fried plantain acrylamide estimation

NIGERIAN ANNALS OF NATURAL SCIENCES, VOLUME 12 (1) December 2011 (pp 1 – 7) PRINTED IN NIGERIA

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ESTIMATION OF NON-ENZYMATIC BROWNING AND ACRYLAMIDE FORMATION IN FRIED

PLANTAIN (MUSA BALBISIANA)

MARSHALL A. AZEKE PHD (BONN)1* AND MAUREEN E. CHUKWUEDO2

Department of Biochemistry, Ambrose Alli University, P. M. B. 14, Ekpoma, Edo State, Nigeria1. Department of Chemistry, Ambrose Alli University, P. M. B. 14, Ekpoma, Nigeria.

Corresponding Author1

*corresponding author. e-mail: [email protected], tel: +234 805 6174857

ABSTRACT

This study was conducted to ascertain the occurrence of non-enzymatic browning and to estimate the levels of the potential carcinogen, acrylamide, in fried plantain. Ripe plantain (Musa balbisiana) pulp was first of all, peeled and sliced The sliced pulp was then treated by blanching for 4 min in boiled water or soaking in dilute citric acid solution. The samples from these treatments were then deep fried with vegetable oil for 0, 2, 4, 6, and 8 min. The level of non-enzymatic browning was determined by absorbance at 420 nm. The values obtained were used in a regression equation to estimate the level of the potential carcinogen acrylamide. The results show that ripe plantain samples treated with dilute citric acid gave fried products with the least non-enzymatic browning. This was followed by the samples blanched for 4 min in boiled water. The same trend was also observed in the estimated acrylamide levels. It was also observed that, regardless of treatment, non-enzymatic browning, and hence, acrylamide levels increased with increase in the time of frying. Dipping of plantain slices into a solution of citric acid at moderate concentrations as well blanching may be viable approaches for the minimisation of acrylamide content, in commercial as well as in homemade foods.

Key words: acrylamide, Maillard browning, plantain, frying. INTRODUCTION

Acrylamide is a suspected human carcinogen (IARC, 1993), formed in fried and baked carbohydrate-rich foodstuffs such as potatoes. In 2002, relatively high amounts of acrylamide were detected in thermally processed carbohydrate-rich foods (http://www.slv.se/engdefault.asp retrieved in January 2011). Numerous studies have been performed to gain insight into acrylamide formation (Becalski et al., 2003; Yaylayan et al., 2003; Stadler et al., 2004).

Reducing sugars, in the case of potatoes mainly glucose and fructose, are required precursors for

the conversion of asparagine into acrylamide (Becalski et al., 2003; Yaylayan et al., 2003; Stadler et al., 2004).

Various factors, such as pH, water activity, reactant concentrations and the ratio between reactants, temperature and time of processing influence the formation of acrylamide in foods (Franke et al., 2005; De Vleeschouwer et al.,

2006). Although in Europe and in the Americas much work has been done on non-enzymatic browning and acrylamide formation in foods, such as wheat flour, potato, coffee and apple (Gökmen and Şenyuva, 2006), Kiwifruit (Wong

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and Stanton, 1989), pear (Cornwell and Wrolstad, 1981), not much has been done on foods in Africa in general and Nigeria in particular. Most of the staple foods in Nigeria are derived from high carbohydrate crops such as cassava, yam, cocoyam, sweet potato, plantain, maize and several others that are locally cultivated. They are frequently processed using simple traditional means, such as boiling, deep fat frying, roasting and baking. Baking, roasting and deep fat frying are high temperature processes that have been confirmed to induce non-enzymatic browning and acrylamide formation (Becalski et al., 2003). For the health

and well being of the consumers, it is important to establish a baseline database for the occurrence of acrylamide in the dry processed foods in Nigeria, commonly eaten after high temperature processing. Consequently, the objective of this work is to study the occurrence of non-enzymatic browning and to estimate the levels of acrylamide in fried plantain. MATERIALS AND METHODS

