Extraction and Isolation of Active Constituents PDF

Extraction and isolation of active constituents Dr. Mosa E. O. Ahmed 11/10/2024 Dr. Mosa E. O. Ahmed 1 Lecture contents This lecture will cover the following topics: Definition of extraction Steps involved in extraction E...

Extraction and isolation of active constituents Dr. Mosa E. O. Ahmed 11/10/2024 Dr. Mosa E. O. Ahmed 1 Lecture contents This lecture will cover the following topics: Definition of extraction Steps involved in extraction Extraction methods of the active plant constituents. Evaporation of the plant extracts Separation and isolation of the active plant constituents. 11/10/2024 Dr. Mosa E. O. Ahmed 2 Extraction of plant material: The term extraction is used pharmaceutically to indicate: The process of separating the medicinally active portions of plant or animal tissues from the inactive or inert components by using selective solvents in standard extraction procedures. 11/10/2024 Dr. Mosa E. O. Ahmed 3  Steps Involved in the Extraction of Medicinal Plants: In order to extract medicinal ingredients from plant material, the following sequential steps are involved: 1. Sample preparation 2. Size reduction 3. Extraction 4. Filtration 5. Concentration 6. Drying 11/10/2024 Dr. Mosa E. O. Ahmed 4  Preparation of plant samples Careful selection of the plant material to avoid the use of adulterated or infected samples. Careful authentication of the selected sample by taxonomic expert. Direct immersion of fresh material in boiling alcohol within few minutes of its collection to stop enzymatic activity. Drying of plant material when required before extraction, should be under controlled conditions. 11/10/2024 Dr. Mosa E. O. Ahmed 5 The instruments used in extraction and preparation of medicinal plants materials include: 1. Disc mill 2. Soxhlet extractor 3. Vacuum column 4. Freeze dryer 5. Rotary evaporator 11/10/2024 MOSA ELHADY Steps Involved in the Extraction of Medicinal Plants: Size reduction: - Hammer or disc mill - Cryogenic milling: new grinding method suitable for thermolabile cpds (volatile oils), provides large surface area and minor loss of cpds. Utilize dry ice, and liquid nitrogen for cooling (- 80 C). 11/10/2024 Dr. Mosa E. O. Ahmed 7 Selection of the solvent for extraction The Selectively dissolve the active material solvent or of choice is that Remove the inactive or inert material which to leave higher concentration of active will compounds in the plant samples. either: 11/10/2024 Dr. Mosa E. O. Ahmed 8 Factors influencing solubilization The most important factor is the polarity of the solvent and solute molecules. As general non-polar solvents will dissolve non-polar compounds such as fats empirical and waxes. rule: Polar solvents dissolve polar compounds such as sugars and alkaloidal salts. The use of solvent mixtures may increase the solubility of certain compounds. 11/10/2024 Dr. Mosa E. O. Ahmed 9 Extraction Processes 1. Infusion The processes like 2. Decoction infusion, decoction and digestion are rarely 3. Digestion used with few exceptions for extraction of drugs. 6. Distillation Maceration and percolation processes are of particular importance in most Pharmacopeias refer to the process of extraction of crude 11/10/2024 Dr. Mosa E. O. Ahmed drugs. 10 Maceration It is based on the immersion of the crude drug in bulk of the solvent menstruum. Repeated m aceration may be more efficient than a single m aceration, since an appreciable amount of active principle may be left behind in the first pressing of the marc. The repeated maceration is more efficient in cases where active constituents are more valuable. 11/10/2024 Dr. Mosa E. O. Ahmed 11 Infusions Infusions are dilute solutions Infusions are liable to fungus containing the readily-soluble and bacterial growth, and it is constituents of crude drugs. necessary to dispense them within 12 hours of their preparation. Formerly, fresh infusions, prepared by macerating the drug for a short period in cold water or boiling water were used. 11/10/2024 Dr. Mosa E. O. Ahmed 12 Infusion: m ethods of preparation The drug is usually coarsely Then add 900 ml of boiling powdered, very fine powder water, cover the vessel being avoided (50 gm). tightly. Moisten the drug in a Allow it to stand for 30 suitable vessel, provided with m inutes. a cover, with 50 ml of cold Then strain the mixture, pass water. enough water to make the Allow to stand for 15 infusion measure 1000 ml m inutes. 11/10/2024 Dr. Mosa E. O. Ahmed 13 Decoction Decoction was used for water After boiling, the liquid is cooled soluble and heat stable filtered, more water is passed constituents of hard and woody through the marc to produce the crude drugs. required volume. Water used as menstruum and At Present no decoction is drug cut in small pieces is Official in Pharmacopeias. actually boiled with the menstruum for the stated time, usually 10 -15 minutes. 11/10/2024 Dr. Mosa E. O. Ahmed 14 Digestion This process is a modified form of maceration in which the extraction of drugs is carried out by applying gentle heat to the substances being extracted. This method is applicable only to those drugs where moderately elevated temperature is not objectionable and the action of the menstruum is increased by gentle heat. 11/10/2024 Dr. Mosa E. O. Ahmed 15 Percolation Percolation is a continuous flow of the solvent through the bed of the crude drug material to get the extract. After imbibition, plant sample is placed in percolator and Sufficient menstruum is also added to maintain a small layer above the drug and allowed to stand for 24 hours. After maceration, the outlet is opened and solvent is percolated at a control rate with continuous addition of fresh volume. 