complete evaporation of solvent the residue was dissolves in 2 ml benzene After re-crystallization using boiling methanol no colorless substance observed Crystalline Lycopene is not isomerized but has a tendency to auto- oxidation, so it was kept in dark glass tube.
  44. 44. Chemical tests: 1.Lycopene dissolves in concentrated H2SO4—indigi blue color appears. 2.On addition of anti- mony chloride in chloroform to a solution of Lycopene in chloroform on intense unstable blue color produces. Identification: UV-wave length—447.2 ,473.2,504.2 nm IR- vibrational wave length 3100 cm-1 2918.92-2851.05 sp3 1670,1640-c=c(trans)
  45. 45. ISOLATION OF CURCUMIN FROM TURMERIC Turmeric of zingiberaceae family wisely cultivated spice in India and others Asian countries. Turmeric is rich in curcuminoids,curcuminnoids shows anti- oxidant,anti-inflammatory,ant-carcinogenic effects, and Hypoglycemic activity. Extraction: Fresh rhizomes were collected washed with deionised water Sliced and dried in sun for one week and again dried at 50°c in hot air oven for 6hrs Dried rhizomes were cut in to small pieces powdered by electric mill Take 20gm of sample in a thimble and placed in Soxhlet apparatus were set up to various solvent from non-polar to polar
  46. 46. 150 ml of solvent added and extracted 6 hrs hexane,B.P 69°C,chloroform 61°C,ethylacetate 77°C,methanol 65°C,acetone56°C,one with hexane 2 hrs Hexane extract was discarded and the powder re extracts with methanol 6 hrs After completion of extraction dark brown extract was then cooled, filtered ,concentrate, using rotatory evaporator finally by vaccum suction to get a crude extract yield calculated.
  47. 47. Identification: Curcumin orange yellow crystalline powder with M.P 183°C,insoluble in cold water, soluble in alcohol, glacial acetic acid, dissolves in H2SO4 yields red color in 0.1N NaoH . 1.TLC: a)Stationary phase:Silicagel-60F254 b)Mobile phase:chloroform:methanol (95:5) Benzene:ethylacetate:18:2 UV:420 nm.
  48. 48. THANK YOU

×