Phytochemical pro ile and medicinal potentials of Lannea Coromandelica stem

In the history of humanity, great ancestors of Indian sub-continent used plants that possessed unique medicinal properties. They identi ied the plants from jungles and derived crude drugs out of them for treating infectious diseases. This treatment process was a tradition. In the recent century, this plant-based drug extraction, processing and re ining began scienti ically by pharmaceutical industries. In this new era, plants with medicinal features are being used in curing broad spectrumof diseases. Lannea Coromandelica (L.C.) is amedicinal plant that belongs to this category. Tribals extensively used it in treating various infectious diseases and common injuries. There have been very few studies on the leaves, barks, lowers, gums and mucilage of this tree. But no preliminary phytochemical composition of L.C. stem has been studied. Therefore, the current work focusses on the screening of the phytochemical pro ile of the stem of L.C. by the sequential ethanolic extract. Stems of L.C. were procured from Mettur, Tamil Nadu. Thirteen different components were qualitatively analyzed using standard procedures from 100g of L.C. stem extract. In this study, seven components were identi ied, and their percentage was estimated. Using GC-MS, 50 components were identi ied of which Pentadecanoic acid, 14-methyl-methyl ester (1.0%) was the major component. The presence of these components in L.C. stem extract can be used in the treatment of different ailments through their antioxidant, anti-arthritic, anti-diabetic, antiin lammatory and antimicrobial activities.


INTRODUCTION
Life on earth predominantly depends upon the natural resources for its survival. Medicinal plants being one of the essential natural resources, its purpose and utility have been well recognized by our great ancestors for many centuries. They used medicinal plants to cure infectious diseases in human beings and animals by traditional methods. Even today in this digital world, medicinal plants contribute extensively for various developments in the form of new drug discovery, drug formulas (Das, 2014) and antibiotics for treating diseases. Globally, there are many rare species of plants still unexplored for its medicinal properties. In India, one such rare medicinal plant is Lannea Coromandelica (L.C.) belonging to the Anacardiaceae family (Vadivel et al., 2012;juan Yun et al., 2014). Traditionally, this plant was used as folk medicine by local tribes in different parts of the India Lee et al. (2017); Islam et al. (2018) and also in other countries like China, Srilanka, Bangladesh (M et al., 2016;Islam and Tahara, 2000), Africa (Premjanu and Jaynthy, 2014;Islam et al., 2018), Nepal and Pakistan. It is widely known as Indian ash tree (Jain et al., 2013;Venkatesham et al., 2014) and by different names based on the growing location of the plant. Tribal people used various parts of this plant for treating pain (Kumar and Jain, 2015;Imam and Moniruzzaman, 2014), wounds (Sathish et al., 2010), cuts, ulcers (Tahara et al., 2002;Ahmed et al., 2013), gastritis, jaundice (Rahmatullah et al., 2010), gout, sprains, diarrhoea (Majumder and Md, 2013), sore eyes, leprosy, impotence (Vadivel et al., 2012) and dysentery. (Pavithra, 2018) in the review article illustrated medicinal importance of Odina, Wodier. She has explained that ethanol extracts of the plant showed to produce a pharmacological response which includes antioxidant activity, antimicrobial, wound healing, anti-diabetic, anti-arthritic activity. It has been found by Reddy and Joy (2011) that L.C. comprised of Flavonoids, tannins, polyphenols, Gallic acid, mucilage, gums and some sterols. The lowers and leaves of L.C. were found to constitute Ellagic acid and Quercetin-3-arabinoside (Subramanian and Nair, 1971). No preliminary studies have been reported so far on the stem of Lannea Coromandelica. Therefore, we have delved the phytochemical compositions of L.C. stem for its pharmacological and therapeutic applications.

Identi ication and Collection
Fresh samples of L.C. stems were procured in April 2019 from Siddha Medicinal Plants, Mettur, Tamil Nadu, India with the help of Dr Padma Sorna, Research of icer -Botanist. The stems of the plant along with leaves, lowers and other plant parts were submitted to Siddha Medicinal Plants Garden, Mettur as herbarium. The sample specimen of L.C. was identi ied and authenticated by Dr P. Radha, Research of icer -Botany.

Flavonoid estimation
Aluminium chloride was used to estimate lavonoid found in the L.C. stem extracts in which Quercetin was used as standard described by Sakanaka et al. (2005). In Ethyl acetate (10 mg/ ml) Quercetin and extracts were prepared. In the test tube, 0.9 ml of distilled water was added along with 0.1 ml of L.C. extract and mixed with 75 µL of 5% sodium nitrate solution. This mixture was allowed to remain for about 6 minutes, and after that 10 µL of 10%, Aluminum chloride was added. After 5 minutes, 0.5 ml of 1M Sodium hydroxide was added to the mixture.
To the mix, distilled water was added to make it up to 2.5 ml and thereby the mixture was shaken well. UV-Visible absorbance was measured at 510 nm. The estimation of lavonoid was expressed in terms of Quercetin equivalence (Q.E.) µg/mg of L.C. extract.

Extract preparation
The stems of Lannea Coromandelica were shade dried for one month and Coarse powdered by using a mixer grinder. The powder was stored at room temperature and covered by plastic wrap to avoid moisture absorption. The inely powdered sample was subjected to sequential extraction using ethanol.   Phenols (Gallic acid) 3.02% 3.

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The extracts were iltered and concentrated to a dry mass separately, evaporated with the help of rotary evaporator at 35 • C and stored in the refrigerator for the phytochemical analysis.

Phenol estimation
Folin Ciocalteu colorimetric method was used to estimate phenol compounds in the L.C. ethanolic extracts. They were with Gallic Acid as standard (10mg/10ml) by Kumar et al. (2009). To different test tubes, 20 to 100 µl of standard solutions were added. 5ml of Folins-Ciocalteu (1:10 dilution) was added to a separate test tube containing extract at a concentration of 10 mg/ml, and the mixture was shaken thoroughly. For half an hour, the mixture was incubated after adding 4ml of 0.7 M sodium carbonate. UV-Visible absorbance was measured at 765nm. The estimation of phenol was expressed in terms of Gallic acid equivalence (G.E.) µg/mg of L.C. extract.

Tannin estimation
Folin-Ciocalteu method was used to estimate the tannin found in the ethanolic extracts of L.C. by Harborne (1973). 1 ml of 35 % Na2CO3, 7.5 ml of distilled water and 0.5 ml of Folin-Ciocalteu reagent was added to the test tube. To this mixture, 0.1 ml of the sample extracts containing 1 mg were added and by adding distilled water to this mixture, it was made up to 10ml. The mixture was kept at room temperature for 30 minutes after being thoroughly shaken. 20, 40, 60, 80 and 100 µg/ml) were used as standard reference solutions for tannic acid. UV/Visible absorbance was measured at 725 nm against the blank solution. The estimation of tannin was expressed in terms of mg of Tannic acid equivalence (T.E.) µg/mg of L.C. extract.

GC-MS analysis of L.C. stem
Using Turbo mass software (5.2.0) and Perkin Elmer Clarus 500, the GCMS analysis was performed. A capillary column of 250 µm column I.D. and 30m length with Elite 5MS (5% phenyl 95% dimethyl polysiloxane) was used in the Perkin Elmer Clarus 500.
The temperature for the mass analyzer was 8ºC/min to 200ºC. The oven temperature was retained at 50ºC with 1-minute holding time in the initial stage of GC The Helium gas was used as a carrier with low rate at 1ml/minute and 1:10 as the split ratio, Mass range was 40-600 amu, the energy of electron used was 70ev, Electron Ionization (E.I.) was used in the MS and the source temperature Transfer line was 180ºC, 200ºC, 1.0µL was injected for the analysis.

RESULTS AND DISCUSSION
The preliminary phytochemical components from the analysis showed that tannins, phenols, lavonoids, alkaloids, saponins, terpenoids and anthraquinones were present in the ethanolic extract of L.C. stems. Table 1 shows the results obtained from the qualitative analysis.
The effect of Flavonoids on the cell membrane permeability and inhibiting enzymes found on the membranes like phospholipase A2 and ATPase has been reported by Hausteen (1983). The medicinal uses of alkaloids found in the plant stems have been known for centuries, and among them, cytotoxicity is one of their biological properties (Nobori et al., 1994). The presence of terpenoid serves as antiin lammatory and anti-fungal properties. The following inferences have been identi ied, 1. The tannins present in the plant stem L.C. exhibits anti-cancer and anti-in lammation properties.
2. The phenols detected from the stems have antioxidative properties.
3. Saponins compound found in L.C. stems known to possess signi icant anti-cancer properties.
In this present work, different compounds showed various percentages which are mainly due to the geographical location of the plant.
The total Flavonoid content from stems of L.C. by the ethanolic extract was 3.05µg/mg (0.30%) expressed in Quercetin equivalence. Figure 1 shows the standard calibration curve of Quercetin. In the stems of L.C., the phenolic content was estimated to be 30.2 µg/mg (3.02%) expressed in Gallic acid equivalence, and the calibration curve is shown in Figure 2. The tannin content in this study was found to be higher 237.5 µg/mg (23.7%) expressed in tannic acid equivalence. The results of tannin standard calibration curve are shown in Figure 3 and given in the Table 2.

Gas Chromatography-Mass Spectroscopy (GCMS)
The GCMS analysis (Figure 4) from L.C. stem illustrates peaks showing a wide range of compounds. An inceptive inquiry of the stem revealed the identi ication of ifty compounds. Results of the compounds are given in Table 3. The primary component was Pentadecanoic acid, 14-methyl-methyl ester (1.0%) ( Figure 5) and other compounds were present in traces. The different compounds present in the L.C. stem is mainly due to climatic and geographical location of the plant (Policegoudra et al., 2012).

CONCLUSION
The presence of these chemical (Flavonoid, alkaloid, saponin, terpenoid, phenol and tannin) components from phytochemical screening in L.C. stem extract can be used in the different ailments through their anti-in lammatory, anti-cancer, anti-arthritic, antioxidant and antimicrobial properties. Further exploration of the L.C. stem could be achieved in the future by isolation and detecting its active compounds.

Funding Support
Nil

Con lict of Interest
Nil