FTIR SPECTRAL ANALYSIS, PHYTOCHEMICAL AND BIOLOGICAL ACTIVITY OF RICINUS COMMUNIS

1. Institute of Biochemistry, University of Sindh, Jamshoro, Sindh, Pakistan. 2. Department of Community Medicine, Gambat Medical College, Khairpur Mirs, Sindh, Pakistan. 3. Water Testing and Surveillance Laboratory Liaquat University medical and Health Science Jamshoro, Sindh, Pakistan. 4. Department of Biochemistry, Gambat Medical College, Khairpur Mirus, Sindh, Pakistan. ...................................................................................................................... Manuscript Info Abstract ......................... ........................................................................ Manuscript History Received: 25 May 2020 Final Accepted: 28 June 2020 Published: July 2020


Estimation of Total Phenolic Compound by Folin Ciocalteu Reagent Method: Preparation of Sample:
For this, 10 gram of sample was taken and dissolved in the 100 ml of 80% methanol in aqueous. It is kept on shaking water bath at room temperature for 24 hours. The solution was filtered through whatmann filter paper no 1 and centrifuged 20 minutes at 6000 rpm. The filtered material was stored in the refrigerator for further analysis of total phenolic compound, tannin and flavonoid.

Standard Solution:
100 µg/ml of the gallic acid was prepared in the 80% of gallic acid was prepared in the 80 % methanol in aqueous. Standard was arranged including (10,20,40, 60, 80 and 100µg/ml).

Procedure:
For this procedure, 0.5 ml plant sample or Standard dissolved in the 2.5 ml of reagent folin-ciocalteu (10 fold diluted in the distilled water). Further 2ml of 7.5% Na 2 CO 3 was added in solution. It is placed in the incubator at room temperature for 30 minutes then absorbance was taken at 760 nm (Maurya S, 2010).
Procedure: 1 ml of extract or standard was taken and diluted with 4 ml of distilled water. 0.3 ml of 5% sodium nitrate was added and placed for 5 minutes. 0.3 ml 10% AlCl 3 was added and to stand for six minutes. Further 2 ml of 1MNaOH was added and final volume 10 ml was made by distilled water and then absorbance was measured at 510 nm ( (pp. Domidar et al., 2011).

Procedure:
In this test, 0.1 ml of samplewas diluted in the 6.9 ml of distilled water. For this, 1 ml of 0.008M of the potassium ferric cyanide was added. Further 1 ml of 0.2 M of ferric chloride in 0.1HCL was added and shaken well. The solution colour blue was formed. Absorbance was taken at 700 nm by spectrophotometer (Sathishkumar T, Baskar R, 2014).

Estimation of total alkaloid by Dragendorff's method:
Dragendorff's reagent: Solution A for this solution, 0.8 gram of the Bismuth Nitrate pentahydrate was dissolved in the 40ml of distilled water then 10 ml glacial acetic acid was added. Solution B for this, 8 gram potassium iodide was taken and 30 ml of water distilled was added. Further solution B and A were mixed.

Standard Solution:
In this, 10 mg of Bismuth Nitrate Pentahydrate was dissolved in the 5 ml concentrated HNO 3 was added and then final volume 100 ml was made in the double distilled water. The standard arranged as (200, 400, 600, 800 and 1000 µg/ml).

Preparation of extract:
For this, 10 g of plant fine powdered was taken and dissolved in 50 ml of 2% aqueous acetic acid and kept on 1177 boiling water bath for 30 minutes. Solution was filtered through whatmann filter paper no-1. The filtrate was collected and residue was extracted again repeated procedure respectively whereas both solutions were mixed and then 100 ml final volume was prepared and the pH 2.5 of extract was maintained.

Procedure:
For this procedure, 5 ml of extract or standard was taken and treated with 2 ml of the reagent Dragendorff's. The precipitates were formed wait few minutes for full precipitation. Further solutions were centrifuged at 4000 rpm for ten minutes. The precipitate was collected and remaining solution was again treated by dragendorff's reagent for checking the precipitation more, if precipitate was formed further precipitates were mixed and washed by alcohol. The precipitates were dissolved in the 2 ml Disodium sulfide after addition brownish black precipitates were formed remaining solution was discarded. The precipitates were treated with 2ml of HNO 3 and 10 ml of double distilled water was added. 1 ml was taken out from prepared solution and 5 ml of thiourea was added and absorbance was measured at 435 nm ( (Sonal P et al., (2011).

