INTRODUCTION:

Plant kingdom is one of the omportant sources of the natural product and food. In present era many life saving, nutritive chemical components are isolated from different plants due to both medicinal and economical values. In India, Ayurveda and Siddha provides major information about medicinal plants, but there are number of plnats which is not investigated thoroughly ⁽¹⁾. Medicinal plants have been used for centuries as remedy for human diseases because they contain the compounds of therapeutic values⁽²⁾. Infectious diseases are the leading cause of death worldwide. Many infectious diseases have been known to be treated with herbal remedies throughout the history of mankind. Natural products, as a pure compound or as a standardized plant extracts provide unlimited opportunities for new drug lead because of the unmatched availability of chemical diversity. An increasing failure of chemotherapeutics and antibiotic resistance exhibited by pathogenic and infectious agents has led to the screening of medicinal plants for their antimicrobial potential.

In recent years, secondary metabolites (phytochemicals) previously with unknown pharmacological activities have been extensively investigated as a source of medicinal plants. Thus, it is anticipated that the phytochemicals with adequate anti-infective efficacy will be used for the treatment of various infections caused by pathogens⁽³⁾. Therefore, there is a need to develop the efficient, safe and inexpensive drugs from plant source are of great importance.

Jatropha curcas, Linn., is an evergreen, large shrub, of 3-4 metre height, found throughout the waste places of India. It is commonly called as “Angular-leaved nut”, and in Tamil, “Kattamanakku”. The seeds are acro-narcotic; oil from the seeds are purgative internally and externally it is depurative and antiseptic; leaves are used as lactogogue; stem juice is haemostatic and root bark is stomachic astringent⁽⁴⁾. It is a multipurpose (both medicinal and biotechnologically important) plant and used as a source for the production of biodiesel⁽⁵⁾. In traditional medicine, it is used to cure various ailments ranging from simple fevers to infectious diseases including sexually transmitted diseases, syphilis, jaundice, mouth infections, guinea worm sores and rheumatism⁽⁶⁾. The leaf has been reported for the presence of β- sitosterol-3-O-β-D-glucopyranoside, 7-keto-β-sitosterol and β-sitosterol⁽⁷⁾.

MATERIALS AND METHODS:

Plant material

The leaves of Jatropha curcas, Linn., was collected from Perambalur, TamilNadu, India, in the month of September. The plant has been authenticated by the plant taxonomist, Dr. A. Tangavelou, Director, Bioscience Research Foundation, Pondicherry and the voucher specimen is procured in the department for future reference. Then, the collected plant materials were subjected to shade drying process and coarsely powdered.

Macroscopical studies

The dried leaves of Jatropha curcas, Linn., were subjected to macroscopical studies which includes organoleptic characters such as size, colour, odour, taste and shape.

Microscopical studies

The plant materials for the proposed study were collected from the Pondicherry vicinity, Pondicherry and extreme care was taken to select healthy plants and for normal organs. The leaf, stem and bark were separated out from the plant and fixed in FAA (Formalin 5ml + Acetic acid 5ml +70% Ethyl alcohol 90ml). After 24 hours of fixing, the specimens were dehydrated with graded series of tertiary-butyl alcohol (TBA) as per the schedule⁽⁸⁾. Infiltration of the specimens was carried out by gradual addition of paraffin wax (melting point 58°-60°c), until this TBA solution attained super saturation. Then, the specimens were castled into paraffin blocks.

Sectioning

The paraffin embedded specimens were sectioned with the help of a rotary microtome, 10-12µm thickness of sections was made. However, dew axing of the sections was done by using customary procedure⁽⁹⁾. The sections were later stained with Toluidine blue⁽¹⁰⁾. Since, Toluidine blue is a polychromatic stain. The staining was remarkably good and yielded varied cytochemical reactions. The dye rendered pink colour to the cellulose walls, blue to the lignified cells, dark green to sobering, violet to the mucilage, blue to the protein bodies etc., wherever necessary, sections were also stained with safranin, fast-green and iodine-potassium iodide (for starch). Cleared sections were then mounted in glycerin for microscopical observation.

Physico-chemical parameters

The various physico-chemical parameters such as total ash, water soluble ash, acid insoluble ash, sulphated ash, water soluble and alcohol soluble extractive value, foreign organic matter and moisture content were carried out^(11,
12).

Preliminary phytochemical screening

Ethanol and aqueous leaf extracts along with leaf powder were also subjected to qualitative examination for an identification of various phytoconstituents⁽¹³⁾.

Fluorescence analysis

The fluorescence analysis of the leaf powder⁽¹⁴⁾ with different chemical reagents and various leaf extracts⁽¹⁵⁾ were carried out by adopting the standard method. The behavior of the samples with different chemical reagents was observed under day blight and UV light at 254 nm.

Antibacterial screening

25 and 50 mg/ml of ethanol leaf extracts of Jatropha curcas, Linn., was subjected to antibacterial screening against microorganisms such as Escherichia coli, Staphylococcus aureus, Pseudomonas aeroginosa, Klebsiella pneumoniae and Micrococcus luteus by disc plate method ⁽¹⁶⁾ using Ofloxacin as a standard (10 µg/ml).

