Heavy Metals Bounding Ability of Pomegranate (Punica granatum) Peel in Model System

Abstract

Pomegranate fruit is an important sources of natural phenolic compounds. In this study, the influence of pomegranate fruit peel on binding of some heavy metals were established by Inductively Coupled Plasma Atomic Emission Spectrometry (ICP-AES). Also, the total phenolic content of methanol extract of pomegranate fruit peel was determined with the Folin-Ciocalteu method. Amounts of 1, 2, 3, 10, 20, 40, and 60 g of ground peel were used in this experiment. After pomegranate samples were weighed, they were added into 500 ml baker. Each sample was treated by several solutions contained the heavy metal elements at different concentrations. The heavy metal contents in the solutions prepared with ground material were decreased with increasing of ground material. At the same time, the bounding rate of heavy metals increased with the increase of ground peel amounts. Lead (Pb) with 99.2% rate for solution obtained from 20 g ground sample was the highest bound element. It can be observed from these results that the amount of metal ions bound by the ground material differed from on fraction to others.

Keywords:

• Pomegranate
• Punica granatum
• Peel
• Heavy metal
• Bind
• ICP-AES

INTRODUCTION

Pomegranate (Punica granatum) is belong to Punicaceae family, and one of the oldest edible fruits. Fruits are valuable sources of natural phenolic compounds, which are known to have beneficial health promoting properties. It is widely grown in many tropical and subtropical countries.[1–3] The plant favors a semi-arid climate and is extremely drought-tolerant. In several countries, the juice is a very popular beverage. The juice of wild pomegranates yields citric acid and sodium citrate for pharmaceutical purposes. Pomegranate juice enters into preparations for treating dyspepsia and is considered beneficial in leprosy. Leaves, seeds, roots and bark have displayed hypotensive, antispasmodic and anthelmintic activity in bioassay. All parts of the tree have been utilized as sources of tannin for curing leather. The trunk bark has 28 g/100 g tannin content, the leaves, 11 g/100 g and the fruit rind as much as 26 g/100 g.[4,5]

Dietary fiber consist of a variety of components, including pectin, hemicellulose, cellulose, lignin, flavonoids, condensed tannins, and hydrolyzable tannins.[6–10] Dietary components differ not only in chemical structure or physiology activity from one vegetable material to another, but also in their capacity for microelement binding.[10] Hemicellulose and pectins have a remarkable ağabeylity to bind heavy metal compounds, which is promising for the promotion of health via fiber-containing food.[11–15] The sorbtion capacity of dietary fiber depends on its chemical structure and proportion of particular elemnts.[11] The literature contains many references to the binding capacity of dietary fibers and dietaryfiber components under simulated gastrointestinal conditions.[16–20] According to Fadavi et al.,[3] the pomegranate juices also contained some important minerals. Concentrations of Mn, Cu, Fe, Zn, Pb, and Cd and the ranges of variations for these minerals were 0.012–0.021, 0.013–0.081, 0.03–0.21, 0.037–0.084, 0.021–0.112 ppm, and trace amounts, respectively. The major class of phytochemical present in pomegranate is the polyphenols and includes flavonoids, condensed tannins and hydrolysable tannins. Hydrolyzable tannins are predominant polyphenols found in pomegranate juice and account for 92% of its antioxidant activity.[8] Studies have also shown that the antioxidant capacity of pomegranate juice is three times that the popular antioxidant-containing beverages such as red wine and gren tea, presumably due to the presence of hydrolyzable tannins in the rind, along with anthocyanins and ellagic acid derivatives.[8]

There is an increased concern in the fruit juice industry about the avbailability of high juice yield pomegranate cultivars with suitable juice composition. So there will be excessive product waste peel from the juice industry. The aim of this work was to establish bounding ability of bind heavy metals of pomegranate fruit peel waste.

MATERIALS AND METHODS

Material

Pomegranate fruits were purchased from the local market in Konya in Turkey in 2007. All of the fruits were brought to the laboratory at the same time. Diseased, bruised and injured fruits were removed from others. Uniform size and appearance were randomly distributed into different lots. Also, all chemicals and solvents were analytical grade and obtained from Merck (Darmstadt, Germany).