Preparation of Mineral Sample by wet acid digestion method:
For this, 0.5 g of fine powdered plant was taken in the volumetric flask and 5 ml of the (Conc:) HNO 3 were added. Place at room temperature for three hour. Volumetric flask was covered by watch glass and heated at 100 o C on a hot plate. Further 5 ml of concentrated HNO 3 were added and heated. Further 2 ml of H 2 O 2 were added and well shaken heated until transparent clear solutions were obtained and heated until near to dryness. 10 ml of deionized water were added and shaken well. Filtered through whatmann filter paper 42 and final volume 25 ml was made in the deionized water and concentration was taken by atomic absorption spectrometer ( AA.800 perkinelmer) ((Lanjwani, AH et al., (2016).

.Estimation of antimicrobial activity by agar well diffusion method: Media for bacterial cultures:
The medium was neutralized at 37 o C for 30 minutes and filtered further then sterilized at 15 lbs for 20 minutes at 121 o C.

Procedure:
Suspension of 24 hours cultures of staphylococcus aurous, bacillus cereus, Escherichia coli and Klebsiella pneumonia was made and sterilized with normal saline. Medium each plate was inoculated by test organism and sterilized by swab rolled of cotton in the suspension to the strip plate surface in formthat lawn growth were formed. The 5 mm diameter cork borer were used for formation of well in the medium plates. 50 µl of plant extract (80 % methanol) were dropped into the each well. The each agar plate was incubated at 37 o C for 24 hours (Banjar G et al., (2014).

Estimation of antioxidant content by ferric reducing antioxidant powermethod: Procedure:
For this procedure, 2.5 ml of plant extract was diluted with buffer phosphate (pH 6.6) and 1 ml potassium ferric cyanides (1%) were added. It was kept in the incubator at 50 ℃ for 20 minutes. It was cooled and diluted with 2.5 ml of Trichloroacetic acids (10 %) and then centrifuged for ten minutes. From above solution, 2.5 ml was taken and diluted in the 2.5 ml of water distilled, further, 0.5 ml of ferric chlorides (0.1%) were added. Above solutions color green was formed. It was stand for ten minutes and absorbance was measured at 593 nm by spectrophotometer (Patel A, 2010).

