INTRODUCTION:

Since ancient period, plants are being used for the development of new drugs or as a phytomedicine for the treatment of diseases¹. Even the World Health Organization (WHO) supports the use of medicinal plants, provided it is proven to be efficacious, safe, less toxic, available and reliable natural resource². The scientific search for new drugs from natural products remains a serious task for scientists worldwide. It is a fact that a large segment of the population in tropical countries relies on traditional medicines for their health care needs³.

Over 80% of population in the developing world makes use of medicinal plant extracts to provide good health. The therapeutic basis of herbal medication are by the presence of diverse bioactive compounds like alkaloids, flavonoids, terpenoids, phenolic compounds, glycosides etc in plants and also for the treatment of diseases which are still incurable, medicinal plants can serve as a source of novel therapeutic agent⁴.

Diospyros virginiana is a persimmon species commonly called the American Persimmon, Common Persimmon, Eastern Persimmon, "'Simmon", "Possumwood", or "Sugar-plum". This is a well-known indigenous tree⁵, growing in woods and fields. The parts of the D.virginiana have a long history of use in the new world. Persimmon have been used medicinally as an astringent and antiseptic and for the treatment of uterine hemorrhage, diarrhea and dysentery, diphtheria, dropsy, fevers, gonorrhea, hemorrhoids, syphilis and thrush⁶. Persimmons have been used to lubricate the lungs and strengthen the spleen and pancreas. They improve energy and contain enzymes that help damaged cells and foreign microbes be broken down. Persimmons have a special affinity for the large intestines and heart⁷. Persimmons have been also used to treat bronchitis, catarrh, cough, , goiter, hangover and hiccoughs. The bark has been used in intermittent and both it and the unripe fruit have been beneficial in various forms of disease of the bowels, chronic dysentery⁸, and uterine hemorrhage; used in infusion, syrup, or vinous tincture.

Preliminary phytochemical screening of the plants is primarily an important aspect in finding the chemical constituents in plant materials. Hence the present study was qualitative analysis and quantitative estimation of phytoconstituents, heavy metals and inorganic elements were also carried out.

MATERIALS AND METHODS:

D.virginiana belongs to the family Ebenaceae was collected from Coonoor, Nilgiris District, Tamil Nadu, India and identified by the special key given Cambell flora. The leaf and bark of D. virginiana were washed with sterile distilled water. After, the leaves and bark were shade dried and powdered by using pestle and mortar. 25g of powder was filled in the thimble and extracted successively with ethanol using a Soxhlet extractor for 48 h. The extracts were concentrated using rotary flash evaporator and preserved at 5°C in airtight bottle until further use.

PHYTOCHEMICAL SCREENING:

Phytochemical screenings of the crude plant samples were carried out to identify the chemical constituents, using standard phytochemical methods as described by³.

Preparation of fat free sample:

2 g of each sample was defatted with 100 ml of diethyl ether using a soxhlet apparatus for 2hr ⁹.

Determination of alkaloids:

5 g of the samples were weighed into 250ml beaker and 200ml of 20% acetic acid was added and covered to stand for 4hr.This was filtered and the extract was concentrated using a water bath to one quarter of the original volume. Concentrated ammonium hydroxide was added drop wise to the extract until the precipitate was complete. The whole solution was allowed to settle and the precipitate was collected by filtration and weighed ¹⁰.

Determination flavonoids:

10g of the plant samples were extracted repeatedly with 100ml of 80% aqueous methanol at room temperature. The extract was filtered through whatman filter paper no. 42 (125mm). The filtrate was later transferred into a crucible and evaporated to dryness over a water bath and weighed ¹¹.

Determination of total phenols:

For total phenolics, the fat free sample was boiled with 50ml of ether for 15 min. 5ml of the extract was pipette into 50ml volumetric flask, then 10ml of distilled water was added. 2ml of ammonium hydroxide solution and 5ml of pentanol were added. The samples were made up to mark and left to react for 30min for color development. The absorbance of the solution was read using spectrophotometer at 505 nm wavelength ¹².