Sample Preparation Samples of ripe plantain (Musa balbisiana) were purchased from Ekpoma Main Market in Edo State, Nigeria. Refined vegetable oil (Grand Soya brand, UAC Nig. Plc, Lagos, Nigeria) was used for deep frying of plantain slices. The plantain samples were peeled and sliced to uniform sizes of approximately 7.50 mm, measured using a micrometer screw gauge. The sliced plantain pulps were divided into four portions. The first portion was subjected to deep frying for 2, 4, 6 and 8 min. The second portion was blanched in boiled water for 4 min before deep frying for 2, 4, 6 and 8 min. The third portion was soaked in 0.5 % citric acid solution for 4 min before deep frying, while the forth portion was not pre-treated and not fried. All samples were subsequently dried in the oven at about 60oC for 18 h. The dried

samples were milled to a particle size of ≤ 0.5 mm and then stored in air-tight containers prior to usage. Solvent selection

The efficiency of four different solvents in extracting browning pigments of fried plantain was determined by extracting the fried, dried and milled plantain with distilled water, 0.2 M NaOH, 1% HCl and 50% ethanol. Determination of non-enzymatic browning

The method of Hendel et al. (1950) was adopted

for the determination of non-enzymatic browning with slight modification. One gram each of sample flour was measured into a 250 ml Erlenmeyer flask and 50 ml of distilled water added to each sample. The samples were allowed to stand for 1 h and filtered using 595 Rundfilter filter paper. The filtered extracts were centrifuged (TOMY CX 250 model) for 30 min at 23,000g. The clear supernatants were acidified with 0.5 ml of 40% v/v acetic acid. The non-enzymatic browning was measured at a wavelength of 420 nm in the Shimadzu spectrophotometer UV-120 against a blank. Sample lots were triplicated with five spectrophotometric readings per replicate. ESTIMATION OF ACRYLAMIDE

The acrylamide level of fried ripe plantain was estimated using the acrylamide estimation curve of Quayson and Ayernor (2007) (figure 1). The values obtained for the non-enzymatic browning for the processed plantain were plotted against their corresponding acrylamide values as reported by Quayson and Ayernor (2007). The graph obtained was used to estimate the acrylamide values of the test samples.



Figure 1: Acrylamide Estimation Curve (Source: Quayson and Ayernor, 2007) Statistical analysis Significance of difference was tested by analysis of variance and Tukey-Kramer test using the GraphPad InStat version 2.04a (GraphPad Software Inc, San Diego, California,

USA, www.graphpad.com). Statistical significance was set at P<0.05. RESULTS

From Fig. 2, NaOH solution was found to be the singular solvent that gave the highest spectrophotometric reading at 420 nm followed by water. This was followed by water while HCl gave the lowest reading.

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Figure 2: Browning extraction (OD 420) efficiency of different solvents

Table 1: Browning levels (OD420) of pre-treated ripe plantain fried at different times

Time of frying (min)

Pre-treatment 0 2 4 6 8

Untreated 0.20a ± 0.01 0.29a ± 0.01 0.36a ± 0.05 0.66a ± 0.03 1.12a ± 0.03

Boiled Water 0.18b ± 0.00 0.20b ± 0.03 0.32b ± 0.03 0.56b ± 0.03 1.04b ± 0.14

Citric acid 0.10c ± 0.00 0.15c ± 0.05 0.21c ± 0.04 0.25c ± 0.03 0.39c ± 0.02

Results are means of three independent determinations ± SD; Values in a column with different alphabets are significantly different (P<0.05). In table 1 the general observation is that browning increased with time of frying for all the untreated and treated samples. Blanching for 4 mins in boiled water resulted in a significant reduction in the degree of browning (P < 0.05). The reduction in browning was even more pronounced with samples treated in dilute citric acid (table 1). For example, the untreated samples fried for 2 min had an absorbance of

0.29 while samples soaked in boiled water or citric acid recorded absorbance of 0.20 and 0.15 respectively. Table 2 shows the acrylamide estimation values of fried plantain, based on the regression of non-enzymatic browning on acrylamide formation. The highest estimated acrylamide was recorded by the untreated sample fried for 8 min. The samples treated with citric acid before frying had the lowest estimated acrylamide followed by the samples blanched in boiled water for 4 mins.



