Pharmacognosy & Phytochemistry (Practical)


A Avinash


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Course Name :

Pharmacognosy & Phytochemistry (Practical)


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Course Objectives

The main purpose of subject is to impart the students the knowledge of how the secondary metabolites are produced in the crude drugs, how to isolate and identify and produce them industrially. Also this subject involves the study of producing the plants and phytochemicals through plant tissue culture, drug interactions and basic principles of traditional system of medicine

Learning Outcomes

To know the modern extraction techniques, characterization and identification of the herbal drugs and phytoconstituents To understand the preparation and development of herbal formulation. To understand the herbal drug interactions To carryout isolation and identification of phytoconstituents

Course Syllabus

Morphology, histology and powder characteristics & extraction & detection of: Cinchona, Cinnamon, Senna, Clove, Ephedra, Fennel and Coriander 2. Exercise involving isolation & detection of active principles a. Caffeine - from tea dust. b. Diosgenin from Dioscorea c. Atropine from Belladonna d. Sennosides from Senna 3. Separation of sugars by Paper chromatography 4. TLC of herbal extract 5. Distillation of volatile oils and detection of phytoconstitutents by TLC 6. Analysis of crude drugs by chemical tests: (i) Asafoetida (ii) Benzoin (iii) Colophony (iv) Aloes (v) Myrrh

Session Plan 1

Experiment 1

To study about the powder microscopy of fennel.

Session 1


The transverse section of mericarp region of fennel shows two prominent surfaces, the dorsal and the commissural surface. The commisural surface has a carpophore and two vittae, and the dorsal surface has a total of five ridges. The mericarp is divided into pericarp, consisting of the epicarp and mesocarp; the testa and the endocarp. Epicarp consists of polygonal cells of epidermis which are tangentially elon-gated and covered by the cuticle. Mesocarp has parenchyma cells with five bicollateral vascular bundles; below each primary ridge a lignified reticulate parenchyma surrounds the vascular bundles. There are four vittae on dorsal surface and two vittae on commisural or the ventral surface. Inner Epidermis or Endocarp shows parquetry arrangement (a group of four to five cells arranged parallelly at acute angles with groups of similar cells in different direction). Testa is a single-layered tangentially elongated cell with yellowish colour. Endosperm consists of thick-walled, wide polyhedral, colourless cells. Cells contain fixed oil, aleurone grains, and rosette crystals of calcium oxalate.

Session 2

Experiment 2

To study about the powder microscopy of coriander.


The transverse section of coriander shows the presence of a dorsal surface and a commissural surface. The dorsal surface consists of two vittae and a carpophore. The dorsal surface has five primary ridges and four secondary ridges. The epicarp consists of a single row of small thick-walled cells with calcium oxalate crystals. The mesocarp has an outer loosely arranged tangentially elongated parenchyma cells and the middle layer consisting of sclerenchyma. The middle layer is again divided into; the outer region of sclerenchyma is represented by longitudinally running fibres, whereas the inner region has tangentionally running fibres. The vascular bundles are present below the primary ridges. The inner layer has polygonal, irregularly arranged parenchyma cells. The endocarp has the parquetry arrangement. In the testa it has single-layered, yellowish cells, and the endosperm is thick, polygonal, colourless parenchyma with fixed oil and aleurone grains.

Session 3

Experiment 3

Isolation of caffeine from tea.



The technique used to separate an organic compound from a mixture of compounds is called Extraction. Extraction process selectively dissolves one or more of the mixture compounds into a suitable solvent. The solution of these dissolved compounds is referred to as the Extract. Here the organic solvent dichloromethane is used to extract caffeine from an aqueous extract of tea leaves because caffeine is more soluble in dichloromethane (140 mg/ml) than it is in water (22 mg/ml). However, the tannins that are slightly soluble in dichloromethane can be eliminated by converting it to their salts (phenolic anions by adding sodium carbonate) (tannins are phenolic compounds of high molecular weight and being acidic in nature can be converted to salts by deprotonation of the -OH group) which remain in the water.

