FORSKOLIN PRODUCTION, ESTIMATION AND UTILIZATION OF PHYTOCONSTITUENTS

FORSKOLIN

Synonym: Forskohlii, Plectranthusbarbalus.

Biological Source: Forskolin is a deterpenes, was obtained from the dried roots of Coleus Forskohlii.

Family: Labiatae.

Geographical Source: Plants grow perennially over the tropical & subtropical regions including India, Pakistan, Srilanka, Brazil and Tropical East Asia.

In India, it occurs at high altitude upto height of 2500m in Himalayas, in the regions from Garhwal, Kumaon to Nepal.

It also occurs in Deccan Peninsula and Parasnath Hills in Bihar.

Plant Habitat

Coleus is a branched, aromatic perennial herb1-2 feet in height leaves have camphor-like flavor with several fasciculate, succulent, radially spread roots about 1000 tonnes of leaves and roots are harvested every year from commercial cultivation in Gujrat.

Method of Cultivation: Vegetative method- Stem Cuttings.

Harvesting: In the month of autumn.

Chemical Constituents

Forskolin is a ditepenes, was first discovered at the Hoechst Research Centre, Mumbai, India and Central Drug Research Institute (CDRI), Lucknow, India. It is also containing various diterpenoid derivatives. The Leaves give a coleon E (Methylenequinone), barbatusin and coleon F. The root contains Coleonol B, Coleonol C, deoxycoleonol and labdane diterpenoids (I, II and III).

Identification test

TLC Method

Stationary Phase                     : Absorbent- Pre-coated Silica gel

Mobile Phase                           :Benzene: Ethyl acetate (85:15)  

Sample                                                : Root Extract

Solvent Form                          : 9cm²

Detecting agent                       : Anisaldehyde-H₂SO₄

Observation                             : Appearance of violet or purple color spots.

Chemistry

Chemically is named as 7β-Acetoxy-8, 13-epoxy-1ά,6β, 9 ά tritrydroxy labd-14-en-11-one.

Forskolin is a diterpenes.

Other active constituents are Coleonol D, E and F.

Coleonol& Forskolin are stereo isomers.

Molecular Formula      : C₂₂H₃₄O₇

Melting Point              : 230°C -232°C

Appearance                 : White crystalline powder.

Solubility                     : Soluble in Chloroform, Benzene and Dichloromethane, sparingly soluble petroleum ether.

Estimation

Forskolin is estimated by two methods:

1. Method 1: HPLC

2.      Method 2: HPTLC

Both the methods are accurate and reproducible

Method 1: HPLC (High Performance Liquid Chromatography)

Chromatographic System

HPLC equipped with i) LC 8A pump and ii) Photo array detector in combination with class LC10A software.

Chromatographic Conditions

Column                       : ODS (Octodecylsilane) C₁₈, 5Um, 250x4.6mm

Mobile Phase               : Acetonitrile: H₂O (50:50)

Wavelength                 : 220nm

Flow Rate                   : 1.6ml/min

Injection volume         : 20μl

Standard Preparation

Dissolve 10mg of forskolin with 15ml of acetonitrile in 25ml volumetric flask and makeup the volume up to 25ml with acetonitrile.

Sample Preparation

Dissolve 250mg of sample (equivalent to 20mg of Forskolin), with 25ml of acetonitrile. Filter the solution & makeup the volume up to 100ml with acetonitrile.

Procedure

Inject the standard and sample preparation (10μl) & record the chromatogram. Now calculate the percentage of Forskolin content from the peak areas.

Method 2: HPTLC (High Performance Thin Layer Chromatography)

Chromatographic System

HPTLC equipped with Densitometer, Applicator and Developing chamber.

Chromatographic Condition

Stationary Phase         : Absorbent- Silica gel 60 F254

Mobile Phase               : Benzene: Ethyl Acetate (85:15)

Solvent Form              : 9 cm²

Detection                    : 550nm, after Spraying C Vanillin H₂SO₄

Standard Preparation

i.           Dissolve 12.5mg of forskolin (Reference Standard) in 100ml volumetric flask with 50ml of CHCl₃& makeup the volume to 1000ml with CHCl₃.

ii.         From this solution, Pipette out 5ml of the solution and makeup the volume to 100ml with CHCl₃.

Sample Preparation

i.           Dissolve the 50mg of sample in 100ml volumetric flask using CHCL₃ as a solvent & makeup the volume to 100ml with the same solvent.

ii.         From this solution, pipette out 5ml of the solution & makeup the volume to 100ml with CHCl₃.

Procedure

i.           Set the applicator as per the condition.

ii.         By means of suitable syringe apply 5,10,15,20μl of standard preparation in 4 different tracks at 1cm height & 4mm band width with 7mm distance between the tracks.

iii.       Apply 5-10μl of sample preparation in another track.

iv.       Allow the plate to develop in the above-mentioned solvent system up to 9cm & remove it.

v.         Dry & Spray it with 10% of vanillin- H₂SO₄.

vi.       Heat the plate at 105°C for 10 minutes.

vii.     Record the Chromatograms of standard & sample.

viii.   Make the calibration curve with the concentration on the X-axis & area on Y-axis.

ix.       Calculate the content of Forskolin in the extract by extrapolation.

Utilization

Forskolin has Vasodilator and Cardio stimulant effects. Forskolin activates/ stimulates the adenylate cyclase, so it is considered as a promising drug for the treatment of glaucoma, congestivecardiomyopathy and asthma.

In Ayurvedic traditional medicine, coleus has been particularly given for the treatment of heart diseases, abdominal colic.

It is also used for the treatment of hypotensive, spasmolytic and platlet aggregation inhibitory activity.