, Volume 44, Issue 5, pp 609–614 | Cite as

Antinutritional factors in ricebean, Vigna umbellata Thunb. (Ohwi and Ohashi) against Callosobruchus maculatus F. (Coleoptera: Bruchidae)



To identify some effective and eco-friendly inhibitors of a serious pest of pulse crops, Callosobruchus maculatus, seeds of 10 genotypes of rice bean, Vigna umbellata and mungbean variety, PAU 911 as susceptible check were screened. The different genotypes of rice bean were found to arrest the growth and development of C. maculatus at grub stages in varying levels. To confirm these results, the seeds of different test genotypes were analysed for the presence of three antinutritional factors (tannins, phytic acid and saponins). The amount of tannins, phytic acid and saponins was recorded highest in rice bean genotype LRB 507 (11.12 mg g−1), LRB 482 (32.30 mg g−1) and LRB 535 (4.17 mg g−1), respectively whereas lowest in mungbean variety, PAU 911 (8.24, 13.51 and 1.12 mg g−1, respectively). These antinutritional factors showed a negative correlation with adult emergence, growth index and adult longevity of C. maculatus but positive correlation with development period. Based on the results of the present study, all the rice bean genotypes were identified as resistant to C. maculatus as compared to susceptible check variety, PAU 911. The role of antinutritional factors in relation to C. maculatus infestation is discussed.


Antinutritional factors Callosobruchus maculatus Genotypes Resistant Vigna umbellata 


Grain legumes popularly known as pulses, play an important role in Indian agriculture. These are referred to as poor man’s meat and rich man’s vegetable (Singh and Singh 1992). Pulse beetles assume greater importance as they damage pulses both in the field and storage (Anonymous 1970). The cowpea weevil, Callosobruchus maculatus F. is a serious pest which infests the pulse seeds at storage. It is widely distributed among the bruchid species, occurring in Africa, Asia, and Australia (Daglish et al. 1993). The estimated losses due to bruchids in various pulses ranged from 30 to 40% within a period of 6 months and the post-harvest seed losses can reach even 100% during severe periods of infestation (Mahendran and Mohan 2002).

Rice bean, Vigna umbellata Thunb. (Ohwi and Ohashi) is a crop of tropical and subtropical regions. The centre of origin and diversity of rice bean is considered to be Indo-China (Tomooka et al. 2000). In India, the distribution of the crop is confined to North-Eastern hills, Western and Eastern Ghats and parts of Himachal Pradesh (Chandel et al. 1978). The dry seeds of rice bean are good source of carbohydrates, proteins, minerals and vitamins. Protein in rice bean is rich in limiting amino acids including methionine and tryptophan (Carvalho and Vieria 1996) and other amino acids including valine, tyrosine and lysine (Mohan and Janardhan 1994). The vegetative parts of the crop serve as nutritive forage for animals. Thus, the suitability of rice bean both for food and fodder makes it a dual purpose crop (Chandel et al. 1988).

Although insecticides are effective against these insects but cause serious problems including residual effects and are carried through the food chain leading to disastrous consequences. This provoked a serious thinking for the development of non-chemical pest management strategies. In this context, host-plant resistance can be a viable alternative to chemical control methods (Khush and Brar 1991). Certain wild species of Vigna are known to have resistance against C. maculatus under storage conditions (Marconi et al. 1997). Role of protease inhibitors in rice bean for providing resistance against bruchids is well known. Ignacimuthu et al. (2000) have also stressed the importance of protease inhibitors including trypsin and chymotrypsin inhibitors in conferring resistance to pulse seeds against C. maculatus. Thus, the present investigation was undertaken to study some other antinutritional factors viz. tannins, phytic acid and saponins responsible for resistance to C. maculatus in different genotypes of V. umbellata under storage conditions.

Material and methods

Procurement of seed

The seeds of 10 genotypes of rice bean (RBL 1, RBL 6, RBL 35, RBL 50, LRB 470, LRB 482, LRB 507, LRB 522, LRB 529, LRB 535) and mungbean variety, PAU 911 (suceptible check) were procured from Department of Plant Breeding and Genetics, Punjab Agricultural University, Ludhiana, India.

Bruchid development on different test genotypes

The experiment was planned to study C. maculatus development on 30 seeds of each test genotype which were taken in glass vials separately for egg laying. Three replications were maintained for each test genotype. One pair of freshly emerged adults was released in each vial. After 24 h of release, the insects were taken out of the vials and only one egg per seed was kept by removing extra eggs. The number of adults emerging from seeds were counted daily and summed up to calculate per cent adult emergence. Total development period was considered from egg laying up to the adult emergence. Growth index was calculated by using the formula given by Sachdeva et al. (1986).
$$ \mathrm{Growth}\ \mathrm{index}=\mathrm{Adult}\ \mathrm{emergence}\ \left(\%\right)/\mathrm{Average}\ \mathrm{development}\ \mathrm{period} $$

The period from the time of emergence of an adult till its death was taken as adult longevity.

Biochemical analysis

Inhibitory activity of three antinutritional factors (Tannins, phytic acid and saponins) was measured by following standard procedures of Sadasivam and Manickam 1992; Zamel and Shelef 1982; Fenwick and Oakenfull 1983, respectively. They were expressed in terms of mg/g of seed materials.

Statistical analysis

The data were subjected to analysis with Completely Randomized Design (CRD) by using suitable transformations and the statistical software CPCS1 (Cheema and Singh 1990). Standard error was also worked out. The biology of C. maculatus and antinutritional factors present in rice bean were tested at 5% level of probability (p < 0.05) using the least significant difference (LSD) test.


Biology of C. maculatus

The experiments on biology of C. maculatus were conducted twice and the pooled mean of both experiments are revealed in Table 1. The data on adult emergence, development period, growth index and adult longevity showed significant effects. Adult emergence in different rice bean genotypes was observed approximately 13–45 times lower than mungbean variety, PAU 911 (99.44%). The delay in development period of C. maculatus was much pronounced in LRB 535 (30.25 days) as against PAU 911 (23.19 days). The genotypes which exhibited lesser adult emergence encountered a prolonged development period. The growth index in different genotypes of rice bean varied from 0.07 in LRB 535 to 0.30 in RBL 6 which was found to be very low as compared to mungbean variety, PAU 911 (4.28). Significantly highest male longevity was recorded in mungbean variety PAU 911 (7.11 days), whereas lowest in genotype LRB 535 (4.00 days). Same pattern was observed in case of female longevity where significantly highest longevity was observed in mungbean variety, PAU 911 (7.28 days) and lowest in genotypes LRB 535 and RBL 1 (4.83 days). Male longevity was found to be more than female longevity among all genotypes in the present study.
Table 1

Biology of C. maculatus on different genotypes of rice bean

Vigna species

Adult emergence (%)

Mean ± standard error

Development period (days)

Growth index

Adult longevity (days)



LRB 470

6.11 (14.21) b

25.91 ± 0.05 d

0.23 ± 0.03 bc

4.62 ± 0.06 bc

5.08 ± 0.09 b

LRB 482

5.00 (12.73) bc

27.67 ± 0.11 b

0.18 ± 0.03 c

5.20 ± 0.12 b

5.50 ± 0.17 b

LRB 507

5.00 (12.73) bc

28.00 ± 0.11 b

0.17 ± 0.00 c

4.87 ± 0.20 b

5.41 ± 0.00 b

LRB 522

4.44 (11.99) c

25.75 ± 0.17 d

0.17 ± 0.02 c

4.75 ± 0.18 b

5.00 ± 0.26 b

LRB 529

3.89 (11.25) c

27.00 ± 0.15 c

0.14 ± 0.02 cd

4.83 ± 0.24 b

5.16 ± 0.08 b

LRB 535

2.22 (7.01) d

30.25 ± 0.21 a

0.07 ± 0.03 e

4.00 ± 0.14 cd

4.83 ± 0.32 bc


6.11 (14.21) b

26.08 ± 0.13 d

0.23 ± 0.03 bc

4.37 ± 0.20 c

4.83 ± 0.27 bc


7.78 (16.11) b

25.17 ± 0.25 e

0.30 ± 0.03 b

5.00 ± 0.17 b

5.16 ± 0.26 b

RBL 35

6.67 (14.95) b

25.25 ± 0.29 e

0.26 ± 0.02 b

4.58 ± 0.16 bc

5.00 ± 0.17 b

RBL 50

6.11 (14.21) b

26.67 ± 0.17 c

0.22 ± 0.02 bc

4.50 ± 0.00 bc

5.00 ± 0.17 b

PAU 911

99.44 (88.21) a

23.19 ± 0.20 f

4.28 ± 0.02 a

7.11 ± 0.35 a

7.28 ± 0.31 a

Figures followed by a common letter(s) are not significantly different by LSD (p = 0.05)

Values represent mean of three replications

Figures in parentheses indicate arcsine transformation

Antinutritional factors

Biochemical analyses were further conducted to study the level of resistance and reason underlying poor development of C. maculatus grubs in different genotypes of rice bean. The study revealed that amount of tannins, phytic acid and saponins was relatively higher in different genotypes of rice bean as compared to mungbean variety, PAU 911 (Table 2). Among rice bean genotypes, the highest amount of tannins was noticed in LRB 507 (11.12 mg g-1) whereas the least amount was recorded in genotypes RBL 6 (8.40 mg g−1). The lowest amount of tannins was observed in mungbean variety, PAU 911 (8.24 mg g−1) among all the test genotypes. The amount of phytic acid was found to be highest in LRB 482 (32.30 mg g−1) which corresponds to more than two times higher quantity than mungbean variety, PAU 911 (13.51 mg g−1). Even LRB 470 which registered the least quantity of phytic acid among all genotypes of rice bean (22.04 mg g−1) had significantly higher amount than PAU 911. Similar was the case with respect to saponin content which showed significant differences and varied from 1.12 mg g−1 (PAU 911) to 4.17 mg g−1 (LRB 535) in seeds of different test genotypes. Among the rice bean genotypes, least saponin content (1.31 mg g−1) was recorded in RBL 1, which was significantly higher than mungbean variety, PAU 911.
Table 2

Amount of various antinutritional factors in different genotypes of V. umbellata

Vigna species

Mean ± standard error

Antinutritional factors (mg/g seed)


Phytic acid


LRB 470

9.48 ± 0.05 bc

22.04 ± 0.14 f

1.44 ± 0.01 ef

LRB 482

8.60 ± 0.12 d

32.30 ± 0.41 a

2.13 ± 0.04 d

LRB 507

11.12 ± 0.05 a

30.01 ± 0.18 b

3.02 ± 0.02 c

LRB 522

9.60 ± 0.05 bc

27.41 ± 0.26 c

3.44 ± 0.14 b

LRB 529

11.05 ± 0.08 a

26.05 ± 0.44 d

1.57 ± 0.06 e

LRB 535

9.90 ± 0.09 b

24.91 ± 0.19 d

4.17 ± 0.10 a


8.81 ± 0.10 cd

23.50 ± 0.49 e

1.31 ± 0.03 f


8.40 ± 0.16 d

25.77 ± 0.54 d

1.63 ± 0.06 e

RBL 35

10.07 ± 0.14 b

29.74 ± 0.66 b

1.49 ± 0.01 e

RBL 50

9.18 ± 0.41 c

31.58 ± 0.09 a

1.53 ± 0.03 e

PAU 911

8.24 ± 0.10 de

13.51 ± 0.39 g

1.12 ± 0.02 g

Figures followed by a common letter(s) are not significantly different by LSD (p = 0.05)

Values represent mean of three replications

Correlation between biochemical seed characteristics and life processes of C. maculatus

Biochemical characteristics of different test genotypes showed a negative correlation with different life processes of C. maculatus except development period where positive correlation was observed (Table 3). Among the antinutritional factor, phytic acid was significantly correlated with adult emergence and growth index (r = −0.78**) at 1% level of significance and with adult longevity (r = −0.61* and −0.64* for male and female, respectively) at 5% level of significance. Saponins were significantly correlated with development period (r = 0.71*) at 5% level of significance.
Table 3

Correlation of biochemical seed characteristics with life processes of C. maculatus

Seed character



Regression line

Tannins to adult emergence



y = −1.544x + 9.71

Phytic acid to adult emergence



y = −14.61x + 28.10

Saponins to adult emergence



y = −1.248x + 2.250

Tannins to development period



y = 0.262x + 2.552

Phytic acid to development period



y = 1.489x - 13.32

Saponins to development period



y = 0.394x - 8.365

Tannins to growth index



y = −0.355x + 9.697

Phytic acid to growth index



y = −3.375x + 27.99

Saponins to growth index



y = −0.286x + 2.24

Tannins to male longevity



y = −0.524x + 12.06

Phytic acid to male longevity



y = −4.047x + 45.88

Saponins to male longevity



y = −0.501x + 4.532

Tannins to female longevity



y = −0.529x + 12.29

Phytic acid to female longevity



y = −4.958x + 52.33

Saponins female longevity



y = −0.460x + 4.517

*Significant at p = 0.05, **Significant at p = 0.01

r Correlation co-efficient, R2 Co-efficient of determination

Co-efficient of determination for different biochemical parameters of test genotypes (Table 3) indicated that phytic acid had maximum contribution toward adult emergence (R2 = 0.61), growth index (R2 = 0.61) and adult longevity (R2 = 0.37 and 0.42 for male and female, respectively), whereas saponins had maximum contribution toward development period (R2 = 0.51) of C. maculatus.


All the test genotypes used in the present study were preferred for oviposition by C. maculatus. However, all the ricebean genotypes failed to support normal development and adult emergence of C. maculatus. Only few adults of smaller size had been emerged with longer development period from ricebean genotypes. On the other hand mungbean variety, PAU 911 supported grub development in a shorter span and considerably higher adult emergence with normal size. Earlier Kashiwaba et al. (2003) observed that several accessions of wild and cultivated V. umbellata completely inhibited adult emergence of C. maculatus. Srinivasan and Durairaj (2007) also support the present findings who reported that 14 out of 17 accessions of V. umbellata, showed no adult emergence of C. maculatus except LRB 113, LRB 282 and LRB 296 which showed long development period (33.33, 32.33 and 38.67 days, respectively) and very low percentage of adult emergence (1.59, 2.08 and 5.14, respectively). In check cultivar CO 6, development period of C. maculatus was comparatively lower (30.90 days) and adult emergence was comparatively higher (28.41%) than rice bean genotypes.

During the present investigations, the death of C. maculatus grubs was observed in the seed cotyledon rather than the seed coat. Previous research by Kashiwaba et al. (2003) also stated that though there was normal egg laying on different accessions of V. umbellata and bruchid grubs were able to penetrate the seed coat but died during first and second instar stage. The possible reason attributed to the non-emergence of adults may be the presence of antinutritional factors in the cotyledons which do not allow the grubs to develop as an adult. The findings of Kashiwaba et al. (2003) also reflected the above inference when they used artificial beans made of V. umbellata flour and found decreased adult emergence and prolonged developmental period of C. maculatus. Furthermore, the biochemical seed characteristics shows a negative correlation with life processes of C. maculatus except development period in the present studies which supported the possible role of antinutritional factors for delay in grub development of C. maculatus and non emergence of adults.

Among the antinutritional factors, the tannin content in all the rice bean genotypes was considerably higher than the susceptible check variety of mungbean, PAU 911 except two genotypes, RBL 6 and LRB 482. Earlier Marconi et al. (1997) reported a range of 7.81–14.87 mg g−1 tannins in different accessions of wild Vigna species (V. oblongifolia, V. racemosa and V. ambacensis) which corroborate the present findings. Saikia et al. (1999) opined 5.13 to 5.72 mg g−1 tannin content in mature seeds of rice bean cultivars which is slightly lower than the amount of tannins observed in present studies. Mubarak (2005) also reported low tannin content in raw mungbean, Phaseolus aureus (3.30 mg g−1 of sample) on dry weight basis. Vincenzo et al. (2005) support the hypothesis that seed coat tannins must be considered in biochemical defense mechanisms which can deter, poison or starve bruchid grubs that feed on cowpea seeds. They also reported that in whole seeds of rice bean genotypes, the total tannin content varied from 4.90 to 8.60 mg g−1 which is in close agreement with the present investigations. Thus seed tannin content may be the possible reason of C. maculatus resistance in V. umbellata genotypes. The higher levels of tannins in different rice bean genotypes might be attributed to seed testa, colour, genotypic make up and variable agro climatic conditions (Awasthi et al. 2011).

The amount of phytic acid was found to be considerably higher in all the rice bean genotypes than the susceptible check variety, PAU 911. Earlier Marconi et al. (1997) reported the phytic acid content (4.30–15.25 mg g−1) in different accessions of eight wild Vigna species. However, the results of our study are at variance, showing higher content of phytic acid in rice bean genotypes. Later Saharan et al. (2002) noticed that the high-yielding varieties of faba bean (VH-82-1) and rice bean (RB-32) registered 10.12–20.18 mg g−1 of phytic acid which is close to the amount of phytic acid noticed in the present studies. In nine cultivars of mungbean, Phaseolus aureus the level of phytic acid was found to be 6.17–9.90 mg g−1 of sample (Tajoddin et al. 2011) whereas in the present studies phytic acid content in mungbean, PAU 911 was little higher (13.51 mg g−1). Thus, in ricebean genotypes, the higher level of phytic acid may be the reason to provide resistance against C. maculatus.

A similar pattern of saponins availability was observed in all the test genotypes as observed in phytic acid content which was considerably higher in all the rice bean genotypes than check variety, PAU 911. Modgil and Mehta (1993) reported that the saponin content in uninfested and 60% infested seeds of chickpea, red gram, and green gram by C. chinensis was 11.19–14.47, 11.60–13.33 and 9.96–12.64 mg g−1, respectively which is comparatively higher than present investigations. Later Saharan et al. (2002) observed that the high yielding varieties of faba bean (VH-82-1) and rice bean (RB-32) showed quite high concentration of saponins (13.13–21.68 mg g−1) when compared with the amount of saponins in present research. The difference may be due to involvement of test genotypes and species in both these studies.

Considering the correlation of biochemical seed characteristics with life processes of C. maculatus, phytic acid content had a highly significant and negative relationship with adult emergence and growth index at 1% level (−0.78**) stressing the importance of phytic acid as a potent antinutritional factor. Moreover the co-efficient of determination in the present studies provide clear cut evidence on the role of phytic acid to a greater extent for conferring resistance in V. umbellata genotypes against C. maculatus followed by saponins among the antinutritional factors studied. Thus the research work emphasizes the presence of resistant genes in the rice bean genotypes which can be used in conventional breeding programmes to incorporate the resistant factors into other crops susceptible to C. maculatus.



The authors wish to express their gratitude to Dr. (Mrs.) Ranjit Kaur Gill, Assistant Plant Breeder, Punjab Agricultural University, Ludhiana for providing seed of different test genotypes for research work.


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Copyright information

© Springer Science+Business Media Dordrecht 2016

Authors and Affiliations

  1. 1.Krishi Vigyan Kendra, S.A.S. Nagar (Mohali)Guru Angad Dev Veterinary and Animal Sciences UniversityLudhianaIndia
  2. 2.Department of Entomology, College of AgriculturePunjab Agricultural UniversityLudhianaIndia
  3. 3.Department of Biochemistry, College of Basic Sciences and HumanitiesPunjab Agricultural UniversityLudhianaIndia

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