Effects of supplemental phytase on growth, nutrient digestibility and anti-oxidant enzyme activity in the gills of juvenile mrigal, Cirrhinus mrigala (Hamilton 1882) fed distillers dried grains with soluble based diets

Figures & data

Table 1. Formulation and chemical investigation of the experimental diets.

	Dietary Phytase level (FTU /kg)
Ingredients^1 (g /kg DM)	D1-0	D2-250	D3-500	D4-750	D5-1000	D6-1250
Fish meal	140	140	140	140	140	140
Soybean meal	140	140	140	140	140	140
Corn DDGS	540	540	540	540	540	540
Wheat flour	70	70	70	70	70	70
Tapioca flour	15	15	15	15	15	15
Fish (cod) liver oil	70	70	70	70	70	70
Vitamin mixture^2	10.0	10.0	10.0	10.0	10.0	10.0
Mineral mixture^2	10.0	10.0	10.0	10.0	10.0	10.0
Chromic oxide (Cr2O3)	5.0	5.0	5.0	5.0	5.0	5.0
Chemical analysis [Dry matter basis (%); mean ± SE, number of determinations = 3].
Moisture	8.2 ± 0.9	7.9 ± 0.4	7.7 ± 0.3	7.9 ± 0.4	7.9 ± 0.3	7.9 ± 0.8
Crude protein^3 (% DM)	33.3 ± 1.4	32.9 ± 1.8	32.8 ± 1.3	32.7 ± 1.6	33.1 ± 1.7	32.9 ± 1.4
Crude lipid (% DM)	9.6 ± 1.5	10.4 ± 1.6	10.2 ± 1.4	10.3 ± 1.6	10.0 ± 1.3	9.7 ± 1.2
Total phosphorus (% DM)	0.61 ± 0.03	0.62 ± 0.05	0.60 ± 0.03	0.58 ± 0.01	0.60 ± 0.03	0.63 ± 0.04
Phytate phosphorus (% DM)	0.25 ± 0.02	0.27 ± 0.01	0.26 ± 0.04	0.25 ± 0.05	0.27 ± 0.03	0.28 ± 0.02
Non-phytate phosphorus (% DM)	0.36 ± 0.04	0.35 ± 0.03	0.34 ± 0.01	0.33 ± 0.02	0.36 ± 0.01	0.35 ± 0.03
Crude fiber (% DM)	13.4 ± 0.52	13.3 ± 1.3	13.5 ± 1.1	13.2 ± 1.2	13.1 ± 0.9	13.4 ± 1.5
Ash (% DM)	6.9 ± 0.5	6.8 ± 0.7	7.3 ± 0.4	7.4 ± 1.3	7.3 ± 0.6	7.8 ± 0.5
NFE^4 (% DM)	29.1 ± 0.4	28.7 ± 0.9	28.8 ± 0.7	28.2 ± 0.8	28.3 ± 0.5	28.5 ± 0.6
Phytase^5 (FTU /kg)	<80 ± 0.2	221 ± 0.1	476 ± 0.3	710 ± 0.2	930 ± 0.4	1215 ± 0.2
Gross energy (kJ g^–1 DM)	21.3 ± 0.2	21.2 ± 0.4	21.3 ± 0.6	21.4 ± 0.7	21.2 ± 0.1	21.3 ± 0.5
Digestible protein^6 (DCP, % DM)	23.92 ± 1.37	27.62 ± 1.52	28.53 ± 1.31	29.78 ± 1.66	30.22 ± 1.83	30.63 ± 1.52
Digestible lipid^7 (DL, % DM)	8.77 ± 1.19	9.68 ± 1.41	9.85 ± 1.20	10.23 ± 1.6	9.94 ± 1.2	9.69 ± 1.32
Digestible phosphorus^8 (DP, % DM)	0.56 ± 0.01	0.53 ± 0.02	0.54 ± 0.01	0.53 ± 0.02	0.55 ± 0.03	0.54 ± 0.01
Digestible energy^9 (DE, kJ g^–1 DM)	11.49 ± 1.27	13.98 ± 1.45	16.15 ± 1.44	17.54 ± 1.39	18.77 ± 1.80	19.77 ± 1.38
DCP/DE (mg kJ^–1)	18.52 ± 1.65	18.76 ± 1.42	18.67 ± 1.59	17.98 ± 1.88	17.70 ± 1.92	17.56 ± 1.40

¹ Feed ingredients were purchased from local markets of Bahawalpur and Karachi, Pakistan

² Mixture of minerals and vitamin were purchased from local markets of Bahawalpur and Karachi, Pakistan

³ Kjeldahl method (Nitrogen × 6.25).

⁴ NFE = nitrogen-free extract [carbohydrate content = 100 – (% moisture + % protein + % fat + % ash + % fiber)].

⁵ FTU = One phytase unit (FTU) is the number of enzymes that liberates 1 micromole of inorganic phosphorus per minute from 0.0051 mol/l sodium phytate at 37 °C and pH 5.50 under the conditions of the test. Phosphorus was determined by spectrophotometric method using molybdovandate reagent.

⁶ Digestible protein = crude protein × protein digestibility/100.

⁷ Digestible lipid = crude lipid × lipid digestibility/100.

⁸ Digestible phosphorus = crude phosphorus × phosphorus digestibility/100.

⁹ Digestible energy = gross energy × energy digestibility/100; Digestible protein to digestible energy ratio = digestible protein mg / digestible energy kJ.

Table 2. The growth rate, feed utilization efficiency and apparent digestibility of protein, lipid and energy % of juvenile mrigal fed different dietary phytase levels for 90 days.

	Dietary Phytase level (FTU /kg)
Parameters	D1-0	D2-250	D3-500	D4-750	D5-1000	D6-1250
Initial weight (g)	12.14 ± 1.49^a	12.12 ± 1.52^a	12.14 ± 1.48^a	12.13 ± 1.50^a	12.11 ± 1.51^a	12.15 ± 1.53^a
Final weight (g)	64.81 ± 1.53^a	73.52 ± 1.34^b	83.63 ± 1.13^c	97.62 ± 1.33^d	97.55 ± 1.80^d	97.62 ± 1.52^d
Weight gain (WG, g)	52.67 ± 1.42^a	61.40 ± 1.27^b	71.49 ± 1.40^c	85.49 ± 1.39^d	85.44 ± 1.28^d	85.47 ± 1.32^d
WG^1 (%)	432.42 ± 6.28^a	501.43 ± 8.30^b	584.41 ± 9.43^c	697.13 ± 5.22^d	696.70 ± 5.45^d	696.61 ± 8.22^d
SGR^2 (% IBW/day)	1.87 ± 0.01^a	1.98 ± 0.02^b	2.24 ± 0.03^c	2.33 ± 0.02^d	2.32 ± 0.04^d	2.32 ± 0.02^d
FI^3 (g DM fish^−1)	639.6 ± 2.11^d	596.35 ± 4.66^c	562.9 ± 3.58^b	525.33 ± 2.73^a	524.8 ± 4.41^a	524.73 ± 6.45^a
FCR^4	1.47 ± 0.01^d	1.19 ± 0.03^c	0.96 ± 0.02^b	0.74 ± 0.03^ab	0.74 ± 0.01^a	0.74 ± 0.03^a
PER^5	2.43 ± 0.03^a	2.57 ± 0.02^b	2.75 ± 0.03^c	3.12 ± 0.01^d	2.78 ± 0.02^c	2.53 ± 0.04^b
ADC^6 (dry matter)	50.34 ± 0.21^a	51.20 ± 1.20^a	52.4 ± 1.55^b	64.5 ± 1.63^c	72.0 ± 0.72^d	81.1 ± 1.45^e
ADC^6 (protein)	71.84 ± 1.83^a	83.95 ± 1.57^b	86.99 ± 1.65^bc	91.10 ± 1.52^c	91.31 ± 1.93^c	93.10 ± 1.44^c
ADC^6 (lipid)	85.11 ± 2.01^a	93.11 ± 1.51^b	96.55 ± 1.62^bc	97.81 ± 1.14^c	98.40 ± 1.43^c	98.42 ± 1.57^c
ADC^6 (phosphorus)	73.17 ± 2.89^a	86.45 ± 3.58^b	91.82 ± 4.02^bc	95.88 ± 2.54^c	96.36 ± 5.13^c	97.11 ± 1.44^c
ADC^6 (energy)	53.98 ± 1.46^a	65.93 ± 1.75^b	75.83 ± 1.55^c	81.94 ± 1.24^d	88.53 ± 1.74^e	92.81 ± 1.43^f
Survival (%)	100	100	100	100	100	100

Mean ± SE, n = 3 and each n consists of 18 fish per replicate. Values of the similar row having different superscripts are found significant (P < 0.05).

¹ Weight gain, WG % =  100 × [(final body weight – initial body weight) / initial body weight]

² Specific growth rate, SGR % =  100 × [ln final body weight – ln initial body weight / days]

³ Feed conversion ratio, FCR = total feed fed (g) / total wet weight gain (g)

⁴ Feed intake, FI = total feed fed as % body weight – total uneaten feed

⁵ Protein efficiency ratio, PER = weight gain / protein intake

⁶ Apparent digestibility coefficient of nutrient or energy, ADC = 100 × [1 − (dietary Cr₂O₃ / fecal Cr₂O₃) × (fecal nutrient or energy/ dietary nutrient or energy)]

Survival rate % =  100 × (final body weight)/ (initial body weight).

Table 3. Total proximate composition (% on wet weight basis) of whole body, muscle, liver and viscera of juvenile mrigal fed various dietary phytase levels for 90 days.

	Dietary Phytase level (FTU /kg)
Parameters	D1-0	D2-250	D3-500	D4-750	D5-1000	D6-1250
Whole body
Moisture	73.45 ± 1.78	72.89 ± 1.67	73.12 ± 1.45	73.34 ± 1.56	73.56 ± 1.56	73.56 ± 2.01
Protein	19.45 ± 0.56	19.78 ± 0.56	19.23 ± 0.67	19.45 ± 0.45	19.78 ± 0.89	19.56 ± 0.78
Lipid	9.34 ± 0.56	9.12 ± 0.57	9.78 ± 0.89	9.67 ± 0.89	8.56 ± 0.66	8.48 ± 0.79
Ash	2.45 ± 0.67	2.34 ± 0.78	2.46 ± 0.13	1.76 ± 0.77	2.25 ± 0.46	2.34 ± 0.86
Phosphorus	0.57 ± 0.01	0.55 ± 0.03	0.56 ± 0.01	0.53 ± 0.02	0.56 ± 0.03	0.58 ± 0.01
Muscle
Moisture	72.89 ± 0.98	72.65 ± 1.33	72.98 ± 1.68	72.55 ± 1.57	72.78 ± 1.35	72.98 ± 1.63
Protein	19.35 ± 1.68	19.56 ± 1.24	19.48 ± 1.88	19.34 ± 1.45	19.57 ± 1.78	19.64 ± 1.45
Lipid	2.87 ± 0.67	2.78 ± 0.45	2.56 ± 0.24	2.45 ± 0.56	2.45 ± 0.68	2.35 ± 0.45
Ash	2.95 ± 0.58	2.95 ± 0.25	2.35 ± 0.34	2.87 ± 0.86	2.88 ± 0.89	2.71 ± 0.56
Phosphorus	0.53 ± 0.02	0.50 ± 0.01	0.52 ± 0.01	0.54 ± 0.03	0.57 ± 0.02	0.56 ± 0.01
Liver
Moisture	59.56 ± 1.52	59.62 ± 1.35	59.72 ± 0.57	59.44 ± 0.35	59.89 ± 0.78	59.58 ± 1.35
Protein	17.33 ± 0.63	17.40 ± 0.62	17.35 ± 0.56	17.12 ± 0.69	17.45 ± 0.95	17.35 ± 0.79
Lipid	10.78 ± 0.67	10.56 ± 0.57	10.46 ± 1.79	9.89 ± 0.65	9.66 ± 0.47	8.99 ± 0.85
Ash	1.85 ± 0.46	1.97 ± 0.47	1.89 ± 0.55	2.23 ± 0.78	1.85 ± 1.25	2.34 ± 0.58
Phosphorus	0.58 ± 0.03	0.54 ± 0.02	0.54 ± 0.02	0.54 ± 0.01	0.56 ± 0.03	0.58 ± 0.01
Viscera
Moisture	48.24 ± 0.68	48.46 ± 1.14	48.57 ± 0.68	48.22 ± 0.67	48.12 ± 0.56	48.35 ± 0.48
Protein^1	19.35 ± 0.57	19.75 ± 0.55	19.23 ± 0.68	19.45 ± 0.45	19.78 ± 0.87	19.57 ± 0.79
Lipid	4.67 ± 0.69	4.59 ± 0.95	4.46 ± 1.33	3.96 ± 1.33	3.75 ± 0.58	3.58 ± 0.85
Ash	1.36 ± 0.58	1.38 ± 0.76	1.45 ± 0.47	1.26 ± 0.54	1.35 ± 0.25	1.17 ± 0.95
Phosphorus^1	0.58 ± 0.02	0.57 ± 0.01	0.57 ± 0.01	0.55 ± 0.02	0.58 ± 0.01	0.57 ± 0.03
Biological analysis
CF^2	3.65 ± 0.04	3.72 ± 0.13	3.73 ± 0.02	3.65 ± 0.03	3.72 ± 0.12	3.65 ± 0.14
VSI^3	8.83 ± 0.45^d	8.24 ± 0.47^c	7.65 ± 0.95^b	7.73 ± 0.13^ab	6.98 ± 0.35^ab	5.98 ± 0.85^a
HSI^4	3.96 ± 0.11^d	3.41 ± 0.22^cd	2.95 ± 0.13^c	2.31 ± 0.32^b	1.62 ± 0.53^b	0.92 ± 0.02^a
MFI^5	5.93 ± 0.12^d	5.71 ± 0.13^cd	5.10 ± 0.76^c	4.63 ± 0.67^b	4.51 ± 0.93^ab	4.01 ± 0.43^a

Mean ± SE, n = 3 and each n consists of 18 fish per replicate. Values of the similar row having different superscripts are found significant (P < 0.05). Initial body proximate composition was: moisture 72.63%, protein 18.66%, lipid 6.38%, ash 1.57%, phosphorus 0.45% and energy 8.95 kJ g⁻¹, and total lipid contents of muscle, liver and viscera were 0.65%, 7.43% and 12.92%, respectively. The condition indices of initial fish were VSI 5.73%, HSI 0.54%, and MFI 1.34%.

¹ Nitrogen or phosphorus retention = 100× [(N and P in final fish × biomass final fish) − (N and P in initial fish × biomass in initial fish)]/(N and P in diet × feed intake);

² CF (Condition factor, K) = 100 × (weight / length³);

³ Viscerosomatic index, VSI = 100 × [wet weight of visceral organs and associated fat tissue (g) / wet body weight (g)];

⁴ Hepatosomatic index, HSI = wet liver weight (g) / empty fish weight (g) × 100;

⁵ Mesenteric fat index, MFI = 100 × [mesenteric fat weight (g) / wet body weight (g)].

Table 4. Enzymatic activity of juvenile mrigal fed different dietary phytase levels for 90 days.

	Dietary Phytase level (FTU /kg)
Item	D1-0	D2-250	D3-500	D4-750	D5-1000	D6-1250
SOD (U/mg protein)	7.79 ± 0.63	7.86 ± 1.24	7.88 ± 0.36	7.89 ± 0.59	7.88 ± 1.0	7.88 ± 0.55
CAT (U/mg protein)	30.18 ± 1.36	28.30 ± 1.26	30.26 ± 1.42	30.59 ± 1.19	31.76 ± 1.25	31.75 ± 1.20
GPx (U/mg protein)	72.52 ± 1.33	74.91 ± 1.20	75.36 ± 1.37	75.39 ± 1.18	76.28 ± 1.39	76.27 ± 1.14

Figure 1. Optimum dietary phytase level (FTU /kg) on weight gain (WG, g) of fish juveniles.

Figure 2. Nitrogen (N) and phosphorus (P) retention (%) in juvenile mrigal fed graded dietary phytase levels for 90 days.