Phytochemicals in Funaria Sp.: A Correlative Analysis for Therapeutic Potential

One of the oldest groups of terrestrial plants, bryophytes date back to the Ordovician epoch (488–444 mya). Mosses were thought to be aborted plant fetuses by some scientists in the 1600s (Crum, 2001). Since ancient times, bryophytes have been utilized in traditional Chinese, Indian, European, and North American medicine to treat burns, bronchitis, tonsillitis, tympanitis, skin disorders, and cardiovascular problems (Khanam et al., 2011). Bryophytes offer a variety of biological activities, including antibacterial, antioxidant, antitoxic, NO production inhibitory, and muscle relaxing qualities, according to their phytochemical composition. Asakawa (2007) Tannins, alkaloids, saponins, cardiac glycosides, steroids, terpenoids, favonoids, phlobatannins, anthraquinones, and reducing sugars are the primary phytochemical constituents found in therapeutic plants. Phytochemical components are generally classified into two groups based on their metabolism activity in the plant: primary constituents, which primarily consist of sugars, amino acids, proteins, and chlorophyll, and secondary constituents, which include alkaloids, favonoids, saponins, tannins, phenolic compounds, and many more. Krishnaiah (2007) Numerous moss species have been isolated in recent years, and it has been found that they include terpenoids, benzoic, pthalic, cinnamic, and a few aromatic compounds that contain nitrogen. These compounds can occasionally share structural similarities with those found in vascular plants (Asakawa et al. 2012).There have been reports of flavonoids, terpenoids, alkaloids, and phenolic compounds. According to a survey of the literature, the phytochemical characteristics of the bryophytes present in the Gautala wildlife sanctuary are not well supported. This study focused on the phytochemical effectiveness of Funaria sp.

MATERIAL AND METHOD:

Sample collecting and plant powder preparation:

The funaria sample was collected from the Gautala wildlife reserve in the Jalgaon region. The plant fragments were collected, cleaned under running water, and then autoclaved to remove any last bits of debris. Fresh plant leaves were allowed to dry at room temperature for 10 days before being crushed in an automated grinder. The dust-free plant powder was stored at 4 °C in an airtight container for later usage (Mehra and De, 2017). After drying, the materials were ground into a fine powder.

Quantitative determination of secondary metabolites:

 Determination of Alkaloids:

This was determined using the method of Nbaeyi-Nwaoha and Onwuka (2014). Twenty milliliters of 20% H2SO4 in ethanol (1:1) were used to dissolve one gram of each of the ground samples, and they were then filtered. Five milliliters of 40% H2SO4 were added to two separate test tubes containing one milliliter of each of the filtrates (leaf and bark), and the mixture was thoroughly mixed. Before taking the measurement, the mixture was covered and left to settle for four hours. The two samples (leaf and bark) were measured at 568 nm using a spectrophotometer. Values were recorded after the process was carried out three times.

Determination of Flavonoids:

The approach of Nbaeyi-Nwaoha and Onwuka (2014) was used to determine this. 200 milliliters of ethyl were used to dissolve one gram of each of the powdered materials, which were then filtered. Five milliliters of the leaf and bark filtrates were placed in two separate test tubes. Five milliliters of diluted ammonia were then added, thoroughly mixed, and let to settle for a few hours. A spectrophotometer was then used to detect the absorbance at 490 nm (Nbaeyi-Nwaoha and Onwuka, 2014). Values were recorded after the process was carried out three times.

Determination of Tannins:

The approach of Nbaeyi-Nwaoha and Onwuka (2014) was used to determine this. One gram of each sample was placed in a separate conical flask, shaken for 30 minutes at 5-minute intervals, and then filtered after 10 milliliters of water had been added. Two distinct conical flasks were filled with a constant amount (2.5 ml) of each filtrate. One milliliter of FollinDenis reagent and Na2CO3 were then added, and everything was well combined. A spectrophotometer was used to measure the absorbance at 720 nm after the mixture had settled for 90 minutes at room temperature. Values were recorded after the process was carried out three times.

Determination of Saponins:

In separate conical flasks, 1g samples were mixed with 10 ml of petroleum ether following the NbaeyiNwaoha and Onwuka (2014) method. After being drained and dried, this was combined with ten milliliters of petroleum ether. Measurements were collected at 550 after adding 5 ml of ethanol to the dry mixture and completely mixing it. Each sample received around 2 ml of the mixture, which was then placed in two separate test tubes and left to settle for 30 minutes. The values were recorded after the process was carried out three times.

Determination of Glycosides:

The approach of Nbaeyi-Nwaoha and Onwuka (2014) was used to determine this. In test tubes, 1 g of materials were thoroughly mixed with 2.5 ml of 15% lead acetate before being filtered. After carefully mixing and letting it settle, two milliliters of chloroform were added to the filtrate. The lower part was gathered and dried by evaporation. After adding three milliliters of glacial acetic acid, 0.1 milliliters of 5% ferric chloride, and 0.25 milliliters of concentrated H2SO4, the dried lower part was thoroughly mixed and allowed to sit for three hours. A spectrophotometer was used to measure the absorbance at 568 nm. The values were recorded after the process was carried out three times.

Determination of Total terpenoid content:

The terpenoid content was calculated using the Ghorai et al. (2012) method. After adding 1.5 mL of chloroform to 200 μL of plant extract, the mixture was thoroughly vortexed. Following the remaining three minutes, 100 μL of concentrated sulfuric acid was added, and the mixture was left to sit at room temperature in the dark for one and a half to two hours. A reddish-brown precipitate formed after incubation, and the supernatant was carefully removed from the solution. The precipitate was thoroughly blended with 1.5 mL of methanol. Using a spectrophotometer, absorbance was measured at 538 nm, with linalool serving as the standard. The amount of terpenoid was stated in milligrams of linalool equivalents per gram (LE/g). of sample in dry weight.

Determination of Total Phenol content:

One milliliter of 80% ethanol was mixed with 0.5 grams of plant extract to quantify total phenolics. After that, the mixture was centrifuged at 12,000 rpm for 15 minutes. The supernatant was then stored in a test tube, and the procedure was carried out six times. Following collection, the supernatant was put in a water bath to dry. The supernatant's volume was increased to three milliliters by adding distilled water. This solution was supplemented with 2 milliliters of 20% Na2CO3. 0.5 ml of Folin Ciocalteau regent was added to this, followed by the addition of 2 ml of (Na2Co3) from 20% Na2Co3 solutions after 5 minutes.  After thoroughly mixing the solution, the test tube was placed in the water bath with boiling water. Their absorbance was measured at 650 nm. As a benchmark, the Catechol was employed (Hagerman et al., 2004).

Statistical analysis:

M.S. Excel was used for all statistical analyses. The phytochemical concentrations in Funaria sp. were subjected to descriptive statistics, which included a mean. The Shapiro-Wilk test for a normal distribution of data was used to evaluate the data's normality. The association between various phytochemicals was assessed using Pearson's correlation analysis. The significance level for all statistical tests was set at 5% (α = 0.05).

Observation Table: