Phytochemical Constituents, Total Saponins, Alkaloids, Flavonoids and Vitamin C Contents of Ethanol Extracts of five Solanum torvum Fruits

Introduction: Phytochemicals are frequently used in chemotherapeutic treatment or may be used as chemo preventive agents with chemoprevention. The study report the quantification of phytochemical constituents and vitamin C contents from ethanol extracts of Solanum torvum fruits. Method: The main objective for this research was to use standard procedures to determine phytochemical and vitamin C content. Results: The estimated alkaloids found in mature fruits were 6.32 ± 0.12 mg/g and 16.94 ± 2.3 mg/g in the immature fruits. Total saponins in mature and immature fruits were 8.60 ± 2.6 mg/g and 16.90 ± 9.4 mg/g respectively. Total flavonoids in mature and immature fruits were 21.14 ± 4.4 mg/g and 14.24 ± 1.8 mg/g respectively. Also vitamin C contents were 11.79 ± 2.0 mg/g in mature fruits and 8.70 ± 0.26 mg/g in immature fruits. With the exception of alkaloids whose difference in the mature and immature was significant, other differences obtained were not significant. Conclusion: The study showed that the extracts contain diversity of phytochemicals in appreciable amount that can expertly keep the body against oxidative stress triggered by free radicals and therefore be used as a source of potent natural products.

Solanum torvum fruits are very rich in vitamin C. In spite of being rich in vitamin C, they also provide phytochemicals such as alkaloids, saponins, sugars, phenols and flavonoids folate, potassium and a lot of fiber for pharmaceutical or food production. 6olanum torvum is a rich source of alkaloids, flavonoids, saponins, glycosides, and tannins. 7According to Pérez-Amador, 8 the percentage constituents of various phytochemical compounds within this fruit is highly appreciable.Regardless of broad research on the medicinal properties of most plants, little information is known about the many humid underutilized plants and fruits in most unindustrialized countries specifically Ghana.One of such fruit is Solanum torvum which is a multipurpose constant slender herb.Consequently, in this research, quantification of phytochemical constituents and vitamin C contents by ethanol extracts of Solanum torvum fruits were investigated to evaluate the potential protective or health benefits of this fruits and their linkage to their medicinal properties.

Determination of people's knowledge on Solanum torvum using questionnaires
A survey was done with the households at Kotokoraba near University of Cape Coast, Cape Coast, Ghana.This was done using questionnaires to help know people's knowledge on S. torvum.The data was collected and analyzed using SPSS analysis.

Plant Material
Five fresh fruits Solanum torvum were collected from the University of Cape Coast botanical garden, Cape Coast, Ghana with voucher numbers UCCBG000445, UCCBG000448, UCCBG000349, UCCBG000387 and UCCBG000543.The taxonomic character of the fruits was determined on 2 nd December, 2014 by a plant taxonomist at the Department of Botany, University of Cape Coast, Ghana.Solanum torvum fruits were then washed under with water to get rid of undesirable dirt and other external constituents.The samples were parched under shade until no moisture left.The parched samples then were ground into powder using a blender.

Ethanol extraction
The ethanol extract from Solanum torvum fruits was also prepared by drenching 50 g of the grinded sample in 500 mL of sterilized distilled water for 2 h in 90°C water bath at the of Biochemistry Research Laboratory, University of Cape Coast, Ghana.The combination was then filtered using Whatman filter paper No 1.The filtrate was concentrated under a reduced pressure using a rotary evaporator at a temperature of 90°C.The resulting extract was weighed and put in storage in impermeable bottles at 35°C for further analysis.
Where, A 0 was the mass of the sample and A 1 was the mass of the crude extract.

Quantitative Analysis Determination of total saponins
The 10 samples were ground and 2 g of each was put into a conical flask and 10 ml of 20% aqueous ethanol was added.The samples were heated over a hot water bath for 4 h with continuous stirring at about 55°C.The mixture was filtered and the residue re-extracted with another 20ml of 20% ethanol.The combined extracts were concentrated over water bath at about 90°C.The concentrate was transferred into a 250 ml separating funnel and 10ml of diethyl ether was added and shaken strongly.The aqueous layer was recovered while the ether layer was cast-off.The purification process was repeated.6 ml of n-butanol was added.The combined n-butanol extracts were washed twice with 1ml of 5% aqueous sodium chloride.The residual solution was heated in a water bath.After evaporation the samples were dried in the oven to a constant weight; the saponin content was calculated. 13

Determination of total alkaloids
Exactly 1g of the sample was weighed into a 250 ml beaker and 40 ml of 10% acetic acid in ethanol was added and covered and allowed to stand for 4 h.This was filtered and the extract was concentrated on a water bath to one-quarter of the original volume.Concentrated ammonium hydroxide was added drop wise to the extract until the precipitation was complete.The whole solution was allowed to settle and the precipitate was collected and washed with dilute ammonium hydroxide and then filtered.The residue is the alkaloid, which was dried and weighed. 14

