UNIT 2.3 METHODS IN NATURAL PRODUCTS CHEMISTRY

Identification of plant material and its active compound Extraction (eg: cold extraction, Soxhlet extraction) Isolation and purification (eg: partitioning, ion-exchange chromatography) Structural elucidation (eg: Mass spectrometry, NMR spectroscopy)

IDENTIFICATION OF BIOLOGICALLY ACTIVE MATERIALS Identity can be achieved by macro- and microscopical examinations. Voucher specimens are reliable reference sources. Outbreaks of diseases among plants may result in changes to the physical appearance of the plant and lead to incorrect identification. At times an incorrect botanical quality with respect to the labeling can be a problem.

Drug may be collected from: 1 - Wild plants. 2 - Cultivated plants. Disadvantage Advantage Scattered in large or unlimited area Present in limited area Difficult to reach Easy to reach The collector must be highly skilled botanists The collector must not be skillful person Deficiency may occur due to continuous collection Continuous supply

The following precautions should be considered during stage of collection: 1.The proper time of the day, time of the year and maturity stage of collection is particularly important because the nature and quantity of constituents may vary greatly in some species according to the season and time of collection 2.The collected plant should be free from any contamination. 3.Collecting plants which are free from diseases (i.e. which are not affected by viral, bacterial, fungal infection).

This may be confirmed by: Authentication of plant material : This may be confirmed by: 1. Establishing the identity by a taxonomy experts. 2.Collection of a common species in their expected habitat by a field botanist. 3. By comparing the collecting plant with a voucher specimen ( herbarium sheet).

3. Inhibit the growth of microorganisms. Drying of plant materials Aim of drying: 1. Ease of transport. 2. Ease of grinding 3. Inhibit the growth of microorganisms. 4. Preservative of active constituents. Drying is done in: Shade and in sunlight (Natural drying). Hot air drying or by freeze-drying (Artificial drying).

EXTRACTION

Extraction and fractionation of constituents : There is no general (universal) method for the extraction of plant materials. The precise mode of extraction depends on: 1- The texture of the plant material. 2- The water content of the plant material. 3- The type of substances to be extracted or nature of active constituents.

EXTRACTION DEFINITION: The separation of medicinally active portion of plants through the use of selective solvent and suitable methods extraction. The principal methods of extraction are: 1 - Macération 2 - Percolation 3 - Infusion 4 - Décoction

5 - Digestion 6 - Continuous hot extraction (Soxhlet extraction procèss). 7 - Liquid-liquid extraction 8 - Solvent-solvent ppt. 9 - Distillation

Definition An extract is a substance made by extracting a part of a raw material, often by using a solvent such as ethanol or water

Method: 1. Percolation This method uses percolator The powdered material is packed in the percolator Enough solvent is than poured to soak the powder Then more solvent is poured and allowed to percolate through the material The extract is collected in a receiver The process is continued until extraction is complete

Method: 2. Soxhlet extraction This is a continuous process of extraction with a hot solvent This extractor is provided with a siphoning system the powdered plant material is packed in a thimble The solvent is boiled in a flask

The evaporated solvent passes through the side tube of the extractor and condensed in the condenser, pitted at the top of the extractor The condensed hot solvent runs into the thimble and soaks the material and extracts the constituents When the chamber holding the thimble becomes full the solvent siphons down to the flask and the process is continuously repeated until complete

MACERATION : 1. In maceration powdered drug is soaked in organic solvent and kept for about 24 hrs , sometimes 3-4 days also, depending upon the part of the plant to be extracted. 2. The solvent is decanted, filtered and concentrated.

DECOCTION : In case of decoction , the powdered plant material is boiled with the solvents .It is collected and filtered. Filtered solvent is concentrated. Eg. Preparation of tea.

Successive solvent extraction example : Petroleum ether (60-80) Benzene Chloroform Acetone Ethanol (95%) Chloroform water

ISOLATION AND PURIFICATION

Bio-assay - Measurement of the concentration or potency of a substance by its effect on living cells or tissues.

STRUCTURAL ELUCIDATION The process of determining the chemical structure of molecules. The following are common methods determining the chemical structure (structure elucidation) X-ray diffraction – crystals are available Proton NMR Carbon-13 NMR Mass spectrometry Infrared spectroscopy

WHAT IS STRUCTURAL ELUCIDATION? Structure elucidation is the process of determining the chemical structure of a compound. For organic compounds, it will often involve the use of nuclear magnetic resonance spectroscopy (NMR spectroscopy). Other characterization techniques include mass spectrometry, infrared spectroscopy and X-ray crystallography.

NUCLEAR MAGNETIC RESONANCE (NMR) Nuclear Magnetic Resonance (NMR) spectroscopy is an analytical chemistry technique used in quality control and research determining the content and purity of a sample for as well as its molecular structure.

INFRARED SPECTROSCOPY Infrared Spectroscopy is the analysis of infrared light interacting with a molecule. This can be analyzed in three ways by measuring absorption, emission and reflection. The main use of this technique is in organic and inorganic chemistry. Used by chemists to determine functional groups in molecules.

X-ray Crystallography X-ray crystallography is a tool used for identifying the atomic and molecular structure of a crystal. In which the crystalline atoms cause a beam of incident X-rays to diffract into many specific directions.

Mass Spectrometry A mass spectrometer can rapidly and accurately measure the molecular weight and quantities of many substances. This technique alone cannot distinguish between two or more substances with the same molecular weight.

1- Name the instruments used is isolation and purification LEARNING CHECK 1- Name the instruments used is isolation and purification 2- State the instruments used in structural elucidation. 3- State the properties of alkaloid. 4- State the difference between primary and secondary metabolites.

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