Determination of saponins:

About 1ml of plant extract was dissolved in 20ml of water and shaken well in a graduated cylinder for 15min.Formation of 1cm layer of foam indicates the presence of saponins.

Determination of gums and mucilage:

About 10ml of plant extract was slowly added to 25ml of absolute alcohol under constant stirring. Precipitation indicates the presence of gums and mucilage.

Determination of carbohydrate and amino acids:

The carbohydrate was determined by molishc’s reagent. Appearance of brown colour indicates the presence. The amino acids were determined by biuret test, ninhydrin test and xanthoprotein test.

TRACE ELEMENT ANALYSIS:

The major elements comprising calcium, sodium, potassium and trace elements (iron and zinc) were determined according to the method of ¹³ with slight modification. The ground samples were sieved with a 2mm rubber sieve and 2 g of each of the plant samples were subjected to dry ashing in a well-cleaned porcelain crucible at 550°C in a muffle furnace. The resultant ash was dissolved in 5 ml of HNO₃ /H₂O₂ (1:1) and heated gently on hot plate until brown fumes disappeared. To the remaining material in each crucible, 5ml of deionized water was added and heated until a colourless solution was obtained. The mineral solution in each crucible was transferred into a 100ml volumetric flask by filtration through a whatman filter paper and the volume was made to mark with deionized water. This solution was used for elemental analysis by atomic absorption spectrophotometer (AAS).Concentration of each element was calculated on percentage of dry matter.

Statistical analysis:

All determinations were replicated three times and results were reported as mean values ± standard deviation.

RESULT AND DISCUSSION:

Phytochemical analysis of the both extract revealed that the presence of alkaloids, carbohydrates, glycosides, polysterols fixed oils and fats, phenolic compounds and tannins, flavonoids, proteins and amino acids and absence of saponins, gums and mucilage and volatile oils in aqueous extract of the plant(Table-1).

The phytochemical analysis of the D.virginiana extracts showed the presence of tannins, alkoloids, flavonoids and phenolic compounds. Tannins have been found to form irreversible complex with proline rich protein synthesis. The activity was exhibited against test organisms with the plant extract ^14.

Table 1: Qualitative Phytochemical screening on extracts of D.virginiana

S.No

Name of Test

Leaf extract

Bark extract

Alkaloids

Carbohydrate

Protein

Fixed oil and fats

Tannin

Saponin

Gum

Volatile oils

Amino acids

Polysterols

Phenolic compounds

Flavonoids

Mucilage

Glycosides

The abundance of flavonoids which are hydroxylated phenolics substances might be responsible for their therapeutic effectiveness against wide array of microorganisms, probably due to their ability to complex with extracellular and soluble proteins and to complex with the bacterial cell wall¹⁷. Flavonoids and other phenolic compounds are potent water soluble antioxidants and free radical scavengers, which prevent oxidative cell damage, have strong anti-cancer activity ¹⁵.

Alkaloids are very important in medicine and constitute most of the valuable drugs. They have marked physiological effect on animals ¹⁶.

An examination of the data from (Table – 2) shows that different medicinal plants contain elements like Mn, Fe, Zn and Cu in various proportions. A number of trace elements play an important role in the metabolism. These elements are called essential. An element is considered essential for a plant if the plant fails to grow normally and complete its life cycle in a medium adequately removed from the element whereas in the presence of the suitable chosen concentration of that element it grows and reproduces normally.

Table-2: Trace element content of leaves and bark of D.virginiana

S.No

Trace element

Leaf (µg/ml)

Bark (µg/ml)

25.20

17.85

9.34

6.00

18.02

15.65

1.24

0.48

These mineral elements are very important in human nutrition. Calcium, potassium and magnesium are required for repair of worn out cells, strong bones and teeth in humans, building of red blood cells and for body mechanisms ^{17, 20}. The biological roles for K and Ca are essential for disease prevention and control and may, therefore, contribute to some of the traditional medicinal influences of the plants.

