The active constituents should be extracted and purified before applying the chemical test.

Preliminary chemical test for active constituentsQualitative test:

1- Test for glycosides and carbohydrate

2 -Test for cardiac glycosides

3 -Test for anthraquinone glycosides

4- Test for flavonoids

5 -Test for saponin

6 -Test for tannins7 -Test for alkaloids

The application of physical constants is applied to the active principles of drugs such as alkaloids, volatile oils, fixed oil..etc.1- Solubility: -We should determine solubility of A.C. in different solvents.-Usually expressed in the following form: 1-g is soluble in ….ml of water, … ml of alcohol, etc.

2- Specific gravity:- Particularly of the fats and volatile oils.-Weight of a given volume of liquid compared with the weight of equal volume of water at specific temperature and pressure in air.

S.G. is not absolutely constant………….why?Because other factors affected it:•maturity of drug from which v.o. is prepared.•Age of v.o.•Method of preparation•PurificationE.g., Jalp is resin is used as purgative is heavier than water.

The specific gravity of anise oil is not less than 0.978 and not more than 0.988.Specific gravity of water =1Most of V.O. is lighter than water, except V.O. of clove, Cinnamon (heavier than water).

3- Optical rotation:Optical activity is the ability of a chiral molecule to rotate the plane of plane-polairsed lightdextro (+) rotary (rotate plane of polarized light to right( clock wise ) levo (-) rotary. rotate plane of polarized light to left ( anti-clock wise )

[α]TD = ± α x 100/LXC

[α]TD = ± value of α/degree of rotation

D: sodium line ≈ 580 mm (light that make rotation).L: Path length of tube in cmC: concentration in gm .

It is measured by polarimeter.

Atropine ± zeroPelocarpine +Iso pelocarpine -

1-Automatic Digital Polarimeter

2-Hand-Held Polarimeter

range ±45°. Polarimeter cell 100 mm, 200 mm and a sodium lamp.

wave length 589nm, measurement range -35°… +35° optical rotation

4- Refractive index:- Particularly of the fixed and volatile oils (crystals, liquids).It is a ratio of = sign angle of incidence/sign angle of refraction of monochromatic beam.RI measure by refractometer. -The RI of peppermint oil is not less than 1.4590 and not more than 1.4650 at 20 ◦C.

5- Congealing point:- Particularly of fixed and volatile oils.- The solidification range of fatty acids in olive oil is between 17 and 26◦C.

6- Melting point:It is of extreme importance, there is a constant range of m.p. using specific solvent. Cocaine m.p. 96-98 ◦C.7- UV:UV light is f importance in several cases, notably in detecting 1- adulterationa- of rhubarb, where rhapontic rhubarb can easily be distinguished from genuine Chinese or Indian rhubarb by its marked fluorescence. Chinese and Indian rhubarb fluorescence brown, while rhubarb rhaponticum is bright blue

b- powdered ergot exhibits a strong reddish fluorescence and can be detected in wheat flour.UV light is used for determining fluorescence and a play of colors in connection with certain extracts such as chlorophyll and the extracts of certain drugs (catechu, senna).

Many alkaloids show distinctive colors under this light, aconitine (light blue), berberine (yellow), emetine (orange).An alkaloids such as quinine show fluorescence in acid solution even in day light, exhibits a much stronger fluorescence under the quartz lamb

Before a new product is introduced on the market, it must be proven both safe and effective in preliminary pharmacologic tests on animals and in quantitative clinical tests on humans.

Bioassay: Estimation of potency of crude drugs or its preparation by study of its effect on living organisms like bacteria, fungal growth or animal tissue or entire animalBioassay is the measure of sample being tested capable of

producing biological effect as that of the standard

1 -Hepatoprotective activity Male\Female albino ratAlcohol, paracetamol, rifampicin, carbon tetrachloride Hepatocyte culture (Hepatic cells)

The parameter used to asses toxicity:

a) Physiological(Hexobarbital hypnosis)b) Biochemical serum estimation of enzymes like SGPT, SGOTc) Histopathological-(liver tissue necrosis and fatty degeneration )

2 -Hypoglycemic activity

Alloxan (180-250 mg\ml) or sterptozocin(induce diabetes )Rabbits, rats or miceHypoglycemic plant extracts like fenugreek Glucose tolerance effect or glucoautoanylsis RIA or ELISA are used for measurement of insulin levels

3 -Anti-inflammatory activityRat ,Albino adult mice Mechanical cause, infection or autoimmune disease Histamine, kinins and prostaglandin causing inflammationCarrageenan irritant substance Reduction of PAW oedema is measured

4 -Anti-ulcer activity

Anti-ulcerogenic plant like liqurice, white cabbage juice

Ulcer inducer:i- Chemicals ( alcohol, drugs)

Ii- stress (immobilization and cold)

Three group of animals1-treated with normal saline

2-treated with ulcerogenic 3 -tested group

4 -reference group

Microbiological activity

A- Cylinder plate method (cup method)Measurement of diameters of microbial growth inhibition surrounding cylinders containing various dilution of test compound and compared with reference

B- Turbidimetric methodBased on inhibition of microbial growth as indicated by measurement of turbidity(transmittance) of suspension of a suitable micro-organism in fluid medium to which have been added tested compoundChange in transmittance produced by tested compound are compared by known concentration of slandered