RESULTS AND DISCUSSION:

Macroscopical studies

Jatropha currcas, Linn., is a large shrub, grown up to 3-4 meter height, abundantly available throughout India. Leaves are 5-7 lobed, broadly ovate, glabrous and are alternately arranged. Flowers in loose panicles of the cymes, yellowish green, fruits are 2.5 cm long, ovoid, black; seeds are ovoid-oblong, dull brownish black.

Figure 1: T.S.of Midrib.

1-Upper epidermis; 2-Palisade layer; 3-Spongy parenchyma;

4-Stomata; 5-Protoxylem; 6-Metaxylem; 7-Bundle sheath;

8-Lacticiferous cells; 9-Cortex; 10-Lower epidermis.

Figure 2: T.S.of Lamina.

1-Upper epidermis; 2-Palisade cells; 3-Secretory cells;

4- Spongy parenchyma; 5- Stomata; 6-Lower epidermis.

Microscopical characters

The transverse section of Jatropha curcas, Linn., divided into midrib (Figure 1, 3) and lamina (Figure 2). It is a dorsiventral type of leaf, consists of protoxylem and metaxylem, laticiferous cells are present in the collenchymas. In lamina (Figure 2), upper epidermis is covered with a cuticle layer, below this palisade cells are present, below this spongy parenchyma and lower epidermis are seen, which contains of paracytic type of stomata.

Figure 3: T.S.of Midrib-Portion enlarged.

Powder microscopy

The powder microscopy reveals that the presence of calcium oxalate crystals (Figure 4), xylem (Figure 5), phloem (Figure 6), multi cellular, covering trichomes (Figure 7) and paracytic type of stomata (Figure 8).

Figure 4: Calcium oxalate crystal.

Figure 5: Xylem.

Figure 6: Phloem.

Figure 7: Multicellular, covering trichomes.

Figure 8: Stomata.

Table 1: Physico-chemical parameters of Jatropha curcas, Linn., leaf

S. No	Parameters	% value
1.	Total ash	5.0
2.	Water soluble ash	2.74
3.	Acid insoluble ash	2.25
4.	Sulphated ash	0.42
5.	Moisture content	6.38
6.	Water soluble extractive value	17.2
7.	Alcohol soluble extractive value	18.6
8.	Crude fibre content	0.86

Table 2: Fluorescence analysis of Jatropha curcas, Linn., leaf powder

S. No	Reagents	Day light	UV light
1.	Powder as such	Light green	Green
2.	Powder + 1N NaoH (Alc)	Pale yellow	Greenish yellow
3.	Powder + 1N NaoH (Aq)	Pale green	Pale green
4.	Powder + 50% Hcl	Green	Deep green
5.	Powder + 50% HNO₃	Green	Deep green
6.	Powder + 50% H₂SO₄	Light green	Green
7.	Powder + 5% Fecl₂	Green	Dark green
8.	Powder + 5% I₂ solution	Green	Yellowish green
9.	Powder + Picric acid	Yellowish green	Yellowish green

Table 3: Fluorescence analysis of Jatropha curcas, Linn., leaf extracts

S. No	Extracts	Day light	UV light
1.	Petroleum ether	Green	Deep green
2.	Benzene	Green	Deep green
3.	Chloroform	Green	Dark green
4.	Acetone	Dark green	Dark green
5.	Ethyl acetate	Light green	Dark green
6.	Methanol	Green	Deep green
7.	Ethanol	Green	Deep green
8.	Aqueous extract	Green	Dark green

Physico-chemical parameters

All the physico-chemical parameter values are depicted in Table 1.

Fluorescence analysis

The leaf powder was treated with various reagents and the observations were recorded in Table 2. The leaf extracts were subjected to fluorescence analysis and chromophores were producing different colours and the results were tabulated in Table 3.

Table 4: Preliminary phytochemical screening of Jatropha curcas, Linn., leaf extracts

S. No	Phytoconstituents	Ethanol extract	Aqueous extract	Powder as such
1.	Carbohydrates	+	+	+
2.	Glycosides	+	+	+
3.	Saponins	-	-	-
4.	Phytosterols	+	+	+
5.	Alkaloids	+	+	+
6.	Proteins	-	-	-
7.	Amino acids	-	-	-
8.	Flavonoids	-	+	+
9.	Tannins	-	+	+
10.	Triterpenoids	+	-	+
11.	Lipids	-	-	+

(+) = Presence of phytoconstituents

(-) = Absence of phytoconstituents

Table 5: Antibacterial screening of Jatropha curcas, Linn., ethanol leaf extracts

S.No	Micro organisms	Zone of inhibition in mm		Ofloxacin
S.No	Micro organisms	25 mg/ml	50 mg/ml	10 µg/ml
1.	Escherichia coli	7	18	24
2.	Staphylococcus aureus	8	11	14
3.	Pseudomonas aeroginosa	4	7	24
4.	Klebsiella pneumoniae	6	11	18
5.	Micrococcus luteus	5	10	14

Preliminary phytochemical screening

Qualitative tests of ethanol and aqueous leaf extracts exhibited the presence of glycosides, saponins, tannins, phytosterols, carbohydrates, flavonoids and fixed oils were found out and enlisted in Table 4.