Trace elements such as manganese, iron and zinc are essential in enzymes metabolism. The concentrations of these elements in the plants are quiet important. Manganese is an important modulator of cells functions and play vital role in the control of diabetes mellitus^18,19. The importance of iron in maintaining the good health has been recognized. Lead is best known for its toxicity even at low concentration.

It showed that the plants hold tremendous promise in providing the variable secondary metabolites and mineral supply that could enhance the curative process of ill health. These findings provide quantitative estimation of the phytochemicals as well as mineral element analysis which are important in understanding the pharmacological and/or toxicological actions of medicinal plants. Further work on extraction and purification of active constituents should be of interest.

REFERENCES:

1. Iwu M, Hand book of African Medicinal plants. CRC press, Boca Raton, Fl. 1993.

2. World Health Organization (WHO): World Health Report. Bridging the gap. 1, WHO, Geneva 1995:118.

3. Sofoware A. Screening plants for Bioactive Agents. In 2nd Edn, Medicinal Plants and Traditional Medicine in Africa. Spectrum books Ltd; Sunshine House, Ibadan, Nigeria, 42-44; 1993:221-229.

4. World Health Organization (WHO): Traditional Medicine. Growing needs and Potential, WHO policy perspective on Medicines, Geneva 2002: 1-6.

5. Anonymous. The persimmon. Gard and Forest. 2(95); 1889: 612.

6. Barram I. A memoir on the distillation of persimmon. Trans.Philos.Soc., 1;1771:231-234.

7. Collins RJ George AP and Mowat AD. The world trade in persimmon. Chron. Hort., 33(2);1993:5-7.

8. Worth CB. Virginia opossums as dissemination of the common persimmon (Diospyros virginiana). J. Mammalogy, 56; 1975:517.

9. Okwu DE and Josiah C. Evaluation of the chemical composition of two Nigerian medicinal plants. Afr. J. Biotech., 5 (4); 2006:357-361.

10. Harborne JB. Phytochemical Methods: A Guide to Modern Techniques of Plant Analysis. Chapman and Hall Ltd. London. 1973: Pp. 49-188.

11. Shahidi F Chavan UD Bal AK and McKenzie DB. Chemical composition of beach pea (Lathyrus maritimus L.) plant parts. Food Chem., 64; 1999: 39-44.

12. Edith A Mofolusho F Omonike O Larry O Dora A.In vivo antimalarial and cytotoxic properties of Annona senegalenisis extract. Afr. J.Trad. Compl. Alt.Med., 3(1); 2005:138-139.

13. Cowan MM. Plant Products as Antimicrobials Agents. Clinical Microbiology Review, 12(4); 1999:564-582.

14. Del-Rio A Obdululio BG Casfillo J Marin FG and Ortuno A. Uses and Properties of citrus flavonoids. J. Agric Food Chem. 45; 1997:4505-4515.

15. Okwu DE. Phytochemicals and Vitamin content of indigenous spices of Southeastern Nigeria. J. Sustain Agric. Environ. 6(1); 2004:30-37.

16. Edeoga HO and Eriata DO. Alkaloid, tannin and saponin contents of some Nigeria medicinal plants. J. Med. Aromatic Plant Sci. 23; 2001:344-349.

17. WHO. Trace elements in Human Nutrition and Health. WHO Technical Report Series, Geneva. 1996: Pp. 199-205.

18. Korc M. Manganese Homeostasis in Human and its Role in Disease states. In: Prasad, A.S. (ed.) Essential and Toxic Trace Elements in Human Health and Disease. Alan R. Liss Inc. New York. 1988 :Pp. 253-264.

19. Vaughan JG and Judd A. The Oxford Book of Health Foods: A Comprehensive Guide to Natural Remedies.1st Edition, Oxford University Press, New York, P. 2003: xvii.

Received on 22.11.2013 Modified on 10.12.2013

Res. J. Pharmacology & P’dynamics. 6(1): Jan.-Mar. 2014; Page 05-07