Result And Discussion:-
Medicinal plants have been used from generation to generation in the traditional system of medicine for treatment various types of diseases.There is a broad range of plant parts possessing a variety of pharmacological properties. The peak at 2926 wave number cm-1 was investigated in the seed and flowers are due to the asymmetric stretching of C-H group of aromatic compounds. This peak indicated the presence of glycoside, tannin, flavonoid and saponin. The peak at 1030 cm-1 indicated the S=O group. It is indication of organosulfur compounds such as, allicin, alliin and diallyl disulphide (Songsungkan J, 2011). The very strong absorption at the region between 2933-2922 cm-1 is due to N-H stretching. The lone C=O stretching vibration band which is corresponding to saturated aliphatic ester present in the all selected parts of the plant. The bands at 900-1350cm-1, 1030 cm-1, in the leaves and 1027 cm-1 in the stem and flowers are attributed to phosphodiester stretching bands region. This peak absorbance due to glycogen, collagen and DNA. The functional group C-O stretch associated with phosphate, glycogen and 1178 oligosaccharides PO-2 stretching modes (Fabian H et al.,(1995). The strong band absorption is observed between 1600 -1660 cm-1 region which indicated the presence of amino acids. The very strong absorption 1601 cm-1, 1605, 1615 were investigated in the selected parts. This result gives the evidence that all parts of solanum surattense indicate the high content of the protein. There is no evidence in the between the region 2220-2260 cm-1 indicates no presence of the cyanide groups in all parts whereas cyanide have toxic effect for consumer (Manju S., (2011). The present screening of phytochemical finding showed thatthe appreciable amount of phytochemical including, carbohydrate, glycoside, phenolic compounds, tannin, flavonoid, saponin, steroid, terpenoid, glycosides amino acid, proteins, fat and oil were investigated whereas the moderate amount of vitamin C and trace of alkaloid was investigated.Present finding showed similarity reported that presence of phenolic compound, flavonoid, tannin, steroid, alkaloid in the stem of Ricinus Communis (Ramesh KS et         FTIR Peak Values of Ricinus Communis stem is shown inthe figure 3. The peak at 3345.77 wave number cm-1 due to the O-H group of hydrogen bounded alcohols and phenols. The peak at 2917.74 wave number cm-1 due to the asymmetric stretching of C-H group of aromatic compounds. It is indicated the presence of glycoside, tannin, flavonoid and saponin. The peak at 1732.87 wave number cm-1 is due to C=O stretch which is evidence of Aldehydes, Ketones, Carboxylic acids, Esters functional group. The peak at 1506 wave number cm-1 is due to C-H (Aromatic ring). The peak at 1421.31, 1371.39 and 1238.90 is due to C-N bonds showed Alkyl ketone, Amines, Amides. The strong band absorption at 1027 cm-1 in the stem is attributed to phosphodiester stretching bands region. It is indicated the presence of glycogen, collagen and DNA. Proximate composition is shown in the table 4. Roots showed highest amount of ash that is good sign because ash possessed minerals deposition in the plants. The stem and root of plants showed rich bio resources of carbohydrate; whereas the highest percentage of protein was observed in the leaves. These parts can play key role to against malnutrition because of Protein and carbohydrate is biggest challenge in Pakistan. Food insecurity of household may be related the protein energy malnutrition. It is evident in the underweight, stunting and wasting whereas affects one-quarter of the world's children (Nations., 2012). One child out of every three children has been malnourished. The 6.2-8.3 million Pakistani children 30-40% have low height for their age called stunting whereas more than 2.9 million Pakistani children greater than 40% have low weight for their height called wasting (Mujib SA et al.,(2004).  Mineral Composition is shown in the table 6. It is investigated that appreciable sources of principle essential macro minerals including Calcium, iron, magnesium, sodium and potassium whereas seeds showed the appreciable sources of mineral composition including Calcium, sodium, magnesium, potassium and iron.The content of minerals in the medicinal plant that depends on the soils abundance, intensity of fertility. The concentration of cadmium, chromium, lead has been indicated below detection limit which is toxic highly even at concentration low. Calcium play most important role to maintaining and building strong teeth and bone as well as important for normal function of blood coagulation, regulation of cell permeability, milk clotting and cardiac muscle (RD, 1994). Magnesium has played most important role in the formation and function of bone and muscle and prevention of high blood pressure and depression (Smith WD and Hammarsten JF, 1958). Sodium and potassium play key role in the ionic balance of the body, carry normal muscle contraction, maintain tissueexcitability and formation of gastric juice in the stomach whereas iron is possessed essential function for formation of Blood haemoglobin, carry oxygen in body, and to make body tendons and ligaments (Shivraj H, Nile and CN N Khobragade, 2009).  (Abhishek, 2011). Castor oil is rich bio resources of antibacterial active phytochemical that can be useful for inhibition the growth of microorganism This plant are rich bio sources of antibacterial phytochemical that can be useful for inhibition the growth of life threatening pathogenic including Escherichia coli leads hemorrhagic colitis Syndrome and haemolytic uremic syndrome whereas flowers of present finding showed very good zone of inhibition Escherichia coli (Zhou B et al., (2017).

Conclusion:-
In the present conclusion it can be noticed that this plantis wealthy resources of active phytochemical that is strongly emphasizedand can be accessiblebio-resources plants derived new drugs. Therefore, the parts this plant could b seen as the rich sources of for useful drugs. Extracts from this plant could be seen as a good source for useful drugs. The traditional medicine practices are strongly recommended and further research work should be carried out isolation, purification and characterization of phytochemical.