Bevarages cover a vast variety of addictive drinks out of which Tea and Coffee are the most popular acceptable drinks. Tea powder is extracted from tea leaves which contain tannins, which are acidic in nature,  a number of colored compounds and a small amount of unrecompensed chlorophyll and an important stimulant called Caffeine. Because of the presence of Caffeine, tea and coffee are gaining popularity as an addictive stimulant. An average 30g of tea can contain 20-ll0 mg of caffeine thereby making  tea  a significant source of caffeine compared to other beverages. Caffeine can stimulate nervous system and can cause relaxation of respiratory and cardiac muscles.  Caffeine is well known to increase both the alertness level and attention span. But like all other addictives, tea also shows withdrawal symptoms like headache, nervousness and insomnia for a regular consuming person.

Caffeine, 1,3,7 - trimethylxanthine, belongs to a wide  class of compounds known as alkaloids. These are plant derived compounds with complex structure containing nitrogen, and usually have roles in physiological activity. The melting point of Caffeine is 238°C.




Extraction is a method used for the separation of organic compound from a mixture of compound. This technique selectively dissolves one or more compounds into an appropriate solvent. The solution of these dissolved compounds is referred to as the extract. In the case of Caffeine extraction from tea powder, the solubility of caffeine in water is 22mg/ml at 25°C, 180mg/ml at 80°C, and 670mg/ml at 100°C. Here the organic solvent Dichloromethane is used to extract caffeine from aqueous extract of tea powder because caffeine is more soluble in dichloromethane (140mg/ml) than it is in water (22mg/ml).The dichloromethane - caffeine mixture can then be separated on the basis of the different densities of dichloromethane and water because dichloromethane is much denser than water and insoluble in it. Residual water is separated from dichloromethane by drain out the dichloromethane through separating funnel, thus dichloromethane passed through the funnel while polar solvents such as water is still remains in the funnel. Water and dichloromethane is slightly soluble in each other. So, after separating the solvents, residual water will remain the organic layer. Mainly anhydrous sodium sulfite is used for the removal of water from organic layer. Anhydrous sodium sulfite is an insoluble inorganic solid which will absorb water, thus drying it.


Session 4

Experiment 4

Isolation of calcium sennoside from senna.

Session 5

Experiment 5

Isolation of curcumin from turmeric.

Session 6

Experiment 6

TLC of Turmeric.

Session 7

Experiment 7

TLC of Caffine.

Session 8

Experiment 8

Separation of sugars by Paper chromatography

Session 9

Experiment 9

Isolation of oil from lemon grass .

Session 10

Experiment 10

TLC of sennoside from senna.

Session 11

Experiment 11

Isolation of Atropine from Belladonna.

Session 8

Experiment 12

Chemical test of aloe


Chemical Tests 

(a) General Tests: For this prepare a 0.1% (w/v) aqueous solution of aloes by gentle heating, add to it 0.5g of Kiesulgur and filter through. Whatman Filter Paper No. 42 and preserve the filtrate for the following tests:

  1. Borax Test (or Schoenteten’s Reaction):To 5 ml of the above test solution add 0.2 g of pure borax and heat gently till it gets dissolved. Transfer a few drops of the resulting solution into a test tube filled with distilled water, the appearance of a green coloured fluoroscence due to the formation of aloe emodin anthranol shows its presence.
  2. Bromine Test:When equal volumes of the test solution and bromine solution are mixed together, it yields a pale-yellow precipitate due to the production of tetrabromaloin.
  3. Modified Borntrager’s Test:It is known that aloin (or barbaloin) belongs to the class of Cglycoside which does not undergo hydrolysis either by heating with dilute acid or alkali, but it may be decomposed with ferric chloride due to oxidative hydrolysis. Hence, the Modified Borntrager’s test employing FeCl3 and HCl is used as stated below:

First of all heat together 0.1 g of powdered aloe with about 2 ml of FeCl3 solution(5% w/v) and 2 ml of dilute HCl (6N) in a test tube over a pre-heated water bath for 5 minutes. Cool the contents and extract the liberated anthraquinone with carbon tetrachloride. Now carefully separate the lower layer of CCl4 and add to it ammonia solution. The appearance of a rose-pink to cherry red colour confirms its presence.


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    A Avinash

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    A.Avinash working as Assistant Professor, Dept of CSE, Centurion University of Technology and Management, Andhra Pradesh . Interested to work on Machine learning,Natural Language Processing,Problem Solving Methodologies , and ChatBot. Programming Skill: C Programming Data Structure Object Oriented Programming using C++ Formal Language Automata Theory Python Web Development(HTML,CSS,PHP) Database Management Systems Compilers