Effect of Gamma Radiation on Microbial Load and Chemical Constituents of Banana Fruits Stored Under Different Temperatures

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1 Research Journal of Agriculture and Biological Sciences, 6(4): , , INSInet Publication Effect of Gamma Radiation on Microbial Load and Chemical Constituents of Banana Fruits Stored Under Different Temperatures 1 Gamal A. Mohamed and 2 Gihan A. Mahmoud 1 Plant Research Dept., Atomic Energy Authority, Abou Zabal 13759, Egypt 2 Horticultural Res. Inst., Agric. Res. Center, Giza, Egypt. Abstract: Banana fruits were exposed to irradiation doses of 0, 0.25, 0.50, 1.0, 1.5, and 2.0 K-Gy. Irradiated and non-irradiated fruits were stored under different temperatures of 5, 10, 15, and. Samples were taken after different storage periods of 7, 14, 21, 28, 35 and 42 days. For all treatments, decay percent of fruits, total bacterial and fungal counts were recorded. In addition, chemical constituents including starch, total sugars, ascorbic acid and peel chlorophyll content were determined. Results revealed that, storing banana fruits under low temperature had the capacity to reduce the decay and decline or inhibit bacterial or fungal growth. In addition, cold storage retarded the changes in chemical constituents during storage. On the other side, the applied irradiation doses exerted positive for maintaining banana fruits and extending its shelf-life. In this connection, the applied irradiation doses reduced the decay and greatly inhibit the microbial growth (total bacterial and fungal count). Furthermore, irradiation doses exhibited good potential for maintaining the chemical constituents near the normal level and in turn keeping its good quality. Irradiation dose of 1.0 K-Gy was found to be the most effective dose to positively maintain the stored banana fruits during the different storage period. Key words: Gamma radiation, banana, microbial load. INTRODUCTION Banana (Musa Sapientum L.) is very well known in Egypt and in many tropical countries as a popular fruits due to its delicious taste and nutritional values as it contain high amounts of sugars and carbohydrate (around 80%). It is considered as a short life commodity [1] due to a rapid senescence process [2] and highly susceptible to various postharvest pathogens. The most important pathogens of banana fruit are Colltotrichum musae, Lasiodiplodia theobromea, and Fusarium spp. which cause crown rot disease as mentioned by [3]. To overcome this problem, chemical fungicides are usually used to protect banana fruits against these diseases. However, the excessive use of such agrochemicals has resulted in serious problems of toxic chemical residues in the environment and high health risks. Thus, an alternative control strategy should be stimulated to replace the utilization of hazardous toxic fungicides. Recently, the ionizing irradiations are very interested to be applied for reducing postharvest losses caused by pathogen infection and fruit fly infestation [4] and prolonging the shelf life fruits by delaying the ripening and senescence processes as declared by [5]. In this respect, gamma radiation which is generated from cobalt-60 has been employed as a fundamental technique during the last 20 years in an extensive research. Gama radiation is considered as a progressive method to preserve fruits by delaying the biological activities to a great extent and lowering the fruit losses from spoilage, deterioration and extending the shelf-life without adverse impact on the organoleptic and nutritional characteristics. Furthermore, an important advantage of gamma radiation which derives from its short wavelength is its ability to penetrate into the tissues. This enables irradiation to reach not only microorganisms in wounds, but also those located within the host, as quiescent or active infection as explained by [6]. For instant, a dose of 1 K-Gy was found to be sufficient to prevent decay by quiescent infection of Botrytis cinerea for several days without causing fruit injury or changes in the ascorbic acid content [7,8]. Generally, the susceptibility of fruits and vegetables to postharvest infection increases as the [6] ripening and senescence processes progress. Therefore, slow senescence will help to maintain the resistance of plant. Sometimes, irradiation can not kill pathogens completely but it may results in cell damage [9], and directly harm the chromosomal DNA of living cell [6]. The damaging of nuclear DNA may cause a mutagenesis and some genetic materials of pathogens probably mutate to be more high or low virulence. The Corresponding Author: Gamal A. Mohamed, Plant Research Dept., Atomic Energy Authority, Abou Zabal 13759, Egypt 431

