Product Test Report
Comparison Test of Cow Feeding Effect on Antimicrobial Peptide Products
2014.11.10
1. Objective:
1) To determine the practical feed effects of Antibacterial Peptide product from Bornsun and M product from Xinbang company;
2) To compare the practical effects of Antibacterial Peptide and M;
3) To determine the research direction of microecological products in our company.
Main constituents of M: yeast culture 600 g/kg, fungal enzyme 200 g/kg, wall-broken yeast 150 g/kg, probiotics (mainly contain Bacillus subtilis+ Saccharomyces cerevisiae, 200 million/g)
2.1 Materials
Antibacterial Peptide(Bornsun) and M(Xinbang)
2.2 Time and place
Time: 2013.7.1-2013.9.1, preparation period was 15 days, trial period was 45 days, total experimental period was 60 days.
Place: Maheli pasture(Huhehaote city)
2.3 Animals
120 healthy cows (similar in body condition, lactation time and birth) were randomly divided into three groups: group 1, control group
and group 3, each group contained 40 cows.
2.4 Diets
These 3 groups were fed same basal diets with same dosage and method, additives are as follows:
Treatment Group 1(southeast cowhouse): basal diets with M 30g/cow a day
Control group (northeast cowhouse): basal diets
Treatment Group 3(southwest cowhouse): basal diets with Antibacterial Peptide 15g/cow a day.
The experimental diets were manufactured by subsidiary company in Huhehaote City: each group needed at least 36 tons
experimental diets, total number of diets were 108 tons.
2.5 Experimental design
Table 1 Sampling details
Items |
Sampling frequency |
Sampling date |
Assayed indexes |
Sampling time point of everyday |
Blood |
4 |
on Day 1st, 15th, 40th and 60th |
serum calcium, serum phosphate, serum IgM, serum IgG |
sampling in the morning without eating |
Milk |
4 |
on day 1st, 15th, 40th and 60th |
milk production, milk protein, milk fat, somatic cells in milk |
sampling in the morning when milking |
Ruminal fluid |
4 |
on day 1st, 15th, 40th and 60th |
ruminal pH value |
at least limosis for 2 hours |
3. Milk indexes analysis
3.1 Effects of Antibacterial Peptide and M on milk production
Table 2 Effect of Antibacterial Peptide and M on milk production(kg/day)
Groups |
Day 1st |
Day 15th |
Day 40th |
Day 60th |
Mean value |
Group 1 |
22.03 |
22.09b |
23.11c |
23.32c |
22.63b |
Control Group |
22.21 |
21.16b |
19.46b |
18.39b |
20.05b |
Group 3 |
23.26 |
25.81a |
27.08a |
26.65a |
25.70a |
P>F |
0.9332 |
0.0436 |
0.0001 |
0.0001 |
0.0001 |
Effects of Antibacterial Peptide and M products on milk production were showed in table 2: in the whole experimental period,
compared with control group, milk production of last three sampling time in group 3 are significantly higher (P<0.01). From each
sampling time point, we could know that milk production of last three sampling time in control group were less than the other two
groups, meanwhile, milk production of group 3 were significantly increased compared with group 1 in last three sampling time (P<0.05),
while milk production of group 1 were significantly increased compared with control group in last two sampling time (P<0.05).
3.2 Effects of Antibacterial Peptide and M products on milk protein
Table 3 Effect of Antibacterial Peptide and M on milk protein(%)
Groups |
Day 1st |
Day 15th |
Day 40th |
Day 60th |
Group 1 |
3.56 |
3.56 |
3.57 |
3.57 |
Control Group |
3.55 |
3.56 |
3.57 |
3.57 |
Group 3 |
3.56 |
3.57 |
3.57 |
3.58 |
P>F |
0.9946 |
0.986 |
0.993 |
0.977 |
Milk protein content of each group were showed in table 3: from the overall data, we could know that, milk protein content of each
group in different time point had no significant difference (P>0.05), all of these data were stably between 3.55% and 3.58%.
Milk protein of four sampling time changed little, however, the values of fourth time were higher than other 3 sampling time in each
group, among which value of group 3 was the highest (3.58%). Possible reason of this result was due to the proximity of October,
so that the milk protein content values became high, moreover, cows in experimental pasture were imported ones which had high
milk protein rates, thus, antibacterial peptide products only have the tendency to improve milk protein content.