Estimation of Rutin in Tephrosia purpurea by HPTLC Method

 

Praveen Patidar*, Sameer Singh, Darshan Dubey and Kamlesh Dashora

Institute of Pharmacy, Vikram University, Ujjain (M.P.).

 

ABSTRACT:

Tephrosia purpurea Linn (Family- Fabeaceae) having anti-hepatotoxic activity, CNS depressant activity and it also show anti-allergic activity. The plant Tephrosia purpurea contains rutin, quercetin, rotenoids, rotenone, tephrosin and lupeol. The powdered aerial part of Tephrosia purpuria was extracted with 95% ethanol by hot continuous extraction method. The prepared extracts were subjected to HPTLC analysis. The marker compounds were estimated by using pre-coated HPTLC aluminium silica gel 60 F254 plates (MERCK) and compared with standard. The methanolic solution of standard rutin 5 µl (1 mg/ ml) and Tephrosia purpuria extract 8 µl (10 mg/ ml) were applied as 7mm band width using CAMAG Linomat IV applicator. The Mobile phase is ethyl acetate: formic acid: glacial acetic acid: water (100:11:11:26). The detection was carried out at 366 nm. The amount of rutin was estimated by the comparing the peak area of standard and the same was present in the extracts. The content of rutin in Tephrosia purpurea extracts was found to be 1.40% w/w. This estimation technique is very much useful for the estimation of quercetin and rutin present in the various medicinal plants.

 

 

INTRODUCTION: ^[1]

Tephrosia purpurea Linn belonging to family- Fabeaceae having anti-hepatotoxic activity, CNS depressant activity and it also show anti-allergic activity. The plant Tephrosia purpurea contains rutin, quercetin, rotenoids, rotenone, tephrosin and lupeol. The aim of this study is to estimate rutin by HPTLC method which is present in Tephrosia purpurea ^[1-5].

 

EXPERIMENTAL METHOD:

Collection of plants

The plant material of Tephrosia purpurea was collected from Madurai district, Tamil Nadu. The collected plant materials was identified and authenticated by Botanical Survey of India (BSI), Coimbtore (TN).

 

Extraction

The powdered aerial parts of Tephrosia purpurea was extracted with 95 % ethanol by hot continuous soxhlet extraction method.

 

Method Development of HPTLC

Standard preparation

10 mg of standard rutin was dissolved in 10 ml of methanol and used for HPTLC quantification at concentration of 1mg/ml.

 

Sample Preparation (extracts)

100 mg of extract was taken and extracted with 10 ml of methanol and slightly warmed. This solution was filtered by whatman filter paper to get a clear solution used for HPTLC analysis at a concentration of 10 mg/ml.

 

Chromatographic Condition ^[6-7]

Stationary phase	:	Precoated Silica gel F 254 Plates (MERCK)
Mobile phase	:	Ethyl acetate: Formic acid: Glacial acetic Acid: water
		(100:  11:   11: 26)
Saturation	:	60 minute
Development chamber	:	CAMAG twin trough development chamber
Applicator	:	CAMAG Linomat IV applicator
Scanner	:	CAMAG Scanner III CATS (4.06), Switzerland
Mode of scanning	:	Absorption (deuterium)
Detection wavelength	:	365 nm
Volume applied standard	:	5 µl
Volume applied samples	:	8 µl each

 

Fig. 1 HPTLC chromatogram of Standard Rutin

 

Fig. 2 HPTLC chromatogram of Tephrosia purpurea extract

RESULT AND DISCUSSION:

Tephrosia purpurea extract

Totally 4 peak observed in the Tephrosia extract, in this second peak with R_f value of 0.29 is matching with standard rutin with R_f value of 0.29. The amount of rutin was estimated by comparing the peak area of standard and the same was present in the sample. The content of rutin was found to be 1.40 % w/w (Fig. 1 and 2).

 

CONCLUSION:

Quantification of rutin in Tephrosia purpurea by HPTLC is very much useful for the estimation of flavonoidal compounds, like rutin present in the various medicinal plant materials, extracts and their formulations. 

 

REFERENCES:

1.       V.U. Ahmad, Z. Ali, S.R. Hussaini, F. Iqbal, M. Zahid, M. Abbas, N. Saba, Flavonoids of Tephrosia purpurea. Fitoterapia. 70; 1999: 443-445.

2.       E Venkata Rao and N Ranga Raju, Two Flavonoids From Tephrosia purpurea. Phyrochemmy. 23(10); 1984: 2339-2342.

3.       A.S. Damre, A.B. Gokhale, A.S. Phadke, K.R. Kulkarni, M.N. Saraf, Studies on the mmunomodulatory activity of flavonoidal fraction of Tephrosia purpurea. Fitoterapia. 74; 2003: 257–261.

4.       Shenoy Smita, Shwetha K, Prabhu K, Maradi R, Bairy KL, Shanbhag T., Evaluation of anti inflammatory activity of Tephrosia purpurea in rats. Asian Pacific Journal of Tropical Medicine. 2010: 193-195.

5.       Ali Rifat Gulpinar, Ilkay Erdogan Orhan, Asuman Kan, Fatma Sezer Senol, Sadiye Ayse Celik, Murat Kartal, Estimation of in vitro neuroprotective properties and quantification of rutin and fatty acids in buckwheat (Fagopyrum esculentum Moench) cultivated in Turkey. Food Research International. 46; 2012: 536–543.

6.       Seth P. D. HPTLC. First edition, CBS Publishers and distributors, New Delhi, 1996.

7.        Wagnar H and Bladt S. Plant Drug Analysis. Second edition, Springer-Verlag Publication, Berlin, 1996. 

 

 

 

Received on 11.10.2012       Modified on 14.10.2012

Accepted on 24.10.2012      © RJPT All right reserved