Content uploaded by K. M. Shakil Rana
Author content
All content in this area was uploaded by K. M. Shakil Rana on Oct 21, 2020
Content may be subject to copyright.
Asian J. Med. Biol. Res. 2020, 6 (3), 482-490; doi: 10.3329/ajmbr.v6i3.49797
Asian Journal of
Medical and Biological Research
ISSN 2411-4472 (Print) 2412-5571 (Online)
www.ebupress.com/journal/ajmbr
Article
Dietary supplementation of wheatgrass powder to assess somatic response of
juvenile grass carp (Ctenopharyngodon idella)
K. M. Shakil Rana1*, M. A. Salam1, Md. Rakib Ahmmed1 and Al Minan Noor2
1Department of Aquaculture, Bangladesh Agricultural University, Mymensingh, Bangladesh
2Department of Fisheries, 13, Shohid CaptainMoonsur Ali Sharani, Matshya Bhaban, Ramna, Dhaka,
Bangladesh
*Corresponding author: K. M. Shakil Rana, Department of Aquaculture, Bangladesh Agricultural University,
Mymensingh-2202. Phone: +8801728300299; E-mail: ranakms@bau.edu.bd
Received: 19 August 2020/Accepted: 23 September 2020/ Published: 30 September 2020
Abstract: Consequence of dietary fish meal substitution with wheatgrass was evaluated by observing growth
response, associated feed cost and survival of grass carp (Ctenopharyngodon idella) fingerlings for sixty days.
Sprouted wheatgrass (Triticum aestivum) was prepared for its inexpensively rich nutrients. Four isonitrogenous
test diets were formulated and applied as treatments (T) in triplicates (R). In the control (T1), basal inclusion rate
of fish meal was 30%, of which 10% was replaced with wheatgrass powder in T2 and in T3 replacement was
20%. In contrast, 30% of basal fishmeal was replaced in T4. Grass carp fingerlings (6.38±0.21 cm and 2.83±0.36
g) were stocked in twelve aquaria (60×40×45 cm³) each containing 75 L water, at 10 fish per aquarium, fed test
diets at 5% of body weight twice daily. Prominent effect of wheatgrass supplementation was found on food
conversion ratio (FCR) and survival rates. The significantly lowest FCR was observed in T3 (2.13±0.42)
followed by T2 (2.89±0.99), T1 (3.01±1.53) and T4 (3.05±0.94). Besides, fish survival rate was significantly
improved in T2 (90%), T3 (93.33%) and T4 (93.33%) compared to the lowest survival in T1 (83.33%). In
conformity, fish tolerance (LT50) to low pH stressor was also increased with wheatgrass supplementation. The
other growth parameters among the treatments were statistically similar with highest specific growth rate and
fish production in T3 (1.13±0.12 %/day and 2.28±0.13 tons/ha). Dietary wheatgrass did not affect the fish
carcass composition rather gave better result to some extents. The significantly highest carcass protein and
lowest moisture was retained in T3 (14.13±0.05% and 74.91±0.25% respectively), whereas comparatively
higher lipid and mineral (ash) content was found in T1 (7.69±0.02% and 2.35±0.27% respectively). Importantly,
feed formulation cost was reduced by 2.61, 4.89, and 7.71% in T2, T3, and T4 respectively compared to T1.
Therefore, wheatgrass could be promising in juvenile grass carp diet.
Keywords: wheatgrass; grass carp; alternate feed source
1. Introduction
Searching alternative protein sources from non conventional feedstuff to sustain the aqua-feed industry has
become a vital trend of research in the aquaculture world. The current reliance on fishmeal or fish oil for
feeding cultured fish has already caused an enormous loss to the wild fisheries resources (Tacon and Metian,
2008; FAO, 2012). Besides, the increased demand, limited supply at soaring price as well as greater propensity
of fish meal to pollute the environment have made it urgent to replace with less expensive protein sources to
improve the sustainability of aquaculture (Martinez-Llorens et al., 2009). In order to satisfy the quest for finding
competitive replacement of fish meal, scientists have switched over a number of dietary ingredients of both
animal and plant origin (Booth and Sheppard, 1984; Goda et al., 2007; Audu et al., 2010). However, in most of
the cases high cost of animal sources or antinutritional factors of some plant origin had limited their wider use in
aqua-feed (Davis, 2015). Although, plant based fish diets always had financial advancement to offer at
comparatively lower price because of their availability (Azeredo et al., 2017; Francis et al., 2001). Therefore, in
Asian J. Med. Biol. Res. 2020, 6 (3)
483
this experiment wheatgrass (freshly sprouted first leaves of the common wheat plant, Triticum aestivum) has
been investigated as a substitute (plant origin) of fish meal in the diet of grass carp fingerlings to assess its
impact on somatic growth and feed cost. Young sprouted wheatgrass are very rich in dietary fiber, antioxidants,
minerals (potassium, iron, zinc, calcium, magnesium, copper, manganese and selenium) and vitamins (A, C, E,
K, B1, B6, B12 and pantothenic acid) (Meyerowitz, 1992; Murphy, 2002; Shirude, 2011). It also contains a
good proportion of basic nutrients viz., lipid, protein and carbohydrates (Devi et al., 2015). Wheatgrass is well
known as “power house of nutrients”, fifteen pounds of which is nutritionally equal to 350 pounds of ordinary
garden vegetables (Mujoriya and Bodla, 2011; Devi et al., 2015). In spite of having huge prospects (locally
available, low cost, nutrient rich), use of wheatgrass as dietary ingredient in fish feed is still under progress.
