HPTLC Fingerprinting of Hemidesmus indicus roots as a Quality Control Parameter in Herbal Formulations
Bhalara Parita V, Savalia Vaibhavi B, Pandya Devang J* School of Pharmacy, RK University, Rajkot, India
Abstract : Hemidesmus indicus roots have Anti-inflammatory, Anti-microbial, Anti-acne, Antioxidant, Hepatoprotective and Anti-arthritic activity. Many of its formulations are available in the market. The present work focuses on developing a simple HPTLC fingerprint of Hemidesmus indicus roots. Methanolic extract of the roots was prepared by maceration. This extract was used to develop a suitable mobile phase for fingerprinting. After mobile phase development involving several pilot TLC, the mobile phase showing distinct spots in TLC was found to be Chloroform: Methanol: Ethyl acetate (13:1:2). It was further subjected to HPTLC fingerprinting where Rf and Area Under Curve were calculated. HPTLC fingerprinting showed9 peaks at 254nm and 4 peaks at 366nm. This work provides a simple technique for standardization and detection of adulteration of Hemidesmus indicus root formulations, many of which are available in the market, consumed by people for treatment of various disease conditions, and also investigated upon continuously considering its wide domain of pharmacological actions. Keywords : Anantmool, Hemidesmus indicus, HPTLC, Quality control, Sarivaa.
Introduction
Hemidesmus indicus(Family – Asclepiadaceae) roots are found commonly across India and are also known as Anantmool (Hindi), Sarivaa (Sanskrit) and Indian Sarsaparilla (English)1,2.According to traditional medicinal texts as well as modern research, they possess anti-venom, anti-inflammatory, anti-pyretic, antimicrobial, anti-acne, antioxidant, hepatoprotective, anti-leprotic, anti-diarrheal, anti-cancer, anti-ulcer, antihyperlipidemic, anti-diabetic, chemoprotective, radioprotective, renoprotective, and anti-arthritic activity1-32 . Many of its formulations are available in the market, including Dashmoolarishta, Manjisthadi Taila and Trifaladi Taila. The present work focuses on developing an HPTLC fingerprint of H. indicus roots.
Pandya Devang J et al /International Journal of ChemTech Research, 2018,11(09): 01-07.
DOI=
http://dx.doi.org/10.20902/IJCTR.2018.110901
Figure 1. Hemidesmus indicus roots
29-43
Experimental
Crude drug material
H. indicusroots were purchased from Sanjivani Aushadhalaya, Bhavnagar, in July 2015 and compared
[1-2]
with standard literature for authentication.
Extraction
50g dry powder of H. indicus roots was macerated with 100ml methanol for 24h at room temperature. Methanolic extract was filtered and evaporated on water bath at 50°C to obtain the dried extract.
Mobile phase development
Pilot TLC were developed for methanol extract using various mobile phases. After observing the pilot results, further TLC were developed by adding ammonia & ethyl acetate for removal of tailing. The mobile phases used were Chloroform : Methanol : Ethyl acetate 5:5:1 to 8:2:1, Chloroform : Methanol : Ammonia
HPTLC fingerprinting of methanolic extract was performed in Dept. of Pharmaceutical Sciences, Saurashtra University, using the mobile phase Chloroform: Methanol: Ethyl acetate (13:1:2), as it gave most appropriate TLC fingerprint, under thefollowing conditions…
Stationary phase:Silica gel 60 F 254 (E. Merck KGaA) Sample application: CAMAG Linomat 5 Detection: CAMAG TLC Scanner 3 Lamp:D2 & W Measurement type: Remission Measurement mode: Absorption Optical filter: Second order Data filtering: Savitsky-Golay 7
Four tracks of same extract at different concentrations were run for the HPTLC fingerprinting and scanned under visible light, UV 254nm and UV 366nm.
Results and Discussion
Nine peaks were detected at 254nm (Table 1, Fig. 2, 3) and four peaks were detected at 366nm (Table 2, Fig. 4, 5) upon HPTLC of methanolic extract of H. indicusroots using mobile phase Chloroform: Methanol: Ethyl Acetate (13:1:2).
