Biochemical Analysis of Mulberry Leaves (Morus alba L.) and Silkworm Bombyx mori Enriched with Vermiwash
A study was carried out to evaluate the effect of vermiwash supplementation on silk worm Bombyx mori. The vermiwash was diluted to different concentration as 10, 25 and 50%. Fresh mulberry leaves were sprayed by each concentration of vermi wash. The leaves were then fed to silk worms from 5th instar stage onwards at four times per day. Group 1 larvae received mulberry leaves without vermiwash spray and served as control, group 2 larvae received 10% vermiwash sprayed mulberry leaves, group 3 larvae received 25% vermiwashsprayed mulberry leaves and group 4 larvae received 50% vermiwash sprayed mulberry leaves. Weight of larvae, silk gland, cocoon, pupa and shell weight of silkworm was significantly increased. Minimum larval weight (2.160 g) was in control and maximum (3.416 g) was in 50% enriched larvae. Silk gland weight of control, 10, 25 and 50% enrichment was 0.511, 0.552, 0.661 and 0.738 g, respectively. The cocoon weight of control, 10, 25 and 50% treatment was 1.065, 1.243, 1.296 and 1.374 g respectively. The shell weight of control (0.171 g) was lower than 10% treatment (0.198 g), 25% treatment (0.247 g) and 50% treatment (0.315 g). There was significant increase in the carbohydrate, protein and lipid of fat body and mulberry leaves. Acid phosphatase and alkaline phosphatase are significantly increased in the body of B. mori.
Received: November 05, 2011;
Accepted: February 17, 2012;
Published: March 27, 2012
Sericulture is one of the oldest industries in India and probably dates back
to the beginning of the Christian era. As far as the Indian scenario is concerned
sericulture is an especially significant industry, the reasons are varied. India
being the second largest producer of silk, next to China at an annual turnover
rate of 20,000 metric tonnes and accounting 20% of world production. About 65%
of total silk production of India comes from Karnataka alone (Singhvi
et al., 2002). During the last fifteen years, the production of mulberry
silk was increased from 2.258 metric tonnes to 6,900 metric tonnes. The Bombyx
mori L. (silkworm) is a phytophagous feeder on Morus alba L. (mulberry
leaves). Scientists have tried alternative hosts for the rearing of silkworm,
but they were not cost effective. Based on the nutritional quality, mulberry
has a great influence on the silkworm growth, silk yield and disease resistance
Earthworms play a vital role in plant growth. Recently, the commercial vermi
culturists have started promoting a product called vermi wash. Vermiwash contains
enzyme cocktail of proteases, amylases, urease and phosphatase and secretions
of earthworms which would stimulate the growth and yield of crops and even develop
resistance in crops receiving this spray. The effect of nitrogen, vitamin and
salts supplementation on the growth of silkworm has been investigated by many
researchers (Mahmood et al., 2002; Kanafi
et al., 2006; Dhiraj and Venkatesh Kumar, 2012),
but the reports on the effect of mulberry leaves enriched with vermiwash were
limited. So, the present study was aimed to find out the effective dose of vermiwash
application to mulberry leaves to find out its effect on increment of weight
of larvae, silk gland, cocoon, pupa and shell weight of silkworm.
MATERIALS AND METHODS
The eggs of silkworm B. mori were collected from Sericulture Training
Institute, Nanjikottai, Thanjavur, Tamil Nadu (India). The culture of larvae
and mulberry leaves (MR2 variety) were standardized by following
the method of Krishnan et al. (1995). The larvae
were reared in plastic trays (20 larvae/tray) and were fed with mulberry leaves.
Fresh mulberry leaves were collected every day morning and stored in wet gunny
bags. The larvae were fed four times a day (8:00, 13:00, 18:00 and 27:00 h).
Bedding and spacing were adopted carefully at the time of rearing (Krishnaswami
et al., 1978). Enough care was taken to maintain the humidity (75-80%)
and temperature (25-28°C) throughout the study period.
The mulberry trees were enriched with the vermiwash in three different concentrations
such as 10, 25 and 50% once in a week by using foliar spray for 4 weeks. For
each concentration of vermiwash application, separate plots were used and untreated
control plot was maintained separately in the neighboring field. The worms were
fed with these enriched leaves every day and were supplied for the entire period
of 5th instar larvae. Finally, the larval weight, cocoon weight, pupa weight
and shell weight were measured and estimation of biochemical parameters of mulberry
leaves and fat body of silkworm larvae were carried out. The carbohydrate was
estimated by the Anthrone method of Seifter et al.
(1950), protein was estimated by the method of Lowry
et al. (1951) and lipid were estimated by chloroform methanol method
(Folch et al., 1957). Comparison was made between
all the treatment groups by drawing bar charts using Microsoft excel.
The changes in the larval and cocoon characteristics of silkworm B. mori
were studied when the larvae fed on mulberry leaf enriched with vermiwash
(Fig. 1, 2). There was increased weight
in enriched larvae when compared to control larvae. Minimum larval weight (2.160
g) was in control and maximum (3.416 g) was in 50% enriched larvae. The silk
gland weight is significantly increased in the vermiwash enriched larvae. Silk
gland weight of control, 10, 25 and 50% enrichment was 0.511, 0.552, 0.661 and
0.738 g, respectively. The mean weight of cocoon was increased in the vermiwash
enriched larvae. The cocoon weight of control, 10, 25 and 50% treatment was
1.065, 1.243, 1.296 and 1.374 g, respectively. The data obtained on the pupal
weight of the control and enriched group were recorded. The increase in the
pupal weight was clearly noticed in the vermi wash enriched groups.
|| Weight of 5th instar
|| Weight of silk gland, cocoon, pupal and shell weight of enriched
Weight of pupa in the control, 10, 25 and 50% was 0.894, 1.045, 1.049 and 1.059
g. The shell weight was significantly increased in the enriched groups when
compared to the control. The shell weight of control (0.171 g) was lower than
10% treatment (0.198 g), 25% treatment (0.247 g) and 50% treatment (0.315 g).