2 pathogenecity of pathogens probably changes if the genetic mutation occurs in the genes associated with virulence of fungi after gamma treatment [10,11]. The present work was carried out to study the effect of different doses of gamma radiation on microorganisms and chemical constituents of banana fruits stored under different temperatures. MATERIAL AND METHODS Studied fruits: Mature green banana (Musa sapientum L. cv. Maghrabi) fruits were collected directly from the orchard without any commercial postharvest treatments. Fruits were taken from trees which are in good physical conditions, uniformity in growth and have subjected to the same cultural practices. Radiation Treatments: Mature green banana fruits were irradiated one day after harvest in a Gamma cell contained cobalt 60 installed at Nuclear Research Center, Inshas. The applied irradiation doses were 0, 0.25, 0.50, 1.0, 1.5 and 2.0 K-Gy at dose rate of 5.7 K-Gy/hr. Storage: Irradiated and non-irradiated banana fruits were stored under temperatures of 5, 10, 15, and. Samples were taken at intervals of 7, 14, 21, 28, 35 and 42 days. Parameters and Analysis: Samples of banana fruits taken at the different intervals were subjected for the following tests and determinations: 1- Decay Assessment: The decay of the banana fruits during storage was recorded and based on visual inspection of each fruit for infection and their percent were calculated according to the formula of [12]. In this regard, infected fruits were arranged as following: 0 = superficial fleck (no decay) 1 = 1-24 % of the surface decayed 2 = % of the surface decayed 3 = % of the surface decayed 4 = 75% or more of the surface decayed. The decay index (DI) for each treatment was obtained from the following formula: Sum. (Number of fruits per category DI = Category number) 100 Total number of infected fruits Severity of Infection % = DI/ Microbiological Studies: The official microbiological methods of the American Spice Trade Association [13] were used. After each storage period, banana fruits were tested for the surviving microflora. Briefly, 1 g of each sample was weighed into a sterilized screw cap bottle under aseptic conditions. Sterile phosphate buffer was added to each bottle and serial subsequent dilutions were made. Total Bacterial Count: To obtain the viable bacterial count, 1 ml of the diluted samples was inoculated into duplicate sterile plates of nutrient agar and incubated at 30 o C for 72 hrs and the number of colony-forming units (CFU) was counted. Total Fungal Count: To obtain the total fungal count, 1 ml of the diluted samples was inoculated into duplicate plates of Wort agar and incubated at 30 o C for 72 hrs and the number of mould colonies was counted. Chemical Analysis: Determination of Starch Content (G/100 Gm D.wt.): Starch content was determined according to the method described by [14]. Determination of total sugars content (g/100 gm D.wt.) : Total sugars were extracted from banana pulp using 80% ethyl alcohol. Total sugars were determined in the ethanolic extract using the colorimetric method of [15] Determination of Ascorbic Acid Content (Mg/100 Gm D.wt.): Ascorbic acid was determined in banana pulp using 2, 6 dichlorophenol indophenol method and 2% oxalic acid according to [16]. Determination of Total Chlorophyll Content: Total chlorophyll content of banana peel was extracted using 80% ethyl alc. and determined according to the equations mentioned by [17]. Statistical Analysis: The method [18] which was used for evaluation of variance (ANOVA) by using randomized complete block design (RCBD) and mean values were compared by L.S.D. test at P = RESULTS AND DISCUSSION 1. Decay Development: The effect of gamma radiation on decay percent and shelf-life extension of mature green banana fruits stored under different rates of temperature are presented in Table (1). The promising effect of gamma radiation to reduce the decay percent and in turn extending the shelf-life was greatly correlated with the imposed storage temperature. For fruits stored at, the irradiated and non-irradiated banana showed no decay up to 14 days of storage, however, only the non-irradiated control exhibited 432