11/10/2024 Dr. Mosa E. O. Ahmed 16 Continuous hot extraction (Soxhlet Extraction) Soxhlet extraction is used where It is used for those drugs small volume of hot menstruum where: is passed through the drug, over The penetration of the time and again to dissolve out menstruum into cellular the active constituents until the tissue is very slow. drug is exhausted. The solute is not readily soluble into the solvent. The quantity of menstruum is very less. 11/10/2024 Dr. Mosa E. O. Ahmed 17 Soxhlet extractor 11/10/2024 Dr. Mosa E. O. Ahmed 18 Distillation Is defined as the separation Distillation failed to isolate of the components of liquid component present in minor mixture by a process quantities and gas involving vaporization and chromatography is routinely subsequent condensation at used. another place. Steam distillation is used to Fractional distillation is used isolate volatile oils and traditionally to isolate the hydrocyanic acid from plant components of volatile oils. materials. 11/10/2024 Dr. Mosa E. O. Ahmed 19 Other m ethods for volatile oils extraction ColdFat Extraction (Enfleurage) Sponge expression Expression: Écuelle à piquer is a cold pressed method of Machine abrasion extraction, which is mostly used in the extraction of citrus essential oils. 11/10/2024 Dr. Mosa E. O. Ahmed 20 Evaporation of crude drug: Evaporation is one of the quality- relevant parameter of the eluate to the soft extract. It is made by cautious vacuum evaporation apparatus (rotary evaporator) and evaporation temperatures not exceeding 55 ̊ C. The temperature in correlation with the evaporation time is of special importance for quality of this step of manufacturing, if the extract contains easily volatile or thermo- labile constituents. 11/10/2024 Dr. Mosa E. O. Ahmed 21 11/10/2024 Dr. Mosa E. O. Ahmed 22 Separation and isolation of plant constituents Isolation and purification of plant constituents become the most difficult operation in phytochemical research. Fractionation techniques are required to separate the compounds belong to the same chemical class. 11/10/2024 Dr. Mosa E. O. Ahmed 23 Separation and isolation of plant constituents Purification of plants constituents is carried out using one or combination of more than one techniques: 1. Sublimation 2. Steam distillation 3. Fractional liberation 4. Fractional crystallization 5. Adsorption chromatography (TLC, GSC) 6. Partition chromatography (PC, GLC, HPLC) 7. Supercritical fluid chromatography 11/10/2024 Dr. Mosa E. O. Ahmed 24 Sublim ation It is sometimes be possible on the whole drug such as isolation of caffeine from tea. 11/10/2024 Dr. Mosa E. O. Ahmed 25 Distillation Fractional distillation is used traditionally to isolate the components of volatile oils. Distillation failed to isolate component present in minor quantities and gas chromatography is routinely used. Steam distillation is used to isolate volatile oils and hydrocyanic acid from plant materials. 11/10/2024 Dr. Mosa E. O. Ahmed 26 Fractional liberation Used to isolate alkaloid salts from aqueous solution by addition of aliquot of alkalis, will give first the weakest base in free state. Also used to isolate organic acids soluble in water immiscible solvent, starting with mixture of acids salts and then fractionated by addition of mineral acids. 11/10/2024 Dr. Mosa E. O. Ahmed 27 Fractional crystallization Utilize the differences in solubility of the components of a mixture in a given solvent. Generally, the derivative of a particular component is used such as picrates of alkaloids and osazones of sugars. 11/10/2024 Dr. Mosa E. O. Ahmed 28 Adsorption chrom atography The fundamental principles of adsorption chromatography is selective adsorption of a particular components on stationary phase. The adsorption chromatographic techniques is originated by Tswett. The use of charcoal for the decolorization and clarification of coloured solution is well known. This consist of two phase stationary phase (solid) and mobile phase (liquid). TLC is an example of adsorption chromatography. 11/10/2024 Dr. Mosa E. O. Ahmed 29 Partition chrom atography It was introduced by Martin and Synge for separation of acetylated amino acids. Separation based on the difference in partition coefficients of the components of a mixture between an aqueous and immiscible organic liquids. The stationary phase is aqueous liquid mixed with suitable carrier (silica gel). Paper chromatography and HPLC are examples for partition chromatography. 11/10/2024 Dr. Mosa E. O. Ahmed 30 Supercritical fluid chrom atography This technique use carbon dioxide as mobile phase. The low viscosity of super critical fluid provides faster flow rate than standard HPLC. This technology producing solvent residue free nutritional and healthcare products. The extract is absolutely solvent residual free as the CO2 is in gaseous state at room temperature. 11/10/2024 Dr. Mosa E. O. Ahmed 31 Supercritical fluid chrom atography According to the physicochemical properties of supercritical CO2, the extraction is carried out at moderate temperature (mainly between 31-60°C), therefore thermo-labile compounds can be obtained without any decomposition. 11/10/2024 Dr. Mosa E. O. Ahmed 32  The end 11/10/2024 Dr. Mosa E. O. Ahmed 33