Table 1. Rf& Area Under Curve of HPTLC of methanol extract at 254nm
Figure 2: HPTLC 2Ddensitometric superimposable chromatogram of methanol extract at 254nm (chloroform: methanol: ethyl acetate-13:1:2)
-0.10 0.00 0.10 0.20 0.30 0.40 0.50 0.60
0.70 [ Rf ] 0.90
Figure 3: HPTLC chromatogram of methanol extract at 254nm(chloroform: methanol: ethyl acetate13:1:2)
Table 2. Rf& Area Under Curve of HPTLC of methanol extract at 366nm
Peak
Rf
Area Under Curve
Area %
1
0.03
14221.9
87.85
2
0.11
701.3
4.33
3
0.13
470.2
2.90
4
0.79
796.0
4.92
Figure 4: HPTLC 2D Densitometric superimposable chromatogram of methanol extract at 366nm (chloroform: methanol: ethyl acetate-13:1:2)
Figure 5: HPTLC chromatogram of methanol extract at 366nm (chloroform: methanol: ethyl-acetate 13:1:2)
The present work can be helpful to herbal industry as an important standardization parameter of H. indicus roots, and especially its alcoholic formulations and extracts, since H. indicus roots are a part of several Ayurvedic and marketed herbal products, as they are indicated in a broad spectrum of diseases6. This work can be specifically useful for authentication of raw material of the roots and in detection of adulteration, which will ultimately benefit the people who consume H. indicus root formulations.
References
1. Kirtikar K. R. and Basu B. D., Indian Medicinal Plants Vol. IV, International Book Distributors, New Delhi, 2005, 277.
2. Khare C. P.,Indian medicinal plants: An illustrated dictionary, Springer, New York, 2007,758.
3. Anonymous, The Ayurvedic Pharmacopoeia of India, Part 1, Vol. I, 1st Edn, Government of India, Ministry of Health and Family Welfare, Dept. of ISM and Homoeopathy, New Delhi; 2001, 107-8.
4. Gogte V. M., Ayurvedic Pharmacology and Therapeutic uses of Medicinal Plants, 1st Edn. Mumbai, Bhartiya Vidhya Bhavan, 2000, 512-3.
5. Gupta N. S., The Ayurvedic System of Indian Medicine, Vol. I, New Delhi, Bharatiya Kala Prakashan, 2006, 96-7.
6. Austin A., A review on Indian Sarsaparilla, Hemidesmus indicus (L.) R. Br., J Biol Sci, 2008, 8(1):1-12.
7. Aneja V., Suthar A., Verma S., Kalkunte S., Phyto-pharmacology of Hemidesmus indicus, Pharmacog Rev, 2008, 2(3):143-50.
8. Ravishankara M. N., Shrivastava N., Padh H., Rajani M., Evaluation of antioxidant properties of root bark of Hemidesmus indicus R. Br. (Anantmul), Phytomed, 2002, 9(1):153–60.
9. Nadana S., Namasivayam N.,Impact of Hemidesmus indicus R.Br. extract on ethanol-mediatedoxidative damage in rat kidney,Redox Report,2007, 12(5):229-35.
10. Kumar G. S., Jayaveera K. N., Kumar Ashok C. K., et al., Evaluation of antioxidant and antiacne properties of terpenoidal fraction of Hemidesmus indicus (Indian sarsaparilla), The Internet Journal of Aesthetic and Antiaging Medicine, 2008; 1 (1).
11. Mahalingam G., Krishnan K., Hypoglycemic activity of Hemidesmus indicus on streptozotocin induced diabetic rats, Int J Diab Dev Ctries, 2008, 28(1):6-10.
12. Mookan P., Rangasamy A., Thiruvengadam D., Protective effect of Hemidesmus indicus against rifampicin and isoniazid-induced hepatotoxicity in rats, Fitoterapia, 2000, 71:55-9.
13. Baheti J. R., Goyal R. K., Shah G. B., Hepatoprotective activity of Hemidesmus indicus R. br. in rats., Indian J. Exp. Biol., 2006, 44(5):399-402.
14. Hiremath S. P., Rudresh K., Badami S., Antimicrobial activity of various extracts of Striga sulphurea and Hemidesmus indicus, Indian J. Pharm. Sci., 1997, 59(3):145-7.
15. Das S., Devaraj S. N., Antidiarrhoeal effect of methanolic extract of Hemidesmus indicus an invitro and invivo study, Indian J Exp Biol, 2003, 41(4):363-6.
