In food research, phenolic compounds with antioxidant activity holds an important position. This is related to the fact that phenolic compounds play a beneficial effect on preventing and/or treating various diseases, such as cancer, diabetes, cardiovascular and neurodegenerative diseases, among others (Cory et al., 2018; Panche et al., 2016; Pandey & Rizvi, 2009). Phenolic is an important bioactive and possess strong antioxidant activity, and fruits and vegetables usually are the primary sources of Phenolic compound (Tungmunnithum et al., 2018). It is widely believed that the ingestion of fresh fruits and vegetables is related to the reduction of cardiovascular and cancer diseases (Aghajanpour et al., 2017).

Pear (Pyrus spp) have been known to contain a number of phytochemical compounds with important bioactive properties like phenolics and flavonoids, which are essential for their health benefits (Öztürk et al., 2015). Those health benefits of polyphenol consumption derive from their antioxidant and anti-inflammatory properties (Borges et al., 2010). Other pro-health properties that are attributed to pears are related to the content of triterpenoids, due to their antioxidative, anti-inflammatory, and anticancer properties (Reiland & Slavin, 2015).

During recent years, some researchers have been focused on analyses and comparison of the nutritional components contained in the edible part of pear fruit such as total sugars, vitamins, minerals, and so on (Slavin & Lloyd, 2012; Kahle et al., 2005; Tanriöven & Ekşi, 2005). A part from some common reported compounds such as arbutin, chlorogenic acid, catechin, quercetin, kaempferol, various hydroxycinnamoylmalic acids, and their ethyl esters, hydroxycinnamoyl malates, procyanidins, and triterpenes compound has also been found in the peel of pear (Lee et al., 2011a; Lee et al., 2011b; Ma et al., 2012).

Pears is one of the fruits imported into Indonesia which is often found in supermarkets or traditional markets. Pears which are widely circulating in Indonesia are Pyrus communis or green pears/European pears and Pyrus pyrifolia or yellow pears/Asian pears (Novera et al., 2015). Pears are usually consumed along with the peels, although consuming the fruit while discarding the peels are also common, because the peels are often viewed as non-beneficial (Sagar et al., 2018). With this fact, understanding the potentials and bioactive compounds of pear peel as a side product of the fruit is very important (Li et al., 2014).

Based on our study in this matter, we found that the information on the composition and constituents of pear peel are limited. Therefore, we propose that it is important for food products research to have a systematic study on pear peel, especially on the phenolic and flavonoid compounds. The aim of this research is to identify the total phenolic and antioxidant contents, and performing qualitative determination of phytochemical contents for the two pear varieties.

Tools and material

The main material used in the study are P. communis and P. pyrifolia. Both samples of pears were obtained from Caringin’s Traditional Market, Bandung, West java that imported from Australia and China, respectively. Other materials used were methanol, ethyl acetate, n-hexane, ethanol, ascorbic acid, gallic acid, quercetin, Folin-Ciocalteu’s phenol reagent (Sigma), aquadest, chloroform, toluene, acetone, 25% ammonia 2 N, 10% hydrochloric acid, Dragendorff reagent, Mayer reagent, Mg powder, amyl alcohol, sodium hydroxide, gelatin, Fe (III) chloride 1%, and anhydrous acetic acid. The tools used are sifters, cups, oven, desiccators, maceration chamber, erlenmeyer flasks, vacuum rotary evaporator, analytical scales, and micropipette. Spectrophotometric measurements were performed on ultraviolet (UV)-1600 spectrophotometer (Shimadzu).

Sample preparation

Both pear peel samples that have been collected and cleaned are dried by drying the cabinet at 40-50°C until smooth and dry.

Extractions

Pear peels were extracted by maceration in 200 ml of n-hexane, ethyl acetate and methanol at room temperature for 24 hours. Residues and extracts are separated by filtering using filter paper. The residue obtained was re-extracted twice with a fresh portion of the extraction solvent. Extracts obtained from the three extraction processes were combined and excess solvents were concentrated using a vacuum rotary evaporator at 50°C. Semisolid extracts obtained quantitatively were transferred to extraction solvents and stored at 10°C until used for further experiments.

Phytochemical screening

Phytochemical compounds were carried out for all extracts with the following methods:

Steroids and terpenoids

As much as 2 ml of extract was added to 2 ml of acetic anhydride and the concentrated H2SO4. The formation of blue with green rings indicates the presence of terpenoids and the brown rings indicate the presence of steroids (Ayoola et al., 2008).

Alkaloids

Extracts were dissolved individually in dilute hydrochloric acid and filtered. Filtrates were treated with Dragendroff’s reagent (solution of Potassium Bismuth Iodide). The Formation of red precipitate indicates the presence of alkaloids (Altemimi et al., 2017).

Saponins

As much as 0.5 g of extract was shaken with 2 ml of water. If foam produced persists for ten minutes it indicates the presence of saponins (Altemimi et al., 2017).

Tannins

To the extract, 1% gelatin solution containing sodium chloride was added. The Formation of white precipitate indicates the presence of tannins (Altemimi et al., 2017).

Phenolics

Extracts were treated with 3-4 drops of ferric chloride solution. The Formation of bluish-black color indicates the presence of phenols (Altemimi et al., 2017).

Flavonoids

Extracts were treated with a few drops of sodium hydroxide solution. Formation of intense yellow colour, which becomes colourless on the addition of dilute acid, indicates the presence of flavonoids (Altemimi et al., 2017).

Determination of total phenolic contents

Total polyphenol was determined by Folin-Ciocalteu method (Aryal et al., 2019). A dilute extract of each plant extract (0.5 ml of 1 : 10 g/ml) or gallic acid (standard phenolic compound) was mixed with Folin-Ciocalteu reagent (5 ml, 1 : 10 diluted with distilled water) and aqueous Na2CO3 (4 ml, 1 M). The mixtures were allowed to stand for 15 minutes and the total phenols were determined by colorimetry at 765 nm. The standard curve was prepared using 0, 50, 100, 150, 200, and 250 mg/L solutions of gallic acid in methanol : water (50 : 50, v/v). Total phenol values are expressed in terms of gallic acid equivalent (mg/g of dry mass), which is a common reference compound.

Determination of total flavonoid contents

Aluminum chloride colorimetric method was used for flavonoids determination (da Silva et al., 2015). Each plant extracts (0.5 ml of 1 : 10 g/ml) in methanol were separately mixed with 1.5 ml of methanol; 0.1 ml of 10% aluminum chloride; 0.1 ml of 1 M potassium acetate; and 2.8 ml of distilled water. It remained at room temperature for 30 minutes. The absorbance of the reaction mixture was measured at 415 nm with UV-Vis spectrophotometer. The calibration curve was prepared by preparing quercetin solutions at concentrations 12.5 to 100 g/ml in methanol.

Data analysis

Results are given as the mean of three independent determinations ± standard deviation using SPSS version 19.0. The data were statistically analyzed by ANOVA and t-tests. The level of statistical significance was set at p <0.05.

Phytochemical screening

Phytochemical screening of the two varieties of peel pears shows that both have the same content, namely phenolic, flavonoids, tannins, saponins, and alkaloids as shown in Table I. All of these phytochemicals are important elements of herbal medicine and are directly related to various health-promoting activities such as anticancer, antifungal, anti-inflammatory, antihyperglycemic, immunomodulatory, and wound healing properties (Forni et al., 2019). However, flavonoid content in P. communis is greater than P. pyrifolia.

Flavonoids have been found to possess antitumoral, antiallergic, and anti-inflammatory activities (Panche et al., 2016). Phenolic content is the greatest content found in both P. communis and P. pyrifolia. In particular, many phenolic compounds have been identified in pear fruits such as arbutin, chlorogenic acid, hyroxycinnamoyl malates, catechins, and procyanidins (Sun et al., 2019).

