Journal of Traditional Medicine & Clinical Naturopathy
Open Access

Since time immemorial, plant-based drugs have been used for various diseases ranging from the common cold to cancer. Traditional herbal medicine is an important component that provides primary health care systems in developing countries [1]. Medicinal plants are playing significant role in the daily lives of many people and are an essential part of the South African cultural heritage [2]. The World Health Organization (WHO) has calculated that 80% of the world’s inhabitants utilize traditional medicine for meeting their primary health care needs and the majority of this therapy requires the use of herbal extracts and their active components. The estimation of 20,000 herbal plants used for medicinal purposes in the world [3]. There are 30,000 species of higher plants in South Africa and about 350 species are traded as medicinal plants [2]. In recent years, there has been an increasing interest in botanical description and biological activities from plants used traditionally to treat many diseases and those used as food [4].

Various medicinal plant bioactive extracts and their isolated active constituents have shown different medicinal pharmacological properties against acute and chronic diseases and disorders [5]. Pain is a direct response to a problematic event in association with inflammation, cancer, tissue damage arising from injury [6].

Inflammation, a confined physical condition, is a complex biological response of vascular tissues to invasion or harmful stimulation by an infectious agent, antigen challenge, chemical, physical, or traumatic impairment [7]. The complex cases and mediators involved in the inflammatory reactions can develop many diseases which include rheumatoid arthritis, chronic asthma, multiple sclerosis, inflammatory bowel disease and psoriasis [8]. Free radicals including reactive oxygen species (ROS) and reactive nitrogen species (RNS) raise a highly reactive state that can damage many biomolecules, leading to several chronic diseases like diabetes mellitus, atherosclerosis, inflammation and cancers [9,10]. Non-steroidal anti-inflammatory drugs (NSAIDs) are often used for treating several inflammatory diseases, but these drugs have side effects that include aspirin-induced asthma and disturbances of the gastrointestinal (GI) tract [11,12].

Diabetes mellitus is the most common of the endocrine disorders which is characterized by chronic hyperglycemia due to relative or absolute lack of insulin secretion or actions [13]. Hyperglycemia resulting from an accumulation of free glucose in the blood that leads to a generation of free radicals due to autoxidation of glucose and glycosylation of proteins [14]. Oxidative stress plays a vital role in the diabetic pathophysiology [13]. Moreover, oxidative stress is thought to be a major contributing factor to the development and progression of diabetic complications, which is also associated with insulin resistance and impaired insulin secretion that results in the development of diabetes mellitus [15].

Currently, there are approximately 347 million people suffering from diabetes, and 90%-95% of all diabetes patients have type 2 diabetes mellitus (T2DM) [16,17]. T2DM results from the ineffective use of insulin, or insulin resistance [17]. Chronic hyperglycemia syndrome, which is a common complication of T2DM, can results from conditions like obesity, kidney failure, nerve damage, fatty liver, hypertension, hyperlipidemia, and cardiovascular diseases [18]. As of 2014, the number of diabetes patients only in China soared to 96 million [16]. Therefore, there is a huge demand for the development of therapeutic agent with minimum side effects for the prevention, control and treatment of diabetes.

The present study investigated the anti-inflammatory and anti-diabetic effects of methanol extract of Pilea microphylla whole plant.

Drugs and chemicals

The standard drug, Metformin hydrochloride was the generous gift samples from Beximco Pharmaceuticals Limited, Bangladesh. Alloxan monohydrate was purchased from Loba Chemie, India. Carrageenan was purchased from Otto Chemika, India. Blood samples analyzed for blood glucose content by using OK meter Match glucose test meter (Hsinchu, Taiwan). Acetic acid was collected from laboratory of Bangladesh University.The standard drug Diclofenac-Na was purchased from Square Pharmaceuticals Limited, Bangladesh.

Experimental animals

Eight weeks-old Swice albino mice (27 g-30 g) purchased from Jahangirnagar University, Dhaka, Bangladesh and were housed in animals’ cages under standard environmental conditions (22°C-25°C, humidity 60%-70%, 12 h light: 12 h dark cycle). The mice were feed with standard pellet diet taken from, Jahangirnagar University, Dhaka. The animals used in this study were cared in accordance with the guidelines on animal experimentation of our institute.

Plant materials

Pilea microphylla plants were collected from Mohammadpur, Dhaka and identified by experts at Bangladesh National Herbarium, Dhaka, Bangladesh.

Drying and grinding

The collected plants were separated from undesirable materials or plants or plant parts. They were dried in the sun for one week after cutting into small pieces. The plant parts were crushed into coarse powder with the help of a suitable grinder. The powder was stored in an airtight container and kept in a cool, dark and dry place until analysis commenced.

Preparation of plant extract

About 350 g of powdered sample was taken in a clean, flat-bottomed glass container and soaked in 1200 ml of 90% methanol. The container with its contents was sealed and kept for a period of 10 days accompanying occasional shaking and stirring. The whole mixture then underwent a coarse filtration by a piece of clean, white cotton material. Then it was through whatman filter paper. The filtrate was kept in an open space to evaporate the solvent thus crude extract was obtained. Fine powders of the plant of Pilea microphylla are dissolved in 90% methanol and then evaporation the solvent.

Anti-inflammatory activity

Inflammation (paw edema) was induced by injecting 0.1 ml of 1% Carrageenan in physiological saline into the sub plantar tissues of the left hind paw of each mouse [19]. The methanol extract of Pilea microphylla 250 mg/kg and 500 mg/kg were administered orally 30 min prior to Carrageenan administration. The paw edema size was measured at 0, 1, 2, 3 and 4 hours by using dial caliper [20]. The percentage reduction of paw edema in drug treated group was compared with the control group. Diclofenac sodium (10 mg/kg p.o.) was used as reference standard. 0 (zero) hour reading was considered as an initial normal paw size. Data was collected from the paw thickness and percentage reduction of paw edema of the treated animals. Percentage reduction of paw edema was calculated by using the formula.

Anti-inflammatory activity (%)=(1-T/C) × 100

Where T is the change of paw diameter in treated group and C is the change of paw diameter in control group.

Anti-diabetic activity

This is done by oral glucose tolerance test (OGTT) in diabetic mice. After fasting 16 hours, diabetes was induced into mice by in intraperitoneal injection (i. p.) of alloxanmonohydride (90 mg/kg) dissolved in saline. After 72 hours, plasma glucose levels were measured by glucometer using a blood sample from tail-vein of mice. Mice with blood sugar higher than 8.5 mmol/L were considered as diabetic. Agematched healthy mice were used to examine the effects of extract on normal mice [21].

All the mice were divided into 4 groups, each group containing 5 mice. The divided groups are NC (normal control), DC (diabetic control), STD (diabetic mice receiving 100 mg/kg Metformin), ME (diabetic mice receiving 250 mg/kg and 500 mg/kg methanol extract). The mice were fasted over-night and next day blood samples were taken from all groups of animals to estimate fasting blood glucose level (0 min). All mice received 1 gm/kg glucose, after 1 h of feeding of extract and drug and four more blood samples were collected at 30,60,90 and 120 minutes intervals and blood glucose level was estimated in all the experiments by using OK meter Match glucose test meter [22].

Extract of Pilea microphylla reduced inflammation in mice

Values were expressed in (Mean ± SEM) value. Each group comprised 4 animals. Control Group received normal water and standard group received 10 mg/kg Diclofenac sodium. Extract administered group was treated with 250 mg/kg and 500 mg/kg of the crude extract of Pilea microphylla (Tables 1, 2 and 3, Figures 1 and 2).

Table 1: Anti-inflammatory effects of the methanol extract of Pilea microphylla extract on 1% Carrageenan induced paw edema on mice.

Table 2: % inhibition of paw volume.

Table 3: Plasma glucose level (mmol/L) of Pilea microphylla extract in alloxan-induced diabetic (mM/L) in mice.

Figure 1: Effects of the methanol extract of Pilea microphylla on Carrageenan induced paw edema in mice.

Figure 2: Percent of inhibition effects of the methanol extract of Pilea microphylla on Carrageenan induced paw edema in mice.

Anti-diabetic effec t o f Pile a microphyl l a extr ac t in allox aninduced diabetic mice

Values were expressed in (Mean ± SEM) value. Each group comprised 4 animals. Control Group received normal water and standard group received 100 mg/kg Metformin Hydrochloride. Extract administered group was treated with 250 mg/kg and 500 mg/kg of the crude extract of Pilea microphylla (Table 4, Figures 3 and 4).

Table 4: % lessening of plasma glucose level.

Figure 3: Effects of the methanol extract of Pilea microphylla on OGTT test in mice.

Figure 4: % lessening of plasma glucose level by the methanol extract of Pilea microphylla on OGTT test in mice.

Alternative medicine is getting more popular for the treatment of inflammation [23]. Plant based drugs have become the main focus of current research due to the greatest therapeutic potential without many of the side effects associated with synthetic drugs [7]. A wide variety of mediators (e.g nitric oxide, eicosanoids and pro- inflammatory cytokines) have been associated critically in the development of inflammatory diseases [24]. Most of the phytochemicals that exert anti-inflammatory effects have been studied extensively for their potential to inhibit pro-inflammatory cytokines [25,26]. Methanol extract of Pilea microphylla extract has been shown to possess good anti-inflammatory potential.

T2DM is a common form of diabetes which is caused by insufficient use of insulin that eventually results in hyperglycemia [27]. Hyperglycemia has abnormal impacts on normal physiological functions in the body as well as results in macro and micro-vascular dysfunction in diabetes [28]. At present available anti-diabetic agents include sulphonylureas and the rapidly-acting secretagogues that stimulate insulin secretion, biguanides that is associated to reduce hepatic glucose production, α-glucosidase inhibitors which delay digestion and absorption, and thiazolidinediones which promotes insulin action. Although there are a large number of drugs to treat T2DM, many of these may cause drug tolerance and severe secondary side-effects such as hepatic disease [29,30]. The present investigation, concentrate of Pilea microphylla had set apart in hypoglycemic impacts in diabetic mice (Tables 3 and 4).

These finding suggests that administration of extract of Pilea microphylla is beneficial in reducing blood glucose levels as well as to alleviate inflammation which may arise from different acute and chronic diseases. However, further research is warranted to find out whether it does modulate the insulin secretion or directly affect the blood glucose in terms of metabolism. Status of cytokines and other targets in the pathway of developing inflammation should be studied following administration of Pilea microphylla extract to elucidate the mechanism of its anti-inflammatory effect.