However, Nath et al., 2014; and Islam et al., 2017; experimented wheatgrass powder in catfish diets with some
promising findings. However, data on carp fish fed wheatgrass based diet is not well in hand yet. Therefore, the
present experiment was designed to evaluate the effect of dietary wheatgrass powder substitution with fish meal
on juvenile grass carp (Ctenopharyngodon idella).
2. Materials and Methods
2.1. Experimental set-up
The “BAUAquaponics Oasis” laboratory of the Department of Aquaculture, Bangladesh Agricultural
University, was devoted to conduct the experiment for 60 days from 6thJune to 6thAugust 2018. The feed based
experiment, contained of four treatments each with three replications, was run in twelve glass aquaria of 100
liters (size: 60×40×45 cm³), containing 75 L of deep tube well water each. Twelve outlets (with one air stone)
from two air pumps (RESUN, Model ACO-003 and 35 watt) were used for continuous aeration in the aquaria.
The aquaria were labeled individually as T1R1, T1R2 , T1R3 , T2R1, T2R2, T2R3 , T3R1 , T3R2 , T3R3 , T4R1 ,T4R2 ,T4R3
and placed following complete randomized block design for scientific justification. Each aquarium was roofed
with net frame to prevent jumping out of fish or predatory attack.
2.2. Stocking of Fish
The fry of grass carp (initial size: 6.38±0.21 cm and 2.83±0.36 g) was collected from local fish hatchery and
stocked at a rate of 10 fish per aquarium. During fry transportation oxygenated plastic bags were used to avoid
stress and injury. Thereafter the experimental fish were subjected to conditioning into glass aquaria at room
temperature ranging 25-30°C for a period of 8-10 days at the beginning of the experiment. Throughout the
conditioning period, fish were fed control diet twice daily (10:00 am and 4:00 pm) at approximately 3% of live
body weight/day.
2.3. Wheatgrass powder preparation
Firstly, 3 clean plastic buckets were taken containing 1 kg of locally collected wheat seeds each for sprouting.
Then the collected wheat seeds were washed and soaked in water for overnight. In the following morning the
soaked seeds were sieved, wrapped with cotton cloth and kept in a perforated bucket covered with cloths for 24
hours. Subsequently, the germinated wheat seeds were spread over each (60x30x8 cm3) of nine previously
prepared (washed and sun dried) sprouting beds (trays). Water was then sprayed over the trays and covered it
for 2 days. Thereafter the sprouted young yellowish wheatgrass were uncovered and brought to sunlight.
Harvesting was done, when the seedlings became 6-7 inch long with dark green colors within 8 days, with the
cut off stems and weighed. Prior to harvest, water was continued to spray over the sprouting bed twice daily
(morning and evening).
After harvesting, blanching was done where the green wheatgrass stems were boiled (over 100 ºC) for 7 minutes
and immediately cooled down in a big bowl by adding ice and one pinch of sea salt (NaCl). As a thumb rule,
blanching is done by boiling objects at 75 to 105ºC temperature for 1to 10minutes, in order to retard enzymatic
activity, removing gases, setting color, improving texture, stopping the changes of flavor and leaching of water
soluble sugars.
The blanched wheatgrass (Figure 1) were kept aerated in room temperature and then dried into a dryer to give it
a crunchy texture. These were then crumbled with scissor and finally blended to produce fine powder to be used
as fish feed ingredients (Figure 1).