Table 2: Estimated acrylamide levels (ppb) in ripe plantain fried at different times

Time of frying (min)

Pre-treatment

0 2 4 6 8

Untreated 1861.9a ± 45.2 2563.5a ± 21.3 3109.2a ± 54.1 5447.6a ± 98.6 9033.2a ± 43.5

Water 1706.1b ± 56.1 1861.9b ± 59.5 2797.3b ± 87.2 4668.1b ± 101.5

8409.62b ± 99.8

Citric acid 1082.5c± 44.2 1472.2c ± 69.7 1939.9c ± 86.5 2251.7c ± 103.4

3343.0c ± 105.6

Results are means of three independent determinations; Values in a column with different alphabets are significantly different DISCUSSION

Although many studies have been done on the non-enzymatic browning and acrylamide content of carbohydrate rich foods, especially in Europe and America, such as potato- and wheat-based foods, there is still little information on these carbohydrate rich foods produced in Nigeria. The finding that NaOH extract gave the highest spectrophotometric reading is not consistent with the report of Quayson and Ayernor (2007) who found that water extract of non-enzymatic reaction products was higher than alkaline extract. This may be due to differences in samples used for assay. However, some other reports have indicated that extraction of browning products and acrylamide at alkaline pH was more efficient than at neutral pH (Eriksson and Karlsson, 2006; Mustafa et al., 2008; Gökmen et al., 2009). Although the alkaline extract gave the highest spectrophotometric reading in the present study, water remains the most appropriate extraction solvent for non-enzymatic reaction products. This is because at alkaline pH secondary Maillard reaction can occur, thereby exaggerating spectrophotometric readings (Martins et al., 2001). The effects of temperature and duration of heating were studied by Maillard (1912), who reported that the rate of the reaction increases with temperature. Many workers have confirmed this observation (Labuza et al., 1994; Ikan, 1990; O’Brien et al., 1998). An increase in

temperature leads to an increase of the reactivity between the sugar and the amino group. Ripe plantain was used in this study

because it contains more sugar than the unripe plantain. Quayson and Ayernor (2007) however, reported that products from unripe plantain browned more than those from ripened plantain. This is difficult to explain considering the fact the sugar is very important for the onset of non-enzymatic browning through pyrolysis, reaction with amino acid (Mazza, 1983; Schallenberger et al., 1959) and caramelisation. The reduction in non-enzymatic browning in the blanched samples may be due to leaching of soluble sugars in the water (Pedreschi et al.,

2009) while the reduced pH of the acid treated samples may have hindered colour formation during Maillard reaction. According Ames (1990), the rate of colour formation during Maillard reaction can be reduced by decreasing the pH. The open chain form of the sugar and the unprotonated form of the amino group, considered to be the reactive forms, are favoured at higher pH. The lower the pH, the more protonated amino group is present in the equilibrium and therefore, less reactive with the sugar. Tareke et al. (2002) showed that increased

heating time for mashed potatoes in a microwave oven increased the amount of acrylamide. Also, in french fries, a similar increase has been reported (Haase, et al., 2003; Rydberg et al., 2003). Becalski et al.

(2003) reported that the level of acrylamide in potato chips increased with increasing frying time. The samples treated with citric acid before frying had the lowest estimated acrylamide followed by the samples blanched



in boiled water for 4 mins. Mestdagh et al.

(2008) reported that chemical pre-treatments such as with citric acid, acetic acid and lactic acid, resulted in a significantly reduced acrylamide content of potato chip. Other methods that have been successfully employed to reduce the acrylamide contents of some processed foods include reduction in temperature and time of processing (Grob et al., 2003), the use of divalent cations (Gökmen

and Şenyuva, 2007) and the use of additives additives (Becalski et al., 2004). CONCLUSION

Based on the results obtained in this study for fried plantain, dipping of plantain slices into a solution of 0.5 % citric acid as well as blanching for 4 min may be viable approaches for the minimization of acrylamide content, in commercial as well as in homemade foods. Citric acid is widely used as a preservative during processing. It could now be used by the food industry to control the formation of acrylamide. Considering the health implications of acrylamide consumption, it would be necessary to subject Nigeria processed carbohydrate rich foods to direct acrylamide assay. This would give a better picture of the level of consumer exposure to acrylamide. REFERENCES

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fried plantain acrylamide estimation

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