Determination of total flavonoids
Exactly 1g of the plant sample was extracted repeatedly with 10 ml of 80% aqueous methanol at room temperature.The whole solution was filtered through whatman filter paper No 42 (125 mm).The filtrate was later transported into a container and evaporated into dryness over a water bath and weighed to a constant weight. 15termination of Vitamin C About 3.5 g of the cleaned fresh 10 samples was measured.The samples were ground with a clean mortar and pestle which had been rinsed with oxalic acid.Exactly 50 ml of oxalic acid was used in each case to grind and dissolve each sample.The mixtures were filtered using cotton wool and collected in separate labeled beakers.After that, 10 ml of each filtrate was pipetted into a conical flask and 1ml of bromine water was added with constant mixing until the colour of the solution had changed to orange yellow.This was to remove phenolic hydrogen atoms in the ascorbic acid.Thiourea was added to the solutions in drops to expel off excess bromine.The volume of each of these sample solutions was made up to 50 ml by the addition of 4% oxalic acid solution.The same procedure was followed for 10 ml of stock ascorbic acid solution which has been prepared earlier to convert it to dehydroascorbic acid by bromination, followed by expelling excess with 10% thiourea.0.05 ml, 0.10 ml, 0.15 ml, 0.20 ml, 0.25 ml, 0.30 ml, 0.35 ml, 0.40 ml, 0.45 ml and 0.5 ml of the dehydroascorbic acid were pipette out into a series of test tubes.Similarly 1 ml of each of the brominated samples were pipetted out into a series of tubes and their volumes made up to 3 ml by adding distilled water.To each of the tubes was added 1ml of 2, 4-dinitrophenylhydrazine (2, 4-DNPH) followed by 2 drops of thiourea.A blank (containing 3 ml of distilled water, 1ml of 2, 4-DNPH, and drops of thiourea) was prepared.The contents were mixed thoroughly and then put in the water bath, at 40 o C for 3 h.After 3 h of incubation, 6 ml of 80% sulphuric acid was added to each solution and allowed to cool slowly in free air.Absorbance of the blank, the samples, and the standards were measured at 540 nm and tabulated.A standard curve was plotted and from it, the various concentrations of vitamin c in the samples were calculated.This was triplicated. 16

Survey
The survey revealed the knowledge of the local people about the fruits of S. torvum.From the survey, the plant is widely known among the local people (Figure 1).It has been assigned various names such as Beware, Kwau Nsusuwa, Samantroba, Ananantroba, etc.

Qualitative determination of the phytochemical constituents of ethanol extract of Solanum torvum
Solanum torvum has been largely considered, precisely its medicinal possessions correlated with stable phytochemicals. 17Several parts of Solanum torvum have been in use for the separation of an extensive collection of compounds.This plant kind is a precise good source of phytochemical components. 7Phytochemical study of the ethanol extract of S. torvum revealed the presence of glycosides, tannins, flavonoids, saponins, alkaloids but no anthraquinones (Table 1).

Quantitative determination of total alkaloid, saponin and flavonoid Contents of ethanol extract of Solanum torvum fruits
There were different compositions of phytochemicals in the fruit samples of S. torvum.There were no significant differences statistically in the levels of flavonoids, and saponins between the mature and young fruits of the S. torvum (Table 2; Figure 2-4).There was a statistically significant difference in the levels of alkaloids between the matured and young fruits of the S. torvum (Table 2; Figure 2-4).

Quantitative analysis of vitamin C content of ethanol extract of Solanum torvum fruits
From the experiment there was no statistical significant difference in the levels of vitamin C contents between the matured and young fruits of the S. torvum (Figure 5)

DISCUSSIONS
Most of the people surveyed used the fruits for food, tea or medicine (Figure 1).There has also been a report of the use of the fruits as food in most Asian countries. 18The advantages in using the fruits for food were well known to the people surveyed.The fruits have been used by the people from about 2 years to 40 years.This gave the basis to experimentally determine the nutritional or medicinal value of the fruits, especially the phytochemicals and vitamin C constituents.Sivapriya and Srinivas 19 and Lakshmi 20 described phytochemical screening of methanolic and ethanol extracts of sun dried S. torvum fruits having the same results as this study had.The presence of these bioactive composites in the extract point out the prospective health remunerations of the plant.][23] Young fruits normally store phytochemicals such as alkaloids to protect them from infections and insects.The alkaloids begin to reduce and their concentrations weaken as the fruits mature and ripen.Environmental influences such as soil type, growing season, geographic location, and mineral status are known to impact intensities of plant secondary metabolites. 24S. torvum fruits that exhibited great altitudes of phytochemicals may be strong to insects and diseases using the phytochemicals as protective chemicals [25][26][27][28] in recent times demonstrated that anti-oxidant possessions of flavonoids and their ability to chelate free metallic ions could be operational in reducing the poisonousness of Doxorubicin (DOX) 39.The presence of flavonoids in S. torvum makes the fruit a very potent antioxidant.The altitudes of vitamin C in vegetables and fruits differ subjected on some factors such as maturity state, position of the fruit on the branch   (Figure 5).1][32] It lowers blood pressure and cholesterol levels. 33he presence of vitamin C in the fruits of S. torvum will help decline the possibility of heart related diseases.

CONCLUSION
The outcomes of the study point out that extracts of Solanum torvum fruits comprise of diversity of phytochemical compounds that can expertly protect the body against oxidative stress caused by free radicals and as a result might be used as a source of potent natural food or medicinal compounds.The vitamin C content of Solanum torvum could substantiate additional analysis of its other constructive natural properties and regulate its protection.   of the stem, temperature, storage, processing, varieties and handling. 29o compare to this study, vitamin C decreases throughout the ripening process.Immature fruit has the highest levels according to previous studies but with this study, matured fruits had the highest content of vitamin C

Figure 5 :
Figure 5: Concentrations of Vitamin C in fruit samples of Solanum torvum.