Biochemical studies from mulberry leaf: In all treatments, the level of carbohydrate is higher than the control. Carbohydrate levels in control leaf was 14 mg, in 10% vermiwash enriched leaf it was 14.40 mg, in 25% enriched leaf it was 14.80 mg and in 50% enriched leaf it was 15.0 mg. The protein content was significantly higher in vermiwash enriched leaf than control. The protein level of the control leaf was 2.693 mg. The protein content of 10% enriched leaf was 2.704 mg, 25% enriched leaf was 2.714 mg and 50% enriched leaf was 2.721 mg. The lipid level of control leaf was 0.052 mg. In 10, 25 and 50% enrichment, it was 0.063, 0.076 and 0.094 mg, respectively (Fig. 3).
Biochemical studies from silkworm: The level of carbohydrate in the total body tissue of the control, 10, 25 and 50% vermiwash enriched larvae was 16, 16.40, 16.80 and 17.40 mg, respectively. The protein level in the control, 10, 25 and 50% enriched larvae was 1.975, 1.989, 1.996 and 2.007 mg, respectively. The lipid level in the control, 10, 25 and 50% enriched larvae was 0.056, 0.062, 0.080 and 0.089 mg, respectively (Fig. 4).
Enzyme studies: The acid phosphatase level in the control larvae was 2.1 IU L-1 and significant increase was observed in 10% (3.5 IU L-1), 25% (7.2 IU L-1) and 50% enriched larvae (9.4 IU L-1). The alkaline phosphatase level in the control was (33 IU L-1) lower than 10% (76 IU L-1), 25% (124 IU L-1) and 50% enriched (163 IU L-1) body tissue of B. mori (Fig. 5).
|| Proximate composition of enriched and control mulberry leaf
|| Proximate composition of enriched and control fifth instar
|| Acid and alkaline phosphatase of enriched and control
In the present study, the fifth instar larvae of B. mori were fed on
mulberry leaves which was enriched with vermiwash in different concentrations.
The increases in the larval, pupal and shell weight of fifth instar larvae of
B. mori were observed. The consumption of mulberry leaf by B. mori
is significantly increased when they enriched with vermiwash and correspondingly
increased the assimilation and production efficiency of the larvae. It was due
to the volume of enzyme secreted in the digestive system resulted in the improvement
of digestion. In the present investigation; larval, silk gland, cocoon, pupal
and shell weight of B. mori were increased when larvae fed with vermiwash
enriched mulberry leaves than the untreated control. Chikkaswamy
et al. (2001) and Mahmood et al. (2002)
reported that application foliar sprays of green leaf significantly improve
the larval, silk gland, cocoon, pupal and shell weight of B. mori. larvae
fed with mulberry leaves enriched with Amway protein showed significant enhancement
larval, cocoon, shell weight (Rani et al., 2011).
Singhvi et al. (2002) reported foliar application
of seriboost significant increase larval weight.
In the present study, biochemical composition of mulberry leaves enriched with
vermiwash and control leaf were analysed. The level of the carbohydrate, protein
and lipid is greater in vermiwash enriched leaves than the control. Foliar sprays
of mulberry leaves increased the biochemical parameters (Chikkaswamy
et al., 2001). The growth rate of plants is also increased in the
vermiwash enriched than control. Dhiraj and Venkatesh Kumar
(2012) reported that foliar sprays influence the performance of mulberry
varieties in terms of physiological parameters of plants viz., carbohydrates,
protein etc. Ashfaq et al. (2000) studied the
various minerals viz., N, P, K, Ca, Mg and Cu, respectively in various combination
were enhanced silk production. Venkataramana et al.
(2009) studied the effect of enrichment of different concentration of vermiwash
and cow dung wash to improve the biochemical and yield attributing parameters
and yield of mulberry. The biochemical analysis of B. mori fed with vermiwash
enriched leaves showed the increase in carbohydrate, protein and lipid content
in the total body tissue of fifth instar larvae than control. These results
are in accordance with the finding of Dhiraj and Venkatesh
Kumar (2012). Singhvi et al. (2002) reported
foliar application of seriboost significant increase in the biochemical parameters
of silk worm in his study. Santhi et al. (2003)
reported that protein content increased with the application of humic acid in
rice, maize and sugarcane. Nandakumar et al. (2004)
also reported that foliar application of Humic acid (HU) in combination with
NPK increased soil nutrients (N, P, K, Fe, Mn, Zn and Cu) which is essential
growth factors of rice. Carbohydrate, protein and lipid are the main sources
of energy at the time of larval-larval, larval-pupal, pupal-adult transformation
Venkataramana et al. (2009). The level of enzymes,
such as alkaline phosphatase and acid phosphatase, in the body of fifth instar
larvae showed significant increase when they fed with vermiwash enriched leaves.
In this study, it is understood that the vermiwash enriched larvae showed an increased larval weight which leads to higher gain of cocoon weight and increased silk production. By knowing this, it is suggested that the farmers are advised for vermiwash enriched larval culture techniques.
The authors are thankful to the Sericulture Training Institute, Nanjikottai, Thanjavur, Tamil Nadu, India and the authorities of AVVM Sri Pushpam College (Autonomous), Poondi for providing the facilities.
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