Phytoconstituents

1. INDUSTRIAL PRODUCTION,
ESTIMATION AND UTILIZATION OF
PHYTOCONSTITUENTS
Taxol, Vincristine & Vinblastine

2. TAXOL
Biological Source: Taxol is a diterpenoid obtained
from the bark of Pacific Yew Tree, Taxus brevifolia,
belonging to family Taxaceae.
2

3. TAXOL
⚫Production:
• Powdered bark extracted with
methanol, filtered & evaporated
1 to dryness.
2
• Partitioned with the mixture
of carbon tetrachloride &
water, filter & evaporated.
3
• Dried CCl4 fraction again
extracted with CCl4 : methanol,
evaporate to obtain crude taxol.
3

4. HPLC Method
Method: Isocratic
Detector: Photodiode array detector
Stationery Phase: C18 Column
Mobile Phase: 0.02 M Potassium dihydrogen phosphate (buffer solution) in
water (pH 4.5 adjusted with potassium hydroxide) and acetonitrile in the ratio
of 60:40 v/v.
Flow rate: 2ml/min.
Run time: 8.0 min
Detection: 230 nm.
Estimation:

5. TAXOL
⚫ Estimation:
HPTLC method
Mob phase- chloroform:methanol (7:3v/v)
Visualizing agent- vanillin sulphuric acid.
⚫ Utilization:
1. Treatment of ovarian, lung, bladder,
esophageal & other types of cancers.
2. Antiproliferative agent.
5

6. VINCRISTINE & VINBLASTINE
⚫ Biological Source: These
are Indole alkaloids
obtained from the dried
whole plant of
Catharanthus roseus or
Vinca rosea, belonging to
family- Apocynaceae.
6

8. ⚫ Production: Plant tissue culture technique.
A two stage fermentation procedure was used.
500 ml Erlenmeyer flasks containing 100 ml medium (0.3%) malt extract, (1.0%)
glucose, (0.3%) yeast extract and (0.5%) peptone) were inoculated with 7 days old
culture and incubated at 28°C on a rotary shaker (240 rpm) for 4–5 days, which was
used as seed culture.
10 ml seed culture was transferred to 500 ml Erlenmeyer flask containing 100 ml
production medium called as vinca medium-1 (Glucose: 3%, Succinic acid: 1%,
Sodium benzoate: 100 mg, Peptone: 1%, Magnesium sulphate: 3.6 mg, Biotin: 1
mg, Thiamine: 1 mg, Pyridoxal: 1 mg, Calcium pentothenate: 1 mg, Phosphate
buffer: 1 ml (pH 6.8), L-Tryptophan: 0.1%, Geranium oil: 0.05%.) which were
incubated at 28°C for 20 days as shake culture (II stage), after which it was
harvested and used for further study. 8

9. Culture filtrates and mycelia were separated with the help of muslin cloth and then
lyophilized.
Lyophilized culture filtrate was extracted using ethyl acetate as a solvent system.
The organic layer was separated from the aqueous layer using separating funnel.
The extraction was repeated thrice and the solvent was dried using anhydrous sodium
sulphate and concentrated under vacuum using rotavapour at 40°C in order to get
crude extract.
Estimation:
⚫ HPLC :
Mob phase- acetonitrile: 0.1 M phosphate buffer.
Wavelength- 254nm.

10. Estimation:
⚫ TLC:
Sample: A small amount of crude extract was dissolved in ethyl acetate.
Stationery Phase: silica gel-G (0.5 mm thickness)
Mobile Phase: Chloroform∶methanol (8∶2) as a solvent system.
Detection: Sprayed with ceric ammonium sulphate reagent.
Vinca alkaloids spots produced brilliant violet color as well as purple color.
Utilization:
1. In chemotherapy regimens
2. Childhood leukemia
3. Immunosuppressant
4. In Hodgkin’s disease