Biological value and protein quality of raw and processed seeds of Mucuna pruriens var. utilis

V Vadivel and M Pugalenthi

Department of Biotechnology, Karpagam Arts and Science College,
Coimbatore - 641 021, Tamil Nadu, India
pugalmuthiah@rediffmail.com

Abstract

The under-utilized legume seeds have better potential to meet the increasing protein requirements of livestock industries, but remain untapped. In the present study, the proximate composition, antinutritional compounds and biological value of seeds of a potential under-utilized legume, Mucuna pruriens (L.) DC. var. utilis (Wall. ex Wight) Baker ex Burck (velvet bean) were investigated.

The mature seeds were found to contain high level of protein (27.3%); lipid (6.06%); fiber (9.7%); ash (5.6%) and carbohydrates (51%). Even though, the raw seeds were found to contain various nutritionally undesirable antinutritional substances, the autoclaving treatment effectively reduced their maximum levels without affecting the nutritional profiles of the velvet bean seeds when compared to soaking, cooking or roasting processing methods. The rats fed with autoclaved seed included diet for 28 days exhibits better growth performance such as higher feed intake (FI) (192 g) and body weight gain (BWG) (51 g). Moreover, the protein quality parameters such as protein efficiency ratio (PER), true digestibility (TD), biological value (BV), net protein utilization (NPU) and utilizable proteins of velvet bean seed proteins were significantly improved by autoclaving processing method when compared to other processing techniques.

Key words: Anti-nutritional compounds, biological value, Mucuna pruriens var. utilis, processing methods, protein quality, velvet beans

Introduction

Most of the developing tropical countries depend on soy bean and other common legume grains as key protein source for feeding livestock animals. But, their low production can not keep phase with the increasing requirements of these conventional protein sources for the expanding livestock industries (Udedibie and Carlini 1998). The heavy demand for these common legumes has given rise to a disproportionate increase in their prices, and consequently, in the cost of the livestock feeds. Hence, the recent research trend has directed to identify and evaluate the under-utilized legume seeds as alternative/additional protein source for livestock animals (Janardhanan et al 2003). Among the various under-utilized pulses, the velvet bean [Mucuna pruriens var. utilis (Wall ex Wight) Baker ex Burck] seeds merits a wide use in South Asian countries and other parts of the tropics as food legume (Pugalenthi et al 2005).

The velvet bean seeds have been reported to contain high level of protein (26-30%) and starch (34-40%), desirable amino acid, fatty acid and mineral composition with good nutritional properties. In India, it has been traditionally used as food by certain ethnic groups, particularly, the Northeastern tribes and Kanikkar tribes in Kerala State and Dravidian tribes in Tamil Nadu State (Pugalenthi et al2005; Vadivel and Pugalenthi 2006). It is easy to cultivate under dry farming and low soil fertility conditions and exhibits many favourable agronomic characters with reliable yield (2.9-6.9 t/ha) (Pugalenthi and Vadivel 2006).

The exploitation and development of such potential non-conventional/under-utilized legume seeds as protein source in animal feeds may offer a good scope to meet the increasing protein requirements in the livestock sector at large, particularly, in the developing countries. However, before recommending such indigenous foodstuffs, their nutritional properties and biological value should be thoroughly investigated. Reports are available on the nutritional quality and biological value of certain under-utilized legumes like Bauhinia purpurea (Vijayakumari et al 1997a); Canavalia ensiformis, Canavalia gladiata, Canavalia maritima, and Canavalia cathartica (Bressani et al 1987; Seena et al 2005; Bhagya et al 2006). But, in the case of velvet beans, although reports are available on the biochemical composition and nutritional value, information regarding the biological value of seed proteins appears to be meager. Hence, the present study was carried out to analyze the biological value and protein quality of raw and differentially processed seed proteins of velvet beans collected from South India.
 

Materials and methods

Collection of the seed materials

The seed samples of velvet beans were collected from Kailasanadu, Idukki District, Kerala, South India. Soon after collection, the immature and damaged seeds were removed and the mature seeds were dried in the sun light for 24 h and stored in plastic containers in refrigerator (5^oC), until further use.

Processing methods

Five separate batches of whole seeds of velvet beans were taken and the first batch was soaked in distilled water for 6 h at room temperature (30 ± 2^oC) in the bean to water ratio of 1:10 (w/v). The second batch of seeds was cooked at 90-95^oC for 1 h in the bean to water ratio of 1:10 (w/v). The third batch of seeds was taken in the bean to water ratio of 1:10 (w/v) in a metal container and autoclaved at 15 lb pressure (121^oC) for 30 min. The fourth batch seeds were roasted for 30 min at 100-110^oC in an iron pot along with clean fine sand to prevent the burning of the seed coat and to ensure the uniform distribution of heat. After each treatment, the treated seeds were rinsed with distilled water, separately, and then dried at 55^oC for 6 h in a hot air oven. The fifth batch of raw seeds was stored as such with out any treatment.

Analytical methods

All the processed as well as raw seeds of separate batches were powdered in a Willey Mill to 60-mesh size and the powdered samples were used for further analysis. The proximate composition such as moisture, crude protein, crude lipid, crude fiber and ash content of raw as well as processed seed flour was determined by following AOAC (1990) method. Nitrogen free extractives (NFE) and calorific values were calculated by following the method of Siddhuraju et al (1992).

The antinutritional compounds such as total free phenolics and tannins content of raw and processed seed samples were extracted and estimated by following the method of Sadasivam and Manickam (1992) and Burns (1971), respectively. The L-Dopa (L-3,4-Dihydroxyphenylalanine) content was quantified according to the method of Brain (1976), whereas, the phytic acid content was determined by following the method of Wheeler and Ferrel (1971) and the oligosaccharides content by Pugalenthi et al (2006) method. The haemagglutinating activity was analyzed according to the method of Makker et al (1997). The trypsin inhibitor activity was determined by casein digestion method (Mulimani and Vadiraj 1993) and α- amylase inhibitor activity was measured according to the Mulimani and Rudrappa (1994) method.

Biological evaluation of protein quality

Fifty numbers of 23 days old male albino rats with an initial body weight of 40 ± 5 g were equally divided into five groups with 10 animals in each group and housed individually in cages. The animals were maintained at 22^oC with 12 h light and 12 h dark at Karpagam Animal House (Approved by Animal Ethical Committee, Government of India). The experimental diets were prepared according to the method of Chapman et al (1959) by including corn starch (80%), corn oil (10%), non-nutritive cellulose (5%), mineral mixture (4%) and vitamin mixture (1%). The raw and differentially processed velvet bean seed flour was incorporated in the diet at the expense of corn starch to the diet containing 10% protein. A diet containing casein as a protein source was used as control diet. The control and test diets along with water were fed to respective animal groups ad libitum for 28 days.

Weighed diet was given daily and unconsumed diet was collected and weighed to calculate the feed intake (FI) values. Weight of the rats was recorded twice in a week and the body weight gain (BWG) was calculated at the end of the experiment. The protein content of the diet was determined by micro-kjeldahl method (AOAC 1990) and the feed efficiency ratio (FER) and protein efficiency ratio (PER) were calculated according to the method of Chapman et al (1959).

The nitrogen balance studies were conducted for 14 days with 60 numbers of male albino rats of 50 ± 6 g body weight. The rats were randomly separated into six groups with 10 animals in each group and individually housed in polypropylene metabolic cages. The animal groups were fed with control casein diet, raw and different processed velvet bean included test diets, separately, and one batch was fed with protein free diet for the determination of endogenous and metabolic nitrogen loss in faeces and urine. After 9 days of acclimatization period, observations were made for nitrogen intake, nitrogen excreted in urine and faeces of experimental animals for 5 days. The values of true digestibility (TD) and biological value (BV) of seed proteins were determined by following the method of Chick et al (1935) and net protein utilization (NPU) by Platt et al (1961), whereas, the level of utilizable proteins was calculated by Gupta et al (1979) method.

Statistical analysis

Results were expressed as mean values ± standard deviations of three separate determinations. The data was subjected to a one-way analysis of variance (ANOVA) and the significance of difference between means at 5 % was determined by Duncan's Multiple Range Test (DMRT) using Irristat software (version 3/93).
 

Results and discussion

Chemical composition

The proximate composition of raw and processed seeds of velvet bean was shown in Table 1.

The crude protein and lipid content of raw seeds (27.3% & 6%) were found to be higher when compared to certain common legumes such as Cicer arietinum (20.7% & 4.16%); Vigna mungo (23.6% & 0.45%); Vigna radiata (24.5% & 0.71%); Vigna aconitifolia (25.3% & 0.69%) and Phaseolus vulgaris (25.1% & 0.9%) (Bravo et al 1999). The protein and lipid content of autoclaved seed samples (25.5% & 6.23%) were higher than the raw and other processed seeds, which is in consonance with the earlier report in the same pulse (Siddhuraju and Becker 2005). The substantial reduction of the ash content in the soaked seeds (45%) when compared to raw seed samples might be due to the leaching of both micro and macro elements into the soaking medium through the enhanced permeability of the seed coats during soaking treatment.

Antinutritional compounds

The effect of various processing methods such as soaking, cooking, autoclaving and roasting on the levels of antinutritional compounds of velvet beans were given in Table 2.

Among the various processing methods employed, the autoclaving was found to significantly (P<0.05) reduce the maximum levels of various antinutritional substances such as total free phenolics (81%), tannins (76%), L-Dopa (79%), phytic acid (77%), oligosaccharides like raffinose (77%), stachyose (73%) and verbascose (75%), haemagglutinating activity (69%), trypsin inhibitor activity (72%) and amylase inhibitor activity (78%). Similarly, significant reduction of various antinutritional compounds during autoclaving treatment was reported for several under-utilized legumes such as Dolichos lablab (Vijayakumari et al 1995); Mucuna monosperma (Vijayakumari et al 1996); Prosopis chilensis (Vijayakumari et al 1997b); Vigna aconitifolia and Vigna sinensis (Vijayakumari et al 1998) and Bauhinia purpurea (Vijayakumari et al 2007).

Growth performance of the animals

The growth performance of the experimental animals fed with diets containing raw and processed velvet bean seeds was illustrated in Table 3.

The FI value (140 g) was significantly lower (p<0.05) for raw velvet bean seeds as compared to casein (276 g) and treated velvet bean seeds (155-192 g). The raw velvet beans show lower FI value than Canavalia ensiformis (225 g) and Canavalia gladiata (244 g) (Bressani et al 1987), but higher when compared to the FI values of vegetable peas (105-120 g) (Saharan and Khetarpaul 1994) and Canavalia maritima (93 g) (Seena et al 2005). The significantly lower FI in rats fed with raw velvet beans than control and processed seeds were probably due to the difference between the diets in protein quality and effects of antinutritional compounds in the raw seeds. Processed velvet beans, particularly the autoclaved seeds containing diet was consumed in larger amount (192 g) than the raw seed included diet. The reduction in the levels of various antinutritional substances under autoclaving treatment might be related to larger FI values for autoclaved velvet bean seeds inclusive diet.

The animal group fed with raw velvet bean included diet shows lowest BWG value (26.5 g) when compared to the control (76.2 g) and test animals fed on treated seeds (32-51 g). The BWG value for raw seeds of the present study is in agreement with an earlier report on vegetable peas (23-28 g) (Saharan and Khetarpaul 1994). Among the different treated seeds, the autoclaved seeds significantly improve the BWG of rats (40 g), which is higher than the BWG values for pressure-cooked seeds of Canavalia ensiformis (28 g) and Canavalia gladiata (32 g) (Bressani et al 1987). Soaking and cooking processing methods were not demonstrated to have a beneficial effect on the growth rate of the animals. It might be due to the presence of heat resistant antinutritional compounds in the velvet beans, which are not destroyed completely by these treatments.

The raw velvet bean seeds exhibit poor FER (0.19) and PER (1.87) values when compared to control and processed seeds (Table 3), which might be due to the presence of high concentration of antinutritional substances and poor quality of proteins of the raw seeds. However, the FER and PER values of raw velvet bean seeds are higher than the faba beans (0.032 & 0.32) (Gupta et al 2005), but lower than vegetable peas (0.22 & 2.17) (Saharan and Khetarpaul 1994). Among the different treatments, the autoclaving results in significant (P<0.05) improvement of FER (0.26) and PER (2.67) of velvet bean seeds. The PER value of autoclaved velvet bean seed is higher when compared to Canavalia ensiformis (1.24) and Canavalia gladiata (1.24) (Bressani et al 1987). FI values were varied among the treatments and so did PER values also to vary. This result showed that the higher the FI, the higher the PER values obtained. This finding is coincided with those of Bender (1956), who has pointed out that the PER determination is depends upon feed consumption.

The low PER value of cooked (2.3), autoclaved (2.67) and roasted (2.33) seeds compared to casein (2.92) could be due to the fact that heat accelerates the millard reaction and makes the protein unavailable (Sagarbieri 1989). Moreover, usually much of the sulphur containing amino acids such as cystine and methionine supplied in the diet were used to synthesize pancreatic enzymes (Fernandez et al 1996). This exacerbated the deficiency of sulphur containing amino acids in legume seeds, were manifested as a lower production of body tissues. Rerouting of retained nitrogen in the animal group fed raw velvet bean seeds may explain why PER in this group was significantly lower than in control, despite the fact that protein content was similar in these two groups.

Protein quality

The protein quality such as true digestibility (TD), biological value (BV), net protein utilization (NPU) and utilizable proteins of raw and processed velvet bean seeds were presented in Table 4.

The TD value of raw velvet bean seeds was found to be lower (67.7%) when compared to control (92.7%) and treated seeds (69.4-78.5 %) and the autoclaved seeds exhibit the highest TD value. The TD level (69.7 %) of raw velvet bean seed is found to be higher when compared to an earlier report on Bauhinia purpurea (46.4%) (Vijayakumari et al 1997a); Canavalia maritima (42.2 %) (Seena et al 2005) and faba beans (63.4%) (Gupta et al 2005) and comparable with vegetable peas (65.8-66.7%) (Saharan and Khetarpaul 1994). The autoclaving treatment significantly improves the TD level of velvet beans (78.5%), which is higher than the values reported for Canavalia ensiformis (76.4%) (Bressani et al 1987) and faba bean (71.4%) (Gupta et al 2005).

The consumption of raw legume seed proteins was reported to increases the endogenous nitrogen loss through the shedding of intestinal mucosa (Sanoja and Bender 1983; Fairweather-Tait et al 1983), an effect that reduces the biological value of raw legume seed proteins. Further, the presence of various antinutritional substances, including trypsin inhibitors, which impede the complete digestion of protein and increases the endogenous faecal excretion of nitrogen (Nestares et al 1996) was also partly responsible for the decrease in protein digestibility value of raw velvet bean seeds.

When compared to the differentially treated seeds, lowest BV value was recorded for raw velvet bean seeds (61.5%), which is comparable with that of BV level of vegetable peas (62.9-63.1%) (Saharan and Khetarpaul 1994) and faba bean (60.4%) (Gupta et al 2005), but found to be higher than Bauhinia purpurea (57.2%) (Vijayakumari et al 1997a). The NPU value of raw velvet bean seeds is also lower (40%) when compared to casein (70.4%) and treated seeds (43.5-54.7%). The NPU level of raw velvet bean seeds is similar with that of earlier reports on vegetable peas (41-42%) (Saharan and Khetarpaul 1994) and faba bean (38.3%) (Gupta et al 2005) but higher than Canavalia maritima (16.8%) (Seena et al 2005). The autoclaved velvet beans exhibit highest level of NPU (54.7%) among the raw and different treated seeds, which is also higher than the previous report on Bauhinia purpurea (46.4%) (Vijayakumari et al 1997a). The highest level of utilizable proteins (16.4%) was registered by autoclaved velvet bean seeds when compared to raw and other processed seeds (8.5-9.2%). The utilizable proteins value of raw velvet bean seeds (8.5%) is comparable with that of vegetable peas (8.4-8.6%) (Saharan and Khetarpaul 1994), but higher than that of Bauhinia purpurea (7.2%) (Vijayakumari et al 1997a)

Among the various antinutritional compounds, the protease inhibitors and tannins were reported to largely decrease the protein quality of seed proteins (Liener 1994). These substances inhibit the proteolytic action of protease enzymes, which leads to the incomplete digestion of proteins with lower amounts of amino acids becoming available for growth (Fernandez et al 1996). The decrease in the trypsin inhibitor activity and other antinutritional constituents as a consequence of autoclaving treatment would have been reducing the faecal nitrogen excretion in rats fed with autoclaved seeds included diet. Improvement in the protein quality after autoclaving treatment might be attributed to reduction on the levels of various antinutrients and to some other factors such as disruption of protein structure and increased accessibility of the seed proteins to enzymatic attack (Nielson 1991).The fact that no such decrease in faecal nitrogen was occurred in the case of roasted seeds may have been due to the protein denaturation, because dry heat treatment cause isopeptide formation (Dutson and Orcutt 1984; Kirk 1984). This further reduces the protein quality, as isopeptides are not hydrolyzed in the intestine, are resistant to proteolytic enzymes and are thus excreted in faeces. As a result, digestibility and availability of some amino acids are reduced (Kirk 1984) and thus gave to poor values of BV, NPU and utilizable proteins for roasted seeds.

Rats that received dietary casein as protein source were able to take full advantage of the nitrogen they retained to favour growth, probably as a result of the more balanced supply of amino acid provided by this diet. In rats fed with raw velvet bean seeds, part of the nitrogen retained may have been rerouted for the synthesis of digestive enzymes such as trypsin and chymotrypsin (Liener 1994) to offset the effects of the high level of protease inhibitors present in the raw velvet bean seeds. The protein quality parameters such as TD, BV, NPU and utilizable proteins of the autoclaved velvet bean seeds are higher than the raw and other processed seeds. This is in agreement with the previous study in cowpea (Dario and Salgado 1994) and Bauhinia purpurea (Vijayakumari et al 1997a). Geervani and Theophilus (1980) also observed that wet heat methods of processing improve the protein quality of Cicer arietinum and Vigna radiata to a greater extent than dry heat methods.
 

Conclusion

The results of the present study indicated that the autoclaving treatment is suitable and more effective in reducing various antinutritional compounds with out affecting the nutritional quality of velvet bean seeds. When considering the biological value, the autoclaved velvet bean seed proteins exhibit better animal growth performance and protein quality when compared to other processing methods of the present study. Hence, such economic and potential processing method could be adapted for the versatile utilization of velvet bean seeds as a protein source. Incorporation of such autoclaved velvet beans in the diets of animals will clearly reduce the over-dependence on common legumes for increasing protein requirements in livestock industries, especially in the developing countries. Investigation on utilization of autoclaved velvet bean seeds as a protein ingredient in the diets of poultry birds is in progress.
 

Acknowledgements

Authors are grateful to the University Grants Commission for giving financial support to a Major Research Project [Sanction No. F. 3 - 42/2004 (SR) dt. 12.01.2004] and thankful to the Management and Administrative authorities of Karpagam Arts and Science College for their encouragement and support.

 

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