Abstract
Effects of electron beam irradiation at doses of 10, 15, 20, 25 and 30 kGy on chemical composition, phytate and tannin contents, and in vivo digestibility of sorghum grain were investigated. Irradiation had no effect (p >0.05) on chemical compositions. Tannin content was reduced (p <0.05) by 28, 30, 42, 83 and 86 percent and phytate content reduced (p <0.05) by 39, 49, 66, 79 and 90 percent in electron beam irradiated compared to control, respectively. Irradiation improved (p <0.05) in vivo digestibilities of dry matter, crude protein, true protein and gross energy. Based upon these results, electron beam irradiation judge to be useful processing method for reducing the anti-nutritional compounds and therefore, improving nutritive value of sorghum grain.
Keywords: Electron beam; Phytate; Tannin; Sorghum; Digestibility
1. Introduction
Grain sorghum outperforms other cereals under various environmental stresses and is thus generally more economical to produce. This grain is cultivated as a staple food for human consumption, animal feed and industrial products such as alcohol. Sorghum components, especially its protein and energy, are less digestible than other cereals for human and monogastric animals (Kondos and Foale, 1983; Chen et al., 1995; Fombang et al., 2005), because of anti-nutritional factors such as tannins and phytic acid. Removal of these undesirable components is essential to improve the nutritional quality of sorghum and effectively utilize its potential as human food or animal feed. The methods (Chen et al., 1995; Duodu et al., 1999; Parker et al., 1999) used to deactivate the anti-nutrients need not necessarily reduce or completely eliminate these compounds; instead some methods reduce the nutritive value of sorghum grain.
Gamma and electron beam irradiation has also been shown to reduce or inactivate some of the anti-nutritional factors in wild leguminous seeds or meals, thereby enhancing their edibility (Siddhuraju et al., 2002; Bhat, et al., 2007). The effectiveness of gamma irradiation on quality of sorghum has been investigated (Fombang et al., 2005). Electron beam has several distinct advantages compared to gamma irradiation, such as short treatment time, no residuals in products, no need to porous packaging, controlled with much precision, eliminated need for rinse water and etc. In the literature, little information is available concerning effects of electron beam irradiation on nutritive value of sorghum. Therefore, in this study, it was aimed to evaluate effects of electron beam irradiation on chemical composition, anti-nutritional contents and in vivo digestibility of sorghum grain.
2. Materials and methods
2.1. Sample irradiation
Seed samples were packed in 30 cm - 40 cm - 5 cm nylon bags (0.5 mm thickness) and exposed to electron beam irradiation at the Yazd radiation processing center (AEOI, Yazd center, Iran) to various doses (10, 15, 20, 25 and 30 kGy) at room temperature by a Rhodotron accelerator model TT200 (IBA Co., Belgium). All samples were irradiated at fixed beam energy of 10 MeV and the required irradiation doses were obtained by adjusting the electron beam parameters (electron beam current, Conveyor speed and etc.). Double side irradiation (exposure to both sides) was performed for uniform dose delivery. The dose was determined with cellulose triacetate films. Similarly packed seed samples without irradiation served as control.
2.2. In vivo digestibility
Cockerels were used in this study as a model for determining dry matter, crude protein, true protein and gross energy digestibilities of untreated and irradiated samples. Ten randomly selected Rhode Island Red cockerels (average age, 40 weeks; average weight, 2.9 kg) were placed individually in metabolic cages with fixed aluminum trays for separating excreta collection. In a completely randomized design, five cockerels as replicates were given untreated or irradiated samples at different doses periodically. Experiment was carried out with 3 days adaptation period, 2 days starvation for depleting digestive tract, then 1 day feeding and following 2 days starvation for complete excretion of undigested material. The samples of dropping avoided during final 72 h period were collected, weighted and frozen (-18°C). Analyses of dry matter, crude protein, true protein and gross energy of untreated and irradiated samples were conducted and calculations were carried out.
2.3. Total protein, fat, moisture, ash and fiber contents
Moisture content was determined from the mass of samples before and after they were stored overnight in an oven at 105°C (Methods 925.09; AOAC, 1995). Nitrogen was determined by using a Dosimat-776 Metrohm apparatus (Metrohm Co., Switzerland) according to AOAC (Method 984.13; AOAC, 1995). The instrument was calibrated each time with ammonium sulfate as a nitrogen standard. Fat content was determined with a Solvent Extractor (Behr Labor-Technik, Düsseldorf, Germany) equipped with six Soxhlet posts. The ether extract was determined according to the method 920.39 (AOAC, 1995). Ash was determined by burning duplicate 2 g samples at 540°C, for 3 h in a muffle furnace (Method 942.05; AOAC, 1995). Crude fiber was determined by treating an oil-free sample by sulphuric acid (0.26 N) and potassium hydroxide (0.23 N) solution using an automatic fiber analyzer (Velp Scientifica, Milan, Italy), followed by oven drying and muffle furnace incineration AOAC (1995). Gross energy of grain and excreta samples were determined by adiabatic bomb calorimeter using Parr-4 Model 1241 Calorimeter. True protein quantification was performed by the method of Bradford (1976). The protein contents of the samples were calculated using a calibration curve obtained for bovine serum albumin standards (0-1.5 mg) treated in the same way. Two extractions were carried out per sub-sample and each sample was analyzed in duplicate.
2.4. Determination of tannin
Quantitative estimation of tannin, as catechin equivalent, was carried out using the modified vanillin-HCl method of Price et al. (1978). Seed flour (1 g) was extracted with methanol (10 ml, 26°C, 12 h), vortexed and decanted. This process was repeated and the supernatant was pooled and made up to 25 ml. The extract (1 ml) was treated with reagent mixture (5 ml of 4% vanillin in methanol and 8% concentrated HCl in methanol, 1:1) and after 20 min, the absorbance was read at 500 nm using CE 2021 spectrophotometer (Cecil instruments, Cambridge, England) using catechin (0-250 µg) as standard.
2.5. Phytate content
Phytate of raw and irradiated samples was determined according to the method described by De Boland (1975). A standard curve was prepared to calculate the ferric ion concentration. The phytate phosphorous was calculated from the ferric ion concentration assuming 4:6 iron to phosphorous molar ratio.
2.6. Statistical analysis
Experimental data were submitted to a Duncan analysis to determine whether the different treatments yielded significantly different results. The Duncan procedure was applied as described by Steel and Torrie (1980), and a difference of p less than 0.05 was considered to be significant.
3. Results and discussion
3.1. Effects on chemical composition
The results of proximate composition of untreated and irradiated sorghum grains are shown in Table 1. There were no significant differences in ether extract, protein and fiber, ash and cell wall contents between the irradiated and non-irradiated grains. These results are in agreement with previous work (Bhat and Sridhar, 2008) on the effect of electron beam irradiation on proximate composition of lotus seed and others (Ebrahimi et al., 2009; Taghinejad et al., 2009) on the effect of gamma irradiation on chemical composition of rapeseed and soybean seed. In former study, irradiation up to 30 kGy and in two later, up to 45 kGy had no significant effect on the proximate composition of seeds.
[Table 1 about here]
3.2. Effects on tannin and phytate contents
Tannin and phytate contents of electron irradiated sorghum are shown in Figure 1. Decrease in tannins was significant at the doses of 10, 15, 20, 25 and 30 kGy compared to control by 28, 30, 42, 83 and 86 percent, and in phytate content by 39, 49, 66, 79 and 90 percent, respectively.
[Figure 1 about here]
The main aim of this study was to evaluate electron beam effects on tannin and phytate contents of sorghum grain. Effect of gamma irradiation on tannin contents of sorghum has been assessed, but no references were found in the literature dealing with the effects of electron beam irradiation on tannin contents of sorghum or other sources. Some reports (Siddhuraju et al., 2002; Bhat et al., 2007) indicated that gamma irradiation increased tannins and others (Abu-Tarboush, 1998; Villavicencio et al., 2000) reported that it decreased tannins in foods or feeds. Mechanism of gamma action on tannin has been related to generation of the hydroxyl and superoxide anion radicals (Riley, 1994), but mode of electron beam action on tannins has not been demonstrated. Further study is needed to clear its mode of reducing in tannin contents.
Phytate content was significantly reduced by electron irradiation, especially eliminated at 25 kGy onwards. Similar observations have been made earlier by Duodu et al. (1999) and Bhat et al. (2007), wherein phytate content of foods or feeds were reduced after exposure to ionizing radiation. Mode of phytate loss by γ-irradiation has been explained in the literature. In this concern, chemical degradation of phytate to lower inositol phosphates and inositol by the action of free radicals (De Boland et al., 1975), and cleavage of the phytate ring (Siddhuraju et al., 2002; Duodu et al., 1999) are reasons of reducing in phytate contents of foods or feeds. Mechanism of phytate reduction by electron beam irradiation has not been demonstrated in the literature.
3.3. Effects on digestibility
The results of in vivo digestibility of untreated and irradiated sorghum grains are shown in Figure 2. At doses higher than 15 kGy, digestibilities of dry matter, crude protein, true protein and gross energy increased significantly compared to control. Results revealed that presence of anti-nutritional factors such as tannins and phytate is the main reason of lowing digestibility in untreated sorghum. Similarly Kondos and Foale (1983) showed that these anti-nutritional factors affect negatively the efficient use of sorghum based diets in pigs.
[Figure 2 about here]
Duodu et al. (2003) reported that tannins have a detrimental effect on the ileal digestibility of protein and amino acids. Due to their hydroxyl groups, tannins may interact with and form complexes with proteins, which may lead to precipitation because of the large size of the tannins. In addition to possibly causing a change in protein conformation, study of Siddhuraju et al. (2002) showed that the tannins may also exert steric effects (due to their large size) and prevent enzymes access to the proteins. Another possible reason for increasing in protein digestibility is modification in the three dimensional structure of sorghum proteins due to irradiation. Studies of Shawrang et al. (2007, 2008) illustrated that protein denaturation occur by irradiation that lead to improvement in intestinal protein digestion.
As stated by Ryden and Selvendran (1993), the phytate molecule is highly charged with six phosphate groups and so is an excellent chelator, forming insoluble complexes with mineral cations, starch and proteins. This leads to reduced protein and starch (energy source) digestibility. The possible formation of a complex between phytate and sorghum proteins, which could lead to reduced protein digestibility, has not been studied in sorghum, but results of Elsheikh et al., (2000) in various legumes and Elkhalil et al., (2001) in sorghum showed that processing treatments could reduce phytic acid content and enhance protein digestibility.
4. Conclusion
The present study reveals that electron beam irradiation has the potential to reduce the adverse effects of antinutritional factors on protein and energy digestibility of sorghum grain. This occurs through decreasing in tannins and phytate contents that could interact with proteins and energy components. Effect of electron beam irradiation on phytate elimination was higher than tannins and it was dose dependent. Electron beam irradiation as a physical method of preservation proved its efficacy in maintaining the nutritive value of sorghum grain.
Acknowledgments
Authors are grateful to Yazd radiation processing center, Nuclear Science and Technology School, Atomic Energy Organization of Iran for the irradiation operations. This study was financially supported by Deputy of Research and Technology, Atomic Energy Organization of Iran (project no: A87A050; 21.10.2008).
