Methods of Plant Extraction and Isolation

In this methods of plant extraction and isolation post we have briefly explained about different types of plant extraction solvents based on various criteria’s for methods of plant extraction.

Plant Extracts, as the term is used pharmaceutically, involves the separation of medicinally active portions of plant or animal tissues from the inactive or inert components by using selective solvents in standard extraction procedures.

The products so obtained from plants are relatively impure liquids, semisolids or powders intended only for oral or external use. The selection of the solvent is crucial for solvent extraction. Selectivity, solubility, cost and safety should be considered in selection of solvents.

Based on the law of similarity and intermiscibility (like dissolves like), solvents with a polarity value near to the polarity of the solute are likely to perform better and vice versa. Alcohols (EtOH and MeOH) are universal solvents in solvent extraction for phytochemical investigation.

Methods of Plant Extraction

Aqueous Extracts

Collected samples were oven-dried at 50ºC for 24 h. They were then lightly triturated in an electric blender and immersed in distilled water in order to obtain stock solutions in a concentration of 5% (w/v). The stock solutions were stored at 4ºC for 24 h in darkness for the solubilisation of active compounds. Subsequently, they were vacuum-filtered through a qualitative paper filter (no. 1; Whatman) to obtain a crude leaf solution of each species (crude extract) in this methods of plant extraction.

Petroleum Extracts

The samples were dried at room temperature for two weeks. The dried leaves (100 g) were powdered in a plant sample grinder at controlled temperature and extracted using petroleum ether in a Soxhlet extraction apparatus attached with a rotary vacuum evaporator. Solvents were removed using rotary vacuum evaporator at 175 mbar at a temperature ranging from 40°C to 60°C.

Chloroform Extracts

The samples were washed with running water. The samples were air-dried for 5 d and then dried in an oven at 40 °C until reaching their constant weight. Dried leaves were powdered using a blender. Leaves powder with amount of 10 g was extracted using chloroform (150 mL of each solvent) using Soxhlet apparatus. Crude extracts were evaporated to dryness using water bath at 60°C. The final extracts were weighed and stored at 4 °C for further use.

E-Acetate Extracts

The dried samples were finely powdered mechanically and 150gm powder extracted first with hexane (500 ml) to remove overall lipid content, then the residue was extracted with ethyl acetate (500ml) using soxhlet apparatus and then filtered by Whatman no.1 filter paper. The excess solvent was removed by a rotary evaporator at 50°C. Then dried crude extract 18.6 gm (12.4% yields) was stored in the refrigerator (below 5°C) prior to use.

Ethanol Extracts

Twenty gram of powdered plant material was kept in 200 mL conical flask and added 100 mL of ethanol. The mouth of the conical flask was covered with aluminium foil and kept in a reciprocating shaker for 24 h for continuous agitation at 150 rev/min for thorough mixing and also complete elucidation of active materials to dissolve in the respective solvent. Then, extract was filtered by using muslin cloth followed by Whatman no 1 filter paper and finally filtered by using vacuum and pressure pump. The solvent from the extract was removed by using rotary vacuum evaporator RE52 with the water bath temperature of 50°C.

n-hexane Extracts

The leaves were dried in a forced air convection oven at 45 °C until constant weight. After drying, they were powdered using mill and the powder was stored protected from light and moisture at 28°C until use. The extract was prepared in a Soxhlet apparatus using 100 g of the powdered leaves and 1 L of n-hexane. The solvent was evaporated at 75 rpm and 64.4 °C in a rotary-evaporator.

Diethyl Ether Extracts

The extracts were obtained by maceration of dry plant material (50 g) with diethyl ether for 48 h at room temperature with continuous agitation. The ratio used in the extraction process was (1 g sample with 12 mL of solvent). Extract solutions were filtered with filter paper and then collected and concentrated using a rotary evaporator at 35-55 °C

Dichloromethane Extracts

3 g of dried samples were soaked in 300 ml of dichloromethane. Afterward, the mixture was filtered using Whatman Paper No. 1 filter paper. The filtrates were evaporated using a rotor evaporator at 40-45º, 1-3 mbar of vacuum pressure and 40 rpm of rotation. Then, the crude extracts were stored at –20º until used.

Acetone Extracts

The sample was extracted in acetone by firstly adding 1 g of sample into 100 mL of solvent. The mixture was then put in an ultrasonic bath with preset conditions: temperature of 35°C, time of 30 min, and power of 150 W. Next, the extract was immediately cooled on ice to room temperature and then filtered using filter paper (Whatman, 11 μm pore size).

n-Butanol Extracts

The samples were washed in tap water, shade dried for 20 days and made into a fine powder of 40 mesh size using the laboratory mill. Following that, 100g of the powder was filled in the thimble and extracted using 500 ml of n-Butanol in soxhlet apparatus for 8 – 10 hours. The extract was filtered through Whatman No.1 filter paper The entire extract was concentrated to dryness using rotary flash evaporator under reduced pressure.

Methanol Extracts

The dried samples were coarsely powdered using a mortar. Then, this coarsely powdered plant was macerated in 80% methanol to obtain the hydro-alcoholic crude extract using Erlenmeyer flask for 3 days at room temperature. After 72 h, the filtrate was separated from the marc by using filter paper (Whatman No.1). Then the alcohol was allowed to evaporate from the filtrate with mild heating on dry oven to dry at 40 °C and then the concentrated extract was stored at 4°C.

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