2.4. Ingredients selection and fish feed formulation
The feeding trial was supported with four different experimental diets that were formulated form the following
ingredients based on their availability, nutrient profiles and market price (Table 1).
Asian J. Med. Biol. Res. 2020, 6 (3)
484
The test diets were formulated with a view to reducing (0, 10, 20, and 30%) dietary inclusion of fish meal (basal
inclusion of 30% in control T1) by adding wheatgrass powder in a progressive manner. Accordingly, the control
diet (T1) contained 30% fishmeal but no wheatgrass powder. Whereas in T2, 10% of fish meal was replaced with
wheatgrass powder; hence it contained 27% fish meal and 3% wheatgrass powder. In order to represent 20%
replacement, T3 contained 24% fishmeal and 6% of wheatgrass powder. In T4 30% of fishmeal was replaced
with 9% inclusion of wheatgrass powder (Table 2).
Pearson square method was followed to calculate the dietary inclusion rate of different ingredients in order to
formulate isonitrogenous (around 31%) test diets. Sinking dry pellet feed (0.5 mm diameter) was prepared with
extruded feed pellet machine and sun dried. Prepared feed were stored in air tight polythene bags at 4°C in
refrigerator before feeding the fish. The proximate composition of the test diets was also determined (AOAC,
1990) that has been shown in Table 3. Some precautionary actions were also taken in preparing the ingredients
for feed formulation such as measured mustard oil cake was soaked overnight and soybean meal was pre-boiled
to minimize their glucocyanate effects.
2.5. Fish feeding trial, sampling and data analysis
Throughout the feeding trial, experimental fish were fed (hand feeding) with the respective test diets twice daily
(10 am and 5 pm) at 5% of their body weight in each aquarium. Siphoning was employed to drain 25% of the
aquarium water daily that ensured removal of uneaten feed and faces. However, same amount of clean water
was also added daily to maintain the water level in the aquarium. Besides daily water exchange, each aquarium
was completely drained fortnightly (during sampling) to keep environmental homogeneity.
Sampling of fish and water quality parameters were done biweekly. Fish were sampled randomly from
individual treatment to observe their average length and weight as well as their response to the test diets by
calculating the growth parameters such as length gain (cm), weight gain (g), percent weight gain, specific
growth rate (SGR, %/day), food conversion ratio (FCR), survival rate (%) and fish production (kg/ha). Fish
carcass profile was also determined following the standard procedure of AOAC (1990). Furthermore, water
quality parameters such as dissolved oxygen (mg/L), water temperature (°C), pH, ammonia and nitrite contents
were measured using portable DO meter, thermometer, pH meter and ammonia testing kits respectively.
Moreover, collected data were loaded in the computer and after final harvest, data were subjected to one-way
ANOVA for statistical analysis (Snedecor and Cochran, 1994). The least significant difference was used for
comparison of the mean values ascertained from different treatments by Duncan’s New Multiple Range Test
(Duncan, 1955).
2.6. Low pH stress test
Tolerance to low pH stressor (pH 3) of the experimental fish was also evaluated to determine the impact of
dietary wheatgrass incorporation on the fish fitness. Therefore, after the completion of feeding trail, 6 fish from
each treatment were randomly selected and transferred to a 20 L container holding water having pH 3. In order
to prepare this low pH water (pH 3), deep tube-well water was strongly aerated for 24 h and gradually mixed
with nitric acid (HNO3). The aquaria for stress test were facilitated with continuous aeration and kept under
ambient temperature. Time required attaining 50% mortality of the test fish was calculated as median lethal time
(LT50).
3. Results and Discussion
3.1. Production of sprouted wheatgrass
The total collection of wheatgrass powder from nine sprouting beds (trays) was 500g. In order to obtain this,
each tray (60x30x8 cm3) was initially sprayed with 250 g of raw wheat seeds. Notably, around 20% wheatgrass
powder was retained from the sprouted live wheatgrass. Therefore, average sprouted wheatgrass production rate
was 277.78 g (live weight) per tray.
3.2. Somatic response of grass carp to test diets
3.2.1. Growth parameters
Dietary substitution of any unconventional ingredients always raises the question about its palatability to the
target fish. In this experiment, progressive inclusion of wheatgrass powder in the test diets were well accepted
by the grass carp fingerlings as they fed steadily but actively and there was almost no feed left over after twenty
minutes of feed delivery. Grass carp are herbivorous in nature but also readily accept formulated pellets under
culture conditions (Ni and Wang, 1999; George, 1982). Therefore, having plant origin might have aided
Asian J. Med. Biol. Res. 2020, 6 (3)
485
wheatgrass contributing to the palatability of the diets. However, wheatgrass powder has also been evaluated in
the diet of catfish without hampering diets’ acceptability (Nath et al., 2014; Islam et al., 2017).
In case of initial length and weight there was no significant difference (P>0.05) among the treatments. After 60
days of grass carp nursing, the highest mean length gain (cm) was observed in T3 (1.05±0.39 cm), followed by
T4 (0.97±0.28 cm), T2 (0.95±0.29 cm) and T1 (0.78±0.81 cm), although the values were statistically similar. In
contrast, fish in T3 attained maximum mean weight gain (g) of 2.68±0.18 g, but the lowest was found in T4
(2.13±0.56 g). However, the differences were insignificant (P>0.05) among the treatments (Table 4). The short
rearing period (60 days) and/or some experimental error could be responsible for statistical non-significance.
In a similar fashion, the highest Specific growth rate (SGR, %/day) and fish production (tons/ha) were
experienced in T3 (1.13±0.12%/day and 2.28±0.13 tons/ha) and the lowest in T4 (0.95±0.19 %/day and
2.04±0.24 tons/ha) without any significant difference (P>0.05). The observed SGR (%/day) values have
outweighed the findings of Islam et al. (2017), who reported SGR values ranged from 0.46 to 0.77 %/day for
stinging catfish (Heteropneustes fossilis) and that of Nath et al. (2014), who documented SGR value 0.29 %/day
for Asian catfish (Clarias batrachus) in wheatgrass based feeding trials. Species variation and environmental
factors might be the reason behind these differences. Again, the fish production in this experiment was
continued to increase with the increase of dietary wheatgrass supplementation, till 20% of fish meal replacement
with wheatgrass powder in T3, but dropped when the replacement rose to 30% in T4. Correspondingly, review of
literature has made it known that substitution of fish meal with plant based ingredients up to a certain level in
fish diets results in positive response but higher dietary substitution causes a reduction in growth and immune
responses (Lin and Luo 2011, Mokrani et al., 2020).
3.2.2. Survival rate
Considering the survival rate, fish fed wheatgrass powder based diets showed significantly improved survival
compared to control diet (T1). Therefore, juveniles in T3 and T4 enjoyed the highest survival rate (93.33%)
followed by T2 (90%) which were statistically different (P<0.05) to the lowest survival in T1 (83.33%) (Table 4).
The fabulous nutritional profile of wheatgrass, loaded with quality minerals (K, Ca, Fe, Mg, Na and S) and
vitamins (A, B, C and E) besides the basic nutrients, had presumably contributed to the better survival with
increasing supplement in the test diets (Mujoriya and Bodla, 2011; Anwar et al., 2015; Devi et al., 2015).
Therefore, the overall findings re-emphasize that fishmeal substitution with non-conventional sources (animal or
plant origin) to a certain level could be feasible in fish diet without limiting growth (Ayoola, 2010; Rana et al.,
2015; Sing et al., 2016; Islam et al., 2017; Daniel, 2018; Osho et al., 2019; Rana et al., 2020).
3.3. Food conversion ratio (FCR) and feed cost
Food conversion ratio (FCR) is an effective parameter to evaluate a fish feed as it determines the required
amount of feed for per unit of somatic growth. In this study, the lowest FCR was 2.13±0.42 found in T3 that was
significantly lower (P<0.05) than the highest FCR in T4 (3.05±0.94). Although, the FCR value of T2 (2.89±0.99)
was statistically similar to other treatments (Table 4). Similarly, Rana et al., 2020; substituted fishmeal with jute
leaf powder in the diet of mrigal (Cirrhinus cirrhosus) fingerlings and reported FCR values ranged between
2.81 and 3.30.
Importantly, feed preparation cost was decreased with the increase of dietary wheatgrass powder in the test
diets. For instance, the expense in formulating T1 diet was the highest (49.85 BDT/Kg), which was subsequently
reduced by 2.61%, 4.89%, and 7.71% in T2 (48.55 BDT/Kg), T3 (47.41 BDT/Kg) and T4 (46.01 BDT/Kg)
respectively (Figure 2). Thus the experiment signifies that replacement of fish meal with low cost wheatgrass
powder in the diet of juvenile grass carp could substantially reduce feed cost and show conformity with the
findings of Rana et al., 2020. Moreover, reduction in feed cost, worth more than 60% of total aquaculture cost
(Gadzama and Ndudim, 2019), has brought wheatgrass powder under the shades of light as a promising
ingredients in the diet of Indian major carps.
3.4. Fish carcass composition
Carcass composition of the tested fish has been shown in Table 5. The findings reveal that, significantly highest
carcass protein and lowest moisture was retained in T3 (14.13±0.05% and 74.91±0.25% respectively). In
contrast, comparatively higher lipid and mineral (ash) content was found in T1 (7.69±0.02% and 2.35±0.27%
respectively) which were statistically similar with other treatments except the lowest lipid content in T4
(6.68±0.27%). Besides, fiber and carbohydrate profiles did not vary significantly among the treatments.
Notably, fiber content was increased with progressive addition of wheatgrass in the test diets. However, the
observed protein contents are slightly lower but lipid and mineral (ash) contents are higher than the findings of
Asian J. Med. Biol. Res. 2020, 6 (3)
486
Ashraf et al., 2011; who reported 74.30±0.07% moisture, 20.00±0.15% protein, 2.52±0.01% lipid and 1.4±0.2%
ash content in grass carp. This variation might be due to the diet and age variation of the fish. To sum up,
addition of wheatgrass in place of fishmeal did not affect the quality of fish greatly rather improved consumer
digestibility to some extent by increasing carcass fiber. Nandeesha et al., 1995; also concluded that plant based
diets had contributed to boost up carcass protein and fat levels in Indian major carps.
3.5. Water quality parameters
The water quality parameters (Table 6) of the trial aquaria ranged between, pH (7.82 and 8.84), dissolved (DO)
oxygen (6.57 and 7.93 ppm), temperature (28 and 29.5 ˚C), ammonia (0.08 and 0.33 mg/L) and nitrite (0.07 and
0.66 mg/L) among the treatments throughout the study period. Consequently, the parameters were within the
acceptable range for grass carp as well as fish culture (Swingle, 1967; Ni and Wang, 1999). Therefore, it could
be summarize that the rearing water was safe for fish wellbeing and had no decisive effect on the response of
fish to the test diets.
3.6. Low pH stress test
Adaptation to stressed condition is an important reference of fish robustness. In this experiment, after the
feeding trials fish were subjected to low pH stress test (pH 3.0). From the literature, it is understood that pH
below 4.0 is lethal to fish, whereas the recommended range is 6.8-9.0 (Swingle, 1967). Results showed that 50%
of the fish in T1 died sooner after 6 minutes (LT50 = 6 minutes) of exposure to low pH stress than fish in other
treatments. In contrast, the values of median lethal time (LT50) for the fish in T2, T3 and T4 were 10, 14 and 16
minutes respectively (Figure 3). These imply that dietary addition of wheatgrass (high mineral content) had
improved resilience of the test fish which conforms with the findings of Rana et al., 2020.
Table 1. Basic nutrients and market price of the selected feed ingredients.
Ingredients
% Crude protein
% Crude lipid
% Carbohydrates
Price (BDT/kg)
Fish meal
56
8.5
2.7
80
Wheatgrass powder
16.5
2
60
25
Mustard oil cake
30
11
35
35
Rice bran
12
12
55
35
Soya bean meal
40
15
30
42
Wheat bran
12
7.5
60
20
Wheat flower
12
2.5
70
25
Soya oil
0
100
0
80
Minerals and vitamin premix
0
0
0
100
Table 2. Dietary inclusion rate (g) of different ingredients used in formulating 100 g of the test diets for
grass carp fingerlings.
Feed
ingredients
T1
(0% replacement of
fish meal with
wheatgrass powder)
T2
(10% replacement of
fish meal with
wheatgrass powder)
T3
(20% replacement of
fish meal with
wheatgrass powder)
T4
(30% replacement of
fish meal with
wheatgrass powder)
Fish meal
30
27
24
21
Wheatgrass
0
3
6
9
Mustard oil cake
20
18
18
17
Rice bran
20
17
17
12
Soya bean meal
10
15
18
23
Wheat bran
10
10
10
10
Wheat flower
5
5
2
3
Soya oil
3
3
3
3
Minerals and
vitamin premix
2
2
2
2
Grand Total
100
100
100
100
Asian J. Med. Biol. Res. 2020, 6 (3)
487
Table 3. Proximate composition (%) of different test diets.
Treatments
Moisture
Crude lipid
Crude protein
Ash
Crude fiber
Nitrogen Free
Extract (NFE)
T1
12.32
6.74
31
12.54
4.34
32.31
T2
12.92
6.54
30.86
11.83
5.14
32.18
T3
12.13
5.53
30.51
11.60
5.79
33.69
T4
12.03
5.78
30.58
11.37
6.03
34.11
Table 4. Growth parameters of grass carp fingerlings fed test diets.
Parameters
T1
(0% replacement
of fish meal with
wheatgrass
powder)
T2
(10%
replacement of
fish meal with
wheatgrass
powder)
T3
(20%
replacement of
fish meal with
wheatgrass
powder)
T4
(30%
replacement of
fish meal with
wheatgrass
powder)
F-value
p-value
Level of
Significance
Mean initial
length(cm)
6.33(±0.20)a
6.44(±0.15)a
6.42(±0.39)a
6.34(±0.09)a
0.15
0.93
NS
Mean final
length(cm)
7.11(±0.96)a
7.38(±0.36)a
7.48(±0.25)a
7.32(±0.48)a
0.25
0.86
NS
Mean length
gain
0.78(±0.81)a
0.95(±0.29)a
1.05(±0.39)a
0.97(±0.28)a
0.16
0.92
NS
Mean initial
weight(g)
2.88(±0.46)a
2.89(±0.12)a
2.78(±0.29)a
2.76(±0.58)a
0.15
0.91
NS
Mean final
weight
5.18(±0.14)a
5.24(±0.75)a
5.47(±0.31)a
4.89(±0.57)a
0.68
0.59
NS
Mean weight
gain
2.30(±0.58)a
2.35(±0.72)a
2.68(±0.18)a
2.13(±0.56)a
0.54
0.67
NS
Percent
weight gain
83.41(±35.24)a
81.35(±24.5)a
97.21(±14.4)a
77.27(±20.58)a
0.36
0.78
NS
FCR
3.01(±1.53)a
2.89(±0.99)ab
2.13(±0.42)b
3.05(±0.94)a
2.85
0.049
*
SGR
(%/day)
0.99(±0.31)a
0.98(±0.22)a
1.13(±0.12)a
0.95(±0.19)a
0.39
0.76
NS
Survival rate
(%)
83.33b
90.00a
93.33a
93.33a
3.83
0.042
*
Fish
production
(tons/ha)
2.16 (±0.06)a
2.19(±0.31)a
2.28(±0.13)a
2.04(±0.24)a
0.682
0.59
NS
Note: Values are mean ±Standard deviation from triplicate groups. Values in a row having similar letters (s) or without
letters do not differ significantly whereas values bearing the dissimilar letter (s) differ significantly as per DMRT
(Duncan’s New Multiple Range Test). * significant at P≤ 0.05; ** significant at P≤ 0.01; NS non-significant at P˃ 0.05
Table 5. Carcass composition (%) of experimental grass carp (wet weight basis) fingerlings.
Parameters
T1
(0%
replacement of
fish meal with
wheatgrass
powder)
T2
(10%
replacement of
fish meal with
wheatgrass
powder)
T3
(20%
replacement of
fish meal with
wheatgrass
powder)
T4
(30%
replacement of
fish meal with
wheatgrass
powder)
F-value
p-value
Level of
Significance
Moisture
75.32(±0.12)bc
75.67(±0.59)ab
74.91(±0.25)bc
76.04(±0.09)a
6.52
0.02
*
Crude lipid
7.69(±0.02)a
7.59(±0.41)a
7.41(±0.23)a
6.68(±0.27)b
8.65
0.007
**
Crude protein
13.73(±0.15)b
13.45(±0.25)b
14.13(±0.05)a
13.47(±0.13)b
11.88
0.003
**
Ash
2.35(±0.27)a
2.20(±0.07)a
2.32(±0.09)a
2.33(±0.11)a
0.56
.66
NS
Fiber
0.48(±0.31)a
0.47(±0.18)a
0.62(±0.13)a
0.61(±0.03)a
0.53
0.68
NS
Carbohydrate
0.42(±0.10)b
0.59(±0.22)ab
0.60(±0.29)ab
0.88(±0.08)a
2.95
0.09
NS
Note: Values are mean ±Standard deviation from triplicate groups. Values in a row having similar letters (s) or without
letters do not differ significantly whereas values bearing the dissimilar letter (s) differ significantly as per DMRT
(Duncan’s New Multiple Range Test). * significant at P≤ 0.05; ** significant at P≤ 0.01; NS non-significant at P˃ 0.05
Asian J. Med. Biol. Res. 2020, 6 (3)
488
Table 6. The water quality parameters over the feeding trial.
Parameters
T1
(0% replacement of
fish meal with
wheatgrass powder)
T2
(10% replacement
of fish meal with
wheatgrass powder)
T3
(20% replacement
of fish meal with
wheatgrass powder)
T4
(30% replacement
of fish meal with
wheatgrass powder)
pH
8.4±0.44
8.2±0.1
8.33±0.32
8.03±0.21
DO (ppm)
7.87±0.06
7.73±0.12
7.33±0.76
7.63±0.38
Temperature (˚C)
28.57±0.40
28.6±0.17
29±0.5
28.4±0.40
Ammonia(mg/L)
0.14±0.06
0.22±0.11
0.14±0.05
0.15±0.08
Nitrite(mg/L)
0.36±0.30
0.29±0.18
0.07±0.00
0.13±0.04
A. Blanched wheatgrass stem
B. Wheatgrass powder
Figure 1. Preparation of wheatgrass powder.
Figure 2. Feed formulation cost in different treatments. (BDT/Kg)
Figure 3. Response of fish to low pH stress test.
4. Conclusions
The overall somatic performance of grass carp (C. idella) fed wheatgrass supplemented test diets was
satisfactory compared to control diet (no dietary wheatgrass inclusion). Notably, fish survival was substantially
improved. The FCR value and feed formulation cost of the respective diets were reduced in a good amount.
Asian J. Med. Biol. Res. 2020, 6 (3)
489
However, the sensitivity to wheatgrass (plant feedstuffs) based diets is species specific and demand extensive
research to certify it as a fish feed ingredient in a broad context.
Conflict of interest
None to declare.
References
Anwar DA, AA El-Yazied, TF Mohammadi and MMF Abdallah, 2015. Wheatgrass Juice and its nutritional
value as affected by sprouting condition. Arab Univ. J. Agril. Sci., 23: 37-49.
AOAC. 1990. Official methods of analysis. 15th edn. Association of Official Analytical Chemists, Washington,
D.C., USA.
Ashraf M, A Zafar, A Rauf, S Mehboob and NA Qureshi, 2011. Nutritional values of wild and cultivated
silver carp (Hypophthalmichthys molitrix) and grass carp (Ctenopharyngodon idella). Int. J. Agric. Biol.,
13: 210–214.
Audu BS, KM Adamu and SA Binga, 2010. The effect of substituting fishmeal diets with varying quantities of
ensiled parboiled beniseed (Sesamum indicum) and raw african locust bean (Parkia biglobosa) on the growth
responses and food utilization of the nile tilapia Oreochromis niloticus. Int. J. Zool. Res., 6: 334-339.
Ayoola AA, 2010. Replacement of fish meal with alternative protein source in aquaculture diets. M.Sc. Thesis,
Graduate Faculty of North Carolina State University, U.S.A. pp. 1-120.
Azeredo R, M Machado, E Kreuz, S Wuertz, A Oliva-Teles, P Enes and B Costas, 2017. The european seabass
(Dicentrarchus labrax) innate immunity and gut health are modulated by dietary plant-protein inclusion and
prebiotic supplementation. Fish Shellfish Immunol., 60: 78–87.
Booth DC and DC Sheppard, 1984. Oviposition of the black soldier fly Hermetia illucens diptera stratiomyidae
egg masses timing and site characteristics. Env. Entom., 13: 421-423.
Daniel N, 2018. A review on replacing fish meal in aqua feeds using plant protein sources. Int. J. Fish. Aquat.
Stud., 6: 164-179.
Davis DA, 2015. Feed and feeding practices in aquaculture. Woodhead Publishing, Cambridge, Britain.
Devi SK, K Hariprasath, GR Nalini, P Veenaeesh and S Ravichandra, 2015. Wheat grass juice - Triticum
aestivum Linn’ a therapeutic tool in pharmaceutical research, an overview. Ijppr. Human., 3: 112-121.
Duncan DB, 1955. Multiple range and multiple F tests. Biometrics, 11: 1-42.
FAO. 2012. Fisheries statistics. Food and Agricultural Organization of the United Nations (FAO), Rome, Italy.
Francis G, HPS Makkar and K Becker, 2001. Antinutritional factors present in plant-derived alternate fish feed
ingredients and their effects in fish. Aquaculture, 199: 197–227.
Gadzama and Ndudim. 2019. Nutritional composition of housefly larvae meal: a sustainable protein source for
animal production – a review. Acta. Sci. Agri., 3: 74-77.
George TT, 1982. The chinese grass carp, Ctenopharyngodon idella, its biology, introduction, control of aquatic
macrophytes and breeding in the Suda. Aquaculture, 27: 317-327.
Goda AM, ER El-Haroun and MAK Chowdhury, 2007. Effect of totally or partially replacing fish meal by
alternative protein sources on growth of African catfish Clarias gariepinus (Burchell, 1822) reared in
concrete tanks. Aquac. Res., 38: 279-287.
Islam T, KMS Rana and MA Salam, 2017. Potential of wheatgrass powder based feed for stinging catfish fry
nursing in laboratory condition. Int. J. Fish. Aquat. Stud., 5: 179-184.
Lin S and L Luo, 2011. Effects of different levels of soybean meal inclusion inreplacement for fish meal on
growth, digestive enzymes and transaminaseactivities in practical diets for juvenile tilapia, Oreochromis
niloticus × O. aureus. Anim. Feed Sci. Technol. 168: 80–87.
Martínez-Llorens, Silvia, AT Vidal, IJ Garcia, MP Torres and MJ CerdÁ. 2009. Optimum dietary soybean meal
level for maximizing growth and nutrient utilization of on-growing gilthead sea bream (Sparus Aurata).
Aquac. Nutr., 15: 320–28.
Meyerowitz S, 1992. “Nutrition in grass”-wheatgrass nature’s finest medicine: the complete guide to using grass
foods & juices to revitalize your health. 6th ed. Great Barrington (Massachusetts): Sproutman publications,
USA. pp. 1-53.
Mokrani A, M Ren, H Liang, Q Yang, K Ji, HC Kasiya and X Ge, 2020. Effect of the total replacement of
fishmeal with plant proteins and supplemental essential amino acids in the extruded diet on antioxidants
genes, enzyme activities, and immune response in juvenile blunt snout bream. Aquacult. Int., 28: 555–568.
Mujoriya R and DRB Bodla, 2011. A study on wheat grass and its nutritional value. Food Sci. Qual. Mngmt.,
2: 1-8.
Asian J. Med. Biol. Res. 2020, 6 (3)
490
Murphy S, 2002. Wheatgrass, healthy for the body and the bank account. ABC Landline. Archived from the
original on 2 December 2002. Retrieved 6 October 2006.
Nandeesha MC, SS De Silva and D Krishna, 1995. Use of mixed feeding schedules in fish culture: performance
of common carp, Cyprinus carpio L. on plant and animal based diets. Aquac. Res., 26: 161-166.
Nath DT, S Hashem, and MA Salam, 2014. Asian catfish fry (Clarias batrachus) rearing with wheatgrass
powder mixed formulated feed in plastic half drum. Int. J. Fish. Aquat. Stud., 1: 162-168.
Ni D and J Wang, 1999. Biology and diseases of grass carp. Ed. Ni D and J Wang, Science Press, Beijing,
China.
Osho FE, EK Ajani, O Orisasona, and T Obafemi, 2019. Replacement of fishmeal with clariid fish offal in the
diet of african catfish, Clarias gariepinus (Burchell, 1822) juveniles. Nig. J. Fish. Aqua., 7: 40 – 48.
Rana KMS, MA Salam, S Hashem and MA Islam, 2015. Development of black soldier fly larvae production
technique as an alternate fish feed. Int. J. Res. Fish. Aqua., 5: 41-47.
Rana KMS, P Biswas, and MA Salam, 2020. Evaluation of jute leaf as substitute of fish meal in the diet of
mrigal (Cirrhinus cirrhosus) fingerlings. Int. J. Agril. Res. Innov. Tech., 10: 117-122.
Shirude AA, 2011. Phytochemical and pharmacological screening of wheatgrass ( Triticum Aestivum L). Int. J.
Pharma. Sci. Rev. Res., 9: 159-164.
Singh P, BN Paul, GC Rana, and SS Giri, 2016. Evaluation of jute leaf as feed ingredient for Labeo rohita
fingerlings. Indian J. Anim. Nutr., 33: 203-207.
Snedecor GW and WG Cochran, 1994. Statistical methods. Oxford and IBH Publishing Company, Calcutta,
West Bengal, India.
Swingle HS, 1967. Standardizations of chemical analyses for water and pond muds. FAO Fish Rep., 4: 397-421.
Tacon AGJ and M Metian, 2008. Global overview on the use of fish meal and fish oil in industrially
compounded aquafeed: trends and future prospects. Aquaculture, 285: 148-158.