Figure 9 (a)

Figure 9 (b)

Figure 9 (c)

Figure 9 (d)

Figure 9 (e)

Figure 9 : Antibacterial activity of ethanolic leaf extract of Jatropha curcas, Linn.,

Antibacterial screening

Ethanol leaf extract (25, 50 mg/ml) of Jatropha curcas, Linn., exhibited significant antibacterial activity against Escherichia coli, Staphylococcus aureus, Pseudomonas aeroginosa, Klebsiella pneumoniae and Micrococcus luteus by disc plate method using Ofloxacin as a standard (10 µg/ml) (Figure 9(a), 9(b), 9(c), 9(d) and 9(e)).

CONCLUSION:

In this present study, various standardization parameters such as macroscopical, microscopical, physico-chemical, fluorescence analysis, phytochemical and antibacterial screening of Jatropha curcas, Linn., was carried out. In this present scenario, pharmacognostical studies have become mandatory for proper identification of the plant. The above parameters are valuable tool in the correct identification of Jatropha curcas, Linn.

ACKNOWLEGEMENT:

The authors are thankful to The Chairman, Dr. K. Varadharaajen, B.A., B.L., and Vice- Chairman, Mr. V. John Ashok, M.A., B.L., Thanthai Roever College of Pharmacy, Perambalur, Tamil Nadu, for providing all the facilities to carry out this work.

REFERENCES:

1. Aswini K, Mangesh K, Kailash V, Pratibha C and Mahavir G. Phrmacognostic investigation on leaves of Citrus maxima (Burm.) Merr. (Rutaceae). International Journal of Pharmaceutical Sciences and Research. 3(12); 2012: 4913-4918.

2. Aliyu AB, Musa AM, Abdullahi MS and Oyewale AO. Phytochemical and antibacterial properties of Ludwigia suffruticosa (Willd.) Oliv. ex O. Ktze (Onagraceae). International Journal of Pharmaceutical and Applied Sciences. 2(4); 2008: 1-5.

3. Parekh J and Chanda SV. In vitro antimicrobial activity and phytochemical analysis of some Indian medicinal plants. Turkish Journal of Biology. 31; 2007: 53-58.

4. Nadkarni KM. Indian Materia Medica, Popular Prakashan, Bombay. 1999; 3 rd ed: pp. 705-706.

5. Jimu L, Nyakudya IW and Katsvanga CAT. Establishment nd early field performance of Jatropha curcas, L. at Bindura University Farm, Zimbabwe. Journal of Sustainable Development in Africa. 10; 2009: 445-469.

6. Aiyelaagbe OO, Hamid AA, Fattorusso E, Scafati OT, Schroder HC and Muller WEG. Cytotoxic activity of crude extracts s well as of pure components from Jatropha species, plants used extensively in African Traditional medicine. Evidence Based Complementary and Alternative Medicine. 2011: 1-7.

7. Momotaz Begum, Sohrab MH, Farhana Afra, Shakila Akter, Hasan CM and Sarwaruddin Chowdhury AM. Phytochemical investigations on the leaves of Jatropha curcas, Dhaka University Journal of Science. 59 (1); 2011: 61- 63.

8. Sass JE. Elements of Botanical Microtechnique, Mc Graw Hill Book Co., New York. 1940; 1st ed : pp. 222.

9. Johansen DA. Plant Microtechnique, Mc Graw and Hill Book Co., New York. 1940; 5 th ed: pp. 523.

10. O’Brien TP, Feder N and Mc Cull ME. Polychromatic staining of Plant Cell walls by Toluidine blue-O, Protoplasma.. 59; 1964: 364-373.

11. Kokate CK, Purohit A and Gokhale S. Practical Pharmacognosy, Vallabh Prakashan, New Delhi. 1994; 10 th ed: pp. 112-114.

12. Anonymous. The Indian Pharmacopoeia, The Controller of Publication, New Delhi. 1996; 4 th ed: pp. A- 53, 54, 89.

13. Trease GE and Evans WC. Pharmacognosy. ELBS Publications, New Delhi. 1989; 12th ed: pp. 49, 126, 132-137, 205, 248.

14. Kokoshi CJ, Kokoshi RJ and Sharma M. Fluorescence of powdered vegetable drugs in ultra violet radiation. Journal of American Pharmaceutical Association. 47; 1958: 715-717.

15. Chase CR and Pratt RJ. Fluorescence of powdered vegetable drugs with particular reference to the development of a system of identification.. Journal of American Pharmaceutical Association. 38; 1949: 324-331.

16. Alam MT, Karim MM and Shakila N. Khan. Antibacterial Activity of Different Organic Extracts of Achyranthes aspera and Cassia alata. Journal of Scientific Research. 1 (2); 2009: 393- 398.

Received on 01.08.2013 Accepted on 27.09.2013

Asian J. Res. Pharm. Sci. 2013; Vol. 3: Issue 4, Pg 195-199