3 Table 1: Effect of gamma radiation on disease (decay) assessment of banana fruits stored under different temperature rates. (Values are means of two successive seasons) Mean Mean Mean Mean Mean L.S.D at 5% level Temperature Gama radiation Storage Interaction

4 decay after 21 days of storage. At storage periods of 28, 35 and 42 days irradiated and non-irradiated treatments showed decay with different magnitudes. The lower rate of decay was exerted by irradiation dose of 1.0 K-Gy followed by those of 0.50 and 1.5 K-Gy. Going up to storage temperature of, again no decay could be recorded up to 14 days of storage and only some decay was noticed with the non-treated banana after 21 days of storage. However, all treatments showed different decay percent with the other storage periods up to the end of the experiment. Again, irradiation dose of 1.0 K-Gy was the superior dose to delay the injury and decay percent followed by treatments of 0.50 and 1.5 k-gy. Raising the storage temperature up to 1, only the non-irradiated banana fruits showed some decay after 14 days of storage. On the other side, different magnitude of decay was noticed with all treatments starting from 21 days of storage and up to the end of experiment. Storage temperature of (room temperature), showed earlier decay symptoms and injuries as compared with the lower storage temperature. The deterioration of banana fruits gradually increased by the lapse of storing days and again irradiation dose of 1.0 K-Gy was the optimum treatment to retard the decay and minimizing the deterioration of the stored banana fruits. Results demonstrated in Table (1) clearly reveal that the higher proportional of decay and injury was registered with the highest temperature storage rate and naturally with the latest sampling date. In other words, lowering storage temperature played an important role for minimizing and retarding the deterioration of both irradiated and non-irradiated banana fruits. It is well known that lowering the storage temperature has an impressive effect for delaying ripening and improving fruits firmness and quality due to different physiological and biochemical aspects as discussed by [19,20,21,22]. On the other hand, the possibility of retarding the decay and extending the shelf life of fresh fruits through the judicious application of gamma rays was established and recorded in previous investigations as mentioned by [4,9,23]. 2. Total Bacterial and Fungal Counts: 2.1. Total Bacterial Count : The imposed gamma irradiation showed noticeable effect on total bacterial count of banana fruits and this effect depended greatly on the applied temperature level as demonstrated by data presented in Table (2). In this concern, nonirradiated banana fruits stored at showed initial bacterial count of CFU.g -1 after 7 days of storage and this count was increased gradually until the end of the experiment as it recorded CFU.g - 1. However, the applied gamma irradiation doses showed remarkable effect to decline the total bacterial count of the stored banana fruits and this effect was much more impressive with increasing the applied doses. In this consideration, the applied doses of 0.25 and 0.50 K-Gy reduced the total bacterial load to the lowest level, however, no count could be detected by irradiation doses of 1.0 and 1.5 K-Gy up to the storage period of 21 and 28 days, respectively. On the other side, a complete inhibition was exerted by the higher dose of 2.0 K-Gy during all the storage periods. Increasing the storage temperature to higher level, more initial counts could be recorded and these counts were increased gradually alongside with the studied storage period. For instance, non-irradiated banana fruits showed initial counts of , and CFU.g -1 for storage 10, 15 and, however, these values were increased to , and CFU.g -1 for the same temperature rates, respectively. The action of the applied irradiation doses on bacterial load showed similar trends for those noticed at storage temperature of. In this regard, the total bacterial counts were reduced gradually by increasing the applied irradiation doses and a complete inhibition was exerted by the higher doses of 1.0, 1.5 and 2.0 K-Gy during the earlier storage periods. From the above mentioned results, it could be deduced that the studied irradiation doses have the capacity to decline or completely inhibit the bacterial growth of the stored banana fruits, however, this effect was correlated by the level of the storage period. The inhibition effect of gamma radiation on the microbial growth was explained by [6,9] who stated that gamma radiation cause cell damage direct harm effect on the chromosomal DNA in addition to protein coagulation of the living cell Total Fungal Count: Total fungal count of the stored banana fruits as affected by gamma irradiation doses is demonstrated by data in Table (3). Data clearly reveal that fungal load of the stored banana fruits was less than that observed for the total bacterial load. In addition, it seems that total fungal count was much more sensitive for the applied gamma irradiation doses rather than of bacteria. Also, the action of gamma radiation to decline or inhibit the fungal load was greatly associated with the level of storage temperature. Naturally and as expected, total fungal count exhibited remarkable reduction in the lower storage temperatures rather than the higher ones as observed previously with total bacterial count. For all the applied storage temperature, gamma irradiation doses reduced total fungal count and this effect was much more impressive as the imposed irradiation dose was increased. In this consideration, a noticeable reduction was exerted by the lower doses of 0.25 and 0.50 K-Gy, however, a complete inhibition was 434