Table I. Qualitative phytochemical screening in P. communis and P. pyrifolia peels

Note: (++) highly presence; (+) presence; (-) absence

Total extraction yields of pear peels

In P. communis peels, total extraction yields from n-hexane, ethyl acetate, and methanol were 11.35%; 17.95%; and 30.85%, respectively as shown in Table II. In P. pyrifolia peels, total extraction yields were 13.50%; 18.75%; and 37.50%, respectively as presented in Table III. In both P. communis and P. pyrifolia peels, the highest yields were observed in methanol extract and higher in the P. pyrifolia peel extracts. The extractable compounds from various plant materials are strongly dependant on the nature of extractable solvents. Methanol is one of the polar solvents used for active component extraction which can attract many compounds such as anthocyanins, terpenoids, saponins, tannins, xanthoxyllines, quassinoids, lactones, flavones, phenones, and polyphenols (Mujeeb et al., 2014).

Table II. Total extraction yields of P. communis peels

Table III. Total extraction yields of P. pyrifolia peels

Total phenolic contents

The antioxidant activity of plant phenolics is due to the reactivity of phenol moieties (hydroxyl group on the aromatic ring) which have the ability to scavenge free radicals via hydrogen donation or electron donation or electron donation. Total phenolics were determined by Folin-Ciocalteu reagent method. In this method, phenols from a blue-colored phosphomolybdic-phosphotungstic complex in the presence of an alkaline solution (Saeed et al., 2012).

The total phenolic contents in P. communis peels were 7.20 ± 0.03 mg/g; 33.35 ± 0.57 mg/g; and 68.86 ± 0.78 mg from n-hexane, ethyl acetate, and methanol extracts, respectively. Statistically significant the lowest yields were observed in n-hexane extracts as compared to ethyl acetate and methanol extract while the latter two have the same significantly different too. In the case of P. pyrifolia peels, the contents from n-hexane, ethyl acetate, and methanol extracts were 8.45 ± 0.11 mg/g; 31.22 ± 0.68 mg/g; and 63.50 ± 0.41 mg/g, respectively. Statistical significance was similar to P. communis peels as presented in Figure 1. Methanol extract have the highest content which were higher in the P. communis peels (68.8 mg GAE/g) than in the P. pyrifolia peels (63.5 mg GAE/g). This result is in agreement with Singh & Rajini (2004) which reported that the maximum antioxidant yield was obtained with methanol compared to acetone and water.

Figure 1. Total phenolic contents of P. communis and P. pyrifolia peels by solvents

Total flavonoid contents

The total flavonoid contents in P. communis peels from n-hexane, ethyl acetate, and methanol were 11.72 ± 0.72 mg/g; 49.75 ± 1.72 mg/g; and 62.84 ± 0.77 mg/g, respectively. In the case of P. pyrifolia peels, the observed contents were 10.91 ± 0.14 mg/g; 40.69 ± 0.84 mg/g; and 50.9 ± 1.45 mg/g, respectively. Analytic results show that n-hexane, ethyl acetate, and methanol extracts on P. communis peels were significantly different from P. pyrifolia peels. From the result, the highest content was observed in methanol extracts there are 62.84 mg QE/g in P. communis peel and 50.9 mg QE/g in P. pyrifolia peel as presented in Figure 2. Methanol is an effective solvent for polyphenols, and then it is commonly used in the laboratory and in industrial extraction process (Zhang et al., 2018).

Figure 2. Total flavonoid contents of P. communis and P. pyrifolia peels by solvents

It can be seen that the total contents of polyphenol and flavonoid have the same trends. Total phenolic content and total flavonoid contents for both the varieties of pear peels showed that methanol extracts are the highest values and P. communis peels contained more phenolic and flavonoid contents than P. pyrifolia peels.

From this study, it is concluded that two varieties of pear peel show the presence of phytochemicals, there are phenolics, flavonoids, tannins, alkaloids, and saponins. Methanol extracts is the highest amount of total phenol and total flavonoids contents both P. communis and P. pyrifolia peels which P. communis peels exhibited more amounts of phenolic and flavonoids than P. pyrifolia peels. The presence of these bioactive compounds in pears varieties establishes themselves as the sources of natural therapeutic agents that can act as potent free radical scavengers.

This project was supported by Ministry of Research and Technology Republic of Indonesia and Universitas Muhammadiyah Bandung in 2019.

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