16. Das S., Devaraj S. N., Antienterobacterial activity of Hemidesmus indicus R. Br. Root extrac, Phytother Res, 2006, 20(5):416-21.
17. Das S., Devaraj S. N., Glycosides Derived from Hemidesmus indicus R. Br. root inhibit adherence of Salmonella typhimurium to Host Cells: Receptor Mimicry, Phytother Res, 2006, 20:784–93.
18. Gopiesh Khanna V., Kannabiran K., Larvicidal effect of Hemidesmus indicus, Gymnema sylvestre and Eclipta prostrata against Culex qinquifaciatus mosquito larvae, Afr J Biotechnol, 2007, 6(3):307-11.
19. Kumar G. S., Jayaveera K. N., Ashok Kumar C. K., et al, Antimicrobial effects of Indian medicinal plants against acne-inducing bacteria, Trop J Pharm Res, 2007, 6(2):717-23.
20. Iddamaldeniya S. S., Thabrew M. I., Wickramasinghe S. M. D. N., Ratnatunge N. and Thammitiyagodage M. G., A long term investigation of antihepatocarcinogenesis potential of an indigenous medicine comprised of Nigella sativa, Hemidesmus indicus and Smilax glabra,J Carcinogenesis, 2006, 5(2):11.
21. Shetty T. K., Satav J. G., Nair C. K., Radiation protection of DNA and membrane in vitro by extract of Hemidesmus indicus, Phytother Res, 2005, 19(5):387-90.
22. Mary N. K., Achuthan C. R., Babu B. H., Padikkala J., In vitro antioxidant and antithrombotic activity of Hemidesmus indicus (L) R.Br, J Ethnopharmacol, 2003, 87:187–91.
23. Mary N. K., Achuthan C. R., Babu B. H., Padikkala J.,Antiatherogenic effect of Caps HT2, a herbal Ayurvedic medicine formulation, Phytomedicine, 2003, 10:474–82.
24. Bopanna K. N., Bhagyalakshmi N., Rathod S. P., Balaraman R., Kannan J., Cell culture derived Hemidesmus indicus in the prevention of hypercholesterolemia in normal and hyperlipidemic rats, Indian J Pharmacol, 1997, 29:105-9.
25. Anoop A., Jegadeesan M., Biochemical studies on the anti-ulcerogenic potential of Hemidesmus indicus R.Br. var. indicus, J Ethnopharmacol, 2003, 84:149-56.
26. Verma P. R., Joharapurkar A. A., Chatpalliwar V. A., Asnani A. J., Antinociceptive activity of alcoholic extract of Hemidesmus indicus R.Br. in mice, J Ethnopharmacol, 2005, 102:298–301.
27. Lakshman K., Shivaprasad H. N., Jaiprakash B., Mohan S., Anti-inflammatory and antipyretic activities of Hemidesmus indicus root extract, Afr J Trad CAM, 2006,3(1):90–4.
28. Kotnis M. S., Patel P., Menon S. N., Sane R. T., Renoprotective effect of Hemidesmus indicus, a herbal drug used in gentamicin-induced renal toxicity, Nephrology, 2004, 9(3):142-52.
29. Rex Jeya Rajkumar S., Sreeraj Kuruppilakathmanikandan, Muthukumar Nadar M.S.A., Qualitative Phytoconstituent Profile of Lobelia trigona Roxb extracts, International Journal of Pharm Tech Research, 2015, 8(10): 47-50.
30. Hemlata, Savita Upadhyay, Shailendra K. Saraf, Development and Validation of Stability Indicating RP-HPLC Method for Simultaneous Estimation of NSAIDS-Antiulcer Agent Combination, International Journal of Pharm Tech Research, 2016, 9(8): 288-300.
31. V.N. Kalpana, V. Devi Rajeswari, Phytochemical and Pharmacological investigation of an indigenous medicinal plant Leucas aspera, , International Journal of Pharm Tech Research, 2016, 9(8): 399-407.
32. Agus Kurniawan, Fadlina Chany Saputri, Rissyelly, Islamudin Ahmad, and Abdul Mun’im, Isolation of Angiotensin Converting Enzyme (ACE) Inhibitory Activity Quercetin from Peperomia pellucida, International Journal of Pharm Tech Research, 2016, 9(7): 115-21.
*****
Extra page not to be printed…
For your Research References requirements , Always log on to www.sphinxsai.com