5 Table 2: Effect of gamma radiation on total bacterial count (T.B.C.) of banana fruits stored under different temperature rates. (Values are means of two successive seasons) recorded by the higher doses of 1.0, 1.5 and 2.0 K-Gy with respect for banana fruits stored at. Similar trends could be detected for the other applied storage temperature. Generally, total fungal count was increased by raising storage temperature and by time lapse of storage. In other words, higher total fungal count was recorded at storage temperature of and the latest sampling date. Thus, the effect of gamma radiation to decline or induced complete reduction was correlated with the above mentioned two parameters. In this connection, the complete inhibited was shown to be more restricted in the earlier storage periods in the highest storage temperature of. The above mentioned results are supported by the findings of [9] who mentioned a complete inhibition of fungal growth by gamma rays due to DNA destruction. Furthermore, [4,24] reported that the high energy rays of irradiation directly damage the DNA of living organisms, inducing cross-linkages and other changes that make an organism unable to grow or reproduce. Additionally, [24] studied the effect of gamma radiation on the mycelial growth of postharvest fungi of banana fruits and revealed that the growth of C. musa, L. theobromea and Fusarium spp. was greatly deteriorated by 2 K-Gy. 435

6 Table 3: Effect of gamma radiation on total fungal counts (T.F.C.) of banana fruits stored under different temperature rates. (Values are means of two successive seasons) Chemical Analysis: 3.1. Starch Content: Starch content (g./100 g. D.W.) of the studied green-mature banana fruits was varied according to the imposed treatments and period of storage according to the data demonstrated in Table (4). In general, starch showed continuous decline during the different storage periods up to the end of the experiment. However, this decline was remarkably retarded in the lower level of storage temperature as compared with higher one. In addition, the applied gamma irradiation doses exhibited further effect to retard the obtained reduction in starch content during the storage of banana fruits. For banana fruits stored at, irradiation dose of 1.0 K-Gy showed higher amount of starch rather than the non-treated fruits, followed by doses of 1.5 and 2.0 K-Gy, respectively. More or less, similar trends could be observed for the fruits stored at the other temperature levels. Irradiation dose of 1.0 K-Gy was the superior treatment to maintain starch content of the stored banana fruits during the different storage periods. The other studied irradiation showed different maintenance for starch content but it was less than that observed with the optimum dose of 1.0 K-Gy. In a previous study, [22] 436

7 Table 4: Effect of gamma radiation on starch content (g/100 g D.W.) of banana fruits stored under different temperature rates. (Values are means of two successive seasons) Mean Mean Mean Mean Mean L.S.D at 5% level Temperature Gama radiation Storage Interaction

8 Table 5: Effect of gamma radiation on sugars content (g/100 g D.W.) of banana fruits stored under different temperature rates. (Values are means of two successive seasons) Mean Mean Mean Mean Mean L.S.D at 5% level Temperature Gama radiation Storage Interaction N.S. 3.4 N.S

9 Table 6: Effect of gamma radiation on ascorbic acid content (mg/100 g D.W.) of banana fruits stored under different temperature rates. (Values are means of two successive seasons) Mean Mean Mean Mean Mean L.S.D at 5% level Temperature Gama radiation Storage Interaction N.S

