In Vitro Antioxidant and Anti-inflammatory Potential Evaluations of Methanolic Extract of Acalypha hispida Burm.f. (Euphorbiaceae)

The principal goal of this research study is to evaluate the antioxidant and anti-inflammatory properties of traditional Bangladeshi medicinal extracts and to examine these activities in relation to their antioxidant content. Acalypha hispida Burm.f. (Euphorbiaceae) is a medicinal plant of Bangladesh and Indian subcontinent, which is widely used as folk medicine for the treatment of many diseases. The aim in the present study was to screen the phytochemical profile and pharmacological activities of methanolic extract of Acalypha hispida leaves. Because each part of Acalypha hispida Burm.f. has different constituents, the pharmacological effects of the plant vary according to the part of the plant evaluated. To investigate pharmacological activities DPPH scavenging assay and HRBC membrane stabilization methods were done for antioxidant and anti-inflammatory potential respectively. The phytochemical analysis of methanolic extract of plant leaves showed that they contained significant presence of flavonoids, phenols, saponins, terpenoids & triterpenes. Alkaloids, glycosides & tannins are also moderately present. Quantitative evaluations show significant presence of phenols than tannin content. The pharmacological studies revealed that the plant extracts may have significant antioxidant effect which is probably mediated by inhibition of DPPH free radical, which is responsible for oxidation. The IC50 values by DPPH scavenging assay observed for standard & leaves were 96.37μg/ml & 493.46μg/ml respectively. There is also moderate anti-inflammatory activity. The IC50 values for anti-inflammatory activity by standard & leaves were 25.03 μg/ml & 843.05 μg/ml respectively.


Introduction
From the very beginning of the civilization, people have depended on nature for their fundamental needs, for the making of li. This one produces in the coastal areas of Bangladesh [1]. Many parts of the plant are used in infectious diarrhoea, pulmonary problems, and in asthma [2]. In Egypt, bishop's weed (Ammi majus) was stated to be used to treat vitiligo, a skin condition categorized by a loss of pigmentation [3]. More recently, a drug (methoxypsoralen) has been produced from this plant to treat psoriasis and other skin disorders, as well as T-cell lymphoma [4]. Alternative medicine demanded to have the healing properties of medicine, but which are contradicted, unproven, or are excessively destructive in relation to their effect [5]. Alternative treatments or identifies are not part of drug or science-based healthcare systems [6]. So, alternative drug consists of a wide diversity of practices, products, and therapiesranging from those that are naturally reasonable and to those with known injurious and toxic properties [7]. Cassia siamea (Lam.) is an extensive medicinal and food plant cultured in Southeast Asia and sub-Saharan Africa. Many traditional claims are stated as medical actions on numerous diseases like constipation, malaria and fevers and jaundice [8]. Moreover, aerial portions of C. siamea are beneficial in ringworm and connected skin diseases [9]. Ethno-botanical examinations also advise antinociceptive and antiviral activities of aerial parts of C. siamea antioxidant and antihypertensive action.
Moreover,a laxative activity, sedative activity and anti-inflammatory of stem bark extract of C. siamea were also stated [10]. Such type actions can be due to the presence of barakol, which has anxiolytic and CNS inhibitory properties.

Plant Material
For this research work, the leaves of A. hispida was collected during July 2018 from the University campus of University OF Chittagong, Bangladesh.

Determination of Total Phenolic Content (TPC)
As it is known that in the alkaline condition phenols ionize absolutely. While Folin-Ciocalteu's reagent is used in this ionized phenolic solution, the reagent will freely oxidize the phenols. Usual color of Folin-Ciocalteu's reagent is yellow and after the oxidation process the solution converts blue. The strength of the color alteration is restrained in a spectrophotometer at 760 nm. The absorbance value will imitate the total phenolic content of the compound [11].

Method of Sample Preparation
In this research work, the total phenolics of the extracts were evaluated using the Folin and Ciocalteu reagent, following the method designated with slight alterations [12]. The test sample (0.2 mL) was variegated with 0.6mL of water and 0.2mL of Folin-Ciocalteu's phenol reagent (1: 1). Subsequently, 5min, 1mL of saturated sodium carbonate solution (8%w/v in water) was added to the mixture and the volume was completed up to 3mL with distilled water. Then the reaction was preserved in the dark for 30min and after centrifuging the absorbance of blue color from dissimilar samples was restrained at 760 nm. All determinations were carried out in triplicate [13].

Method of Sample Preparation
In this research work, fifty micro liters (µl) of tannins extract for each sample was occupied in test tube and volume was completed to 1.0 ml with distilled water. Then, 0.5 ml Folin Ciocalteu reagent was added and varied accurately. Then 2.5 ml 20 per cent sodium carbonate solution was added and varied accurately and kept for 40 minutes at room temperature. Moreover,the optical density was reserved at 725 nm in UV spectrophotometer and concentration was assessed [14].

Phytochemical Screening
In Table 1

Anti-inflammatory Activity
Percent inhibition of protein denaturation was calculated as follows [15]: Control-Sample inhibition= ×100 Control %    In this research work, the method of HRBC membrane stabilization was selected to estimate anti-inflammatory activities ( Tables   4-7).
In this research work, it exposed that the plant extracts might have moderate anti-inflammatory effect which is possibly reconciled by HRBC membrane stabilization.

Antioxidant activity
Here, the free radical-scavenging action of extracts was assessed with the DPPH assay [16]. In this research work, it exposed that the plant extracts may have important antioxidant effect which is maybe reconciled by inhibition of DPPH free radical, which is accountable for oxidation (Tables 8-11).    In this research work, it exposed that the plant extracts may have important antioxidant effect which is maybe reconciled by inhibition of DPPH free radical.

Conclusion
In this research work it was observed that qualitative assessments indication weighty existence of flavonoids, phenols, saponins, terpenoids and triterpenes. It was also observed that alkaloids, glycosides and tannins are moderately present. Quantitative assessments show weighty presence of phenols than tannin content. There is an excellent antioxidant activity in the methanolic extract. There was also moderate anti-inflammatory activity in the methanolic extract of leaves. Each part of Acalypha hispida has altered constituents, the pharmacological properties of the plant differ in relation to the part of the plant assessed. Quantitative assessments show weighty existence of phenols than tannin content.
The IC50 values by DPPH scavenging evaluate observed for standard and leaves were 96.37 µg/ml and 493.46µg/ml separately.
Thus, there is a tremendous antioxidant activity in the methanolic extract. Moreover, there was also moderate anti-inflammatory activity in the methanolic extract of leaves. So, it is evaluated that the IC50 values for anti-inflammatory action by standard and plant leaves were 25.03 µg/ml and 843.05 µg/ml separately.