Fruit Handling and Pre-treatment

The pomegranate fruits were kept refrigerated (+4°C) during analyses. Fruits for analyses were peeled by hand. The total phenolic content of methanol extract of pomegranate fruit peel was estimated with the Folin-Ciocalteu method.[21] Portions of 1, 2, 3, 10, 20, 40, and 60 g of dried and ground peels were used in experiment. Pomegranate samples were weighed in an appropriate portions and added into 500 ml conical flasks which were then treated with amounts of appropriate model solution. Concentrations of metal ions were measured by ICP-AES. The peels were dried in oven (70°C) (MMM Medcenter GmbH D-82152 Planegg/München/Type: Venticell 222, Seri No:960325, Germany). The ground materials were separately added into beaker with 500 ml volume, which were then treated with amounts of appropriate model solution. Standard solutions contained heavy metals were added into sample. Formulations (A, B and C) are given as; (A) 10 g ground material was added into 200 ml 5 ppm Standard solutions, 20 g ground material was added into 200 ml 5 ppm Standard solutions; (B) 40 g ground material was added into 200 ml 2 ppm Standard solutions, 60 g ground material was added into 200 ml 5 ppm Standard solutions; (C) 1 g ground material was added into 50 ml 5 ppm Standard solutions, 2 g ground material was into 50 ml 5 ppm Standard solutions, 3 g ground material was added into 50 ml 5 ppm Standard solutions. The initial concentration of heavy metals in the Standard solutions were prepared as 2 and 5 ppm for models. Amounts of Standard solutions were separately mixed with each ground and weighed pomegranate sample. Each solutions were measured by ICP-AES. Results were obtained according to curves of standard metal ion concentrations.[22]

Mineral Contents of Samples

The solutions contained 10, 20, 40, and 60 g ground material were shaked for 24 h, and other samples were shaked for 1 h. Distilled deionized water and ultrahigh-purity commercial acids were used to prepare all reagents, standards, and samples. Then, samples were filtrated through whatman No 42. The filtrates were collected in 50 ml Erlenmayer flasks and analysed by ICP-AES. The mineral contents of the samples were quantified against standard solutions of known concantrations which were aralysed concurrently by ICP-AES.[22]

Statistical Analysis

The data were analysed for statistical significance by analyses of variance.[23] The data from experiment were subjected to ANOVA using randomized complete block design with statistical analysis system-ANOVA procedure.[24]

RESULTS AND DISCUSSION

The influence of pomegranate fruit peel on bound heavy metals such as Cd, Cr, Pb, etc. are given in the Tables 1 and 2. The heavy metal binding by the ground pomegarnate in the absence of industrial fractions were determined. It can be observed from these values that the amount of metal ions bound by the ground material differed from one fraction to another of the different concentrations of the ground material, 60 g have bound the largest amounts of Cr, Cd, Ni and Pb. It is worth noting that the polyphenol component of the ground material was found to be 10 g ground bark solutions contained 2.32 ppm Cd, 1.87 ppm Cr, 1.36 ppm Cu, 1.54 ppm Fe, 2.94 ppm Ni, 0.05 ppm Pb and 2.37 ppm Zn. In addition, 20 g ground bark solutions treated with 5 ppm Standard solutions contained 1.64 ppm Cd, 1.40 ppm Cr, 0.87 ppm Cu, 1.04 ppm Fe, 2.58 ppm Mn, 1.78 ppm Ni, 0.04 ppm Pb and 1.89 ppm Zn. As seen in Table 1, heavy metal concentrations of solutions were decreased with increasing of ground bark material. At the same time, the binding rate of heavy metal increased with increasing of ground bark amount (Table 2). The highest bound element was Pb with 99.2% for 20 g ground pomegranate sample.

Table 1 Bounding amount of heavy metals by ground pomegranate peel*

	Heavy metals (ppm)
Sample amounts (g)	Cd	Cr	Cu	Fe	Mn	Ni	Pb	Zn
1	3.25 ± 0.13**	2.85 ± 0.03	2.08 ± 0.02	1.91 ± 0.01	3.27 ± 0.13	2.35 ± 0.09	0.14 ± 0.03	3.14 ± 0.17
2	2.40 ± 0.07	1.92 ± 0.10	1.56 ± 0.01	1.63 ± 0.02	2.85 ± 0.17	2.00 ± 0.08	0.06 ± 0.01	2.44 ± 0.21
3	1.17 ± 0.02	1.57 ± 0.06	1.21 ± 0.03	1.54 ± 0.07	2.49 ± 0.11	1.85 ± 0.10	0.06 ± 0.01	2.15 ± 0.09
10	2.32 ± 0.11	1.87 ± 0.05	1.36 ± 0.01	1.54 ± 0.13	2.94 ± 0.16	2.10 ± 0.17	0.05 ± 0.01	2.37 ± 0.13
20	1.64 ± 0.12	1.40 ± 0.01	0.87 ± 0.01	1.04 ± 0.11	2.58 ± 0.09	1.78 ± 0.02	0.04 ± 0.01	1.89 ± 0.06
40	0.67 ± 0.01	0.72 ± 0.01	0.23 ± 0.01	0.41 ± 0.08	1.06 ± 0.04	0.80 ± 0.01	0.02 ± 0.00	0.89 ± 0.03
60	0.53 ± 0.02	0.62 ± 0.01	0.21 ± 0.01	0.36 ± 0.06	0.91 ± 0.01	0.62 ± 0.01	0.02 ± 0.00	0.80 ± 0.02
*Dry basis; and **mean ± standard deviation.