10 Table 7: Effect of gamma radiation on peel chlorophyll content (mg/100 g D.W.) of banana fruits stored under different temperature rates. (Values are means of two successive seasons) Mean Mean Mean Mean Mean L.S.D at 5% level Temperature Gama radiation Storage Interaction

11 mentioned that lowering storage temperature has positive effect to keep starch content of banana fruits which should be attributed to reducing the enzymes activity which convert the starch into simple sugars. Also, the promising effect to maintain starch content of the stored banana fruits is due to its effect for reducing the biological activity to a great extent and inturn delaying the natural ripening as stated by [5,9] Total Sugars Content: Total sugars content of non-irradiated and irradiated banana fruits stored under different levels of temperature is presented in Table (5). Total sugars content was found to be increased alongside with the different storage period. That holds true for the fruits stored under the different imposed temperature. However and as might be expected, the rate of increase in total sugars content was interrupted by lowering storage temperature. Thus, banana fruits stored at contained less sugars rather than those stored at 15 or. Regarding the influence of the studied gamma irradiation doses on total sugars content of banana fruits, it could be easily deduced that the imposed treatments retarded the increase in sugars content as compared with the non-treated control during the different storage periods and temperature levels. However, this influence was varied according to the applied irradiation dose. Irradiation dose of 1.0 K-Gy seemed to be the most effective dose to decline the increase in sugars content of the stored banana fruits, and this effect was much more pronounced in the higher temperature of storage and the latest sampling date. The above mentioned results came in agreement with those of [22] who declared that storing banana fruits at low levels of temperature was effective to retard the conversion of starch into simple sugars and inturn keeping sugars content in lower content. Again, the influence of gamma radiation to maintain sugars content in stored banana fruits and retarding its increase is due to delaying the ripening and reducing the biological activity including a reduction in starch conversion into simple sugars as discussed previously [9,23] Ascorbic Acid Content: Ascorbic acid was declined progressively in the stored banana fruits with increasing storage periods as shown from results in Table (6). However, the rate of decrease was much more obvious in banana fruits stored at higher temperature level rather than the lowest one. In other words, storing the banana fruits in low temperature had the capacity to maintain ascorbic acid content and inturn keeping the quality and nutritional value in a good level. On the other side, exposing banana fruits to gamma rays showed also a promising effect to maintain ascorbic acid content and keeping its good quality. Thus, the irradiated banana fruits showed higher ascorbic acid content rather than the nonirradiated ones. Irradiation dose of 1.0 K-Gy was the superior treatment to maintain ascorbic acid content near the normal level. That holds true for banana fruits stored under the different imposed temperature level. With concern of the positive effect of cold storage for maintaining ascorbic acid, [22] mentioned that lowering storage temperature should reduce rate of metabolic and biological processes that lead to a vit. C degradation. On the other hand, several investigators mentioned the promising effect for maintaining ascorbic acid and keeping the good quality of the stored fruits to a high level [21,25,23] Peel Chlorophyll Content: Chlorophyll content of banana peel as affected by the applied irradiation treatments and stored under different temperature levels is shown in Table (7). In general, total chlorophyll content was decreased as storage periods increased. There was a linear relationship between the chlorophyll decline and the progress of storage period. However, the rate of chlorophyll degradation was faster in the higher storage temperature rather than the lower ones. So, chlorophyll content of banana peel exhibited higher proportions in the cold storage. Regarding the effect of gamma radiation, it could be easily deduced that the irradiated banana peels contained more chlorophyll rather than the non-irradiated control. In other words, the imposed gamma irradiation treatments had the capacity to retard the decline in chlorophyll content and that holds true for all the storage periods and temperatures. Irradiation dose of 1.0 K-Gy was found to be the most effective one to maintain chlorophyll content in banana peel. The other doses showed different maintenance for chlorophyll peel but still less than 1 K-Gy treatment. 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