Table 2 Bounding rate of heavy metals by ground pomegranate peel (%)

	Heavy metals
Sample amounts (g)	Cd	Cr	Cu	Fe	Mn	Ni	Pb	Zn
1	43.0	35.0	58.4	61.8	34.6	53.0	97.2	37.2
2	61.6	52.0	68.8	67.4	43.0	60.0	98.8	51.2
3	68.6	30.6	75.8	69.2	50.2	63.0	98.8	57.0
10	62.6	53.6	72.8	69.2	41.2	58.0	99.0	52.6
20	72.6	67.2	82.6	79.2	48.4	64.4	99.2	62.2
40	64.0	66.5	88.5	79.5	47.0	60.0	99.0	55.5
60	69.0	73.5	89.5	82.0	54.5	69.0	99.0	60.0

In other treatments, 40 and 60 g ground pomegranate peel were used in experiment. Solutions of both samples contained lower heavy metal compared with 10 and 20 g ground pomegranate solutions. Solutions obtained from 40 g ground samples contained 0.72 ppm Cr, 0.67 ppm Cd, 0.23 ppm Cu, 0.41 ppm Fe, 1.06 ppm Mn, 0.80 ppm Ni, 0.02 ppm Pb, and 0.89 ppm Zn. Second solution for 60 g sample contained 0.62 ppm Cr, 0.53 ppm Cd, 0.21 ppm Cu, 0.36 ppm Fe, 0.91 ppm Mn, 0.62 ppm Ni, 0.02 ppm Pb and 0.80 ppm Zn. The solutions of 40 and 60 g levels of ground pomegranate peel were found to be the most effective metal binder.

Also, it was treated to the effect of solutions obtained from 1, 2, and 3 g ground pomegranate peel contained 5 ppm standard solutions in 50 ml. While solutions filtered from 1 g ground sample contain 2.85 ppm Cr, 3.25 ppm Cd, 2.08 ppm Cu, 1.91 ppm Fe, 3.27 ppm Mn, 2.35 ppm Ni, 0.14 ppm Pb and 3.14 ppm Zn, solutions contained 3 g sample contained 1.57 ppm Cr, 1.97 ppm Cd, 1.21 ppm Cu, 1.54 ppm Fe, 2.49 ppm Mn, 1.85 ppm Ni, 0.06 ppm Pb and 2.15 ppm Zn (Table 1). Pb was the highers bound in all samples (Table 2).

Phenolic compounds widely distributed in plants, attract significant scientific interest due to their bio-functional health-promoting properties.[25–31] As a result, of all the ground material used in experiment, 60 g was the most efective and 10 g the least efective metal ion binders. The total phenolic content of methanol extract of pomegranate fruit peel was determined with the Folin-Ciocalteu reagent, and established as 137.3 GAE mg/g. This analysis confirm that pomegranate fruit peel had a higher phenolic content. Al-Musatafa and Al-Thunibat (2008) reported that the total phenolic content of pomegranate peel contained of 98.6 and 103 GAE mg/g in aqueous and methanol extracts with the Folin-Ciocalteu method, respectively. Various phenolic compounds have different responses to Folin-Ciocalteau assay.[32] Previous work found that the HPLC analysis of the methanolic extract of Punica granatum peel have detected some polyphenols, like gallic acid (34.03%) and catechin (3.31%).[33]

Borycka and Zuchouski[11] established that the blackcurrant preparation showed the best binding capacity with respect to cadmium and lead at pH 6.0. As a result, decreasing of heavy metals in solutions is probably due to phenolic compounds of pomegranate bark. Polyphenols were found to bind considerable amounts of lead ions.[32] According to Fadavi et al.,[3] the pomegranate juices also contained some important minerals. Concentrations of Mn, Cu, Fe, Zn, Pb, and Cd and the ranges of variations for these minerals were 0.012–0.021, 0.013–0.081, 0.03–0.21, 0.037–0.084, 0.021–0.112 ppm, and trace amounts, respectively. The content of lead in all solutions was very low.

CONCLUSIONS

Solutions of both samples contained lower heavy metal compared with 10 and 20 g ground pomegranate solutions. The heavy metal concentrations of solutions were decreased with increasing of ground pomegranate material. As seen Table 1, ground pomegranate peel can be used as bind to the heavy metals as filtrating agent in several industry. As a result, of all the ground material used in experiment, 60 g was the most efective and 10 g the least efective metal ion binders. The total phenolic content of methanol extract of pomegranate fruit peel was determined with the Folin-Ciocalteu reagent, and established as 137.3 GAE mg/g. This analysis confirm that pomegranate fruit peel had a higher phenolic content.

ACKNOWLEDGMENT

The authors thank to Mrs Perihan Özcan due to her help for material preparation, and this work was supported by Selçuk Üniversity Scientific Research Project (S.Ü.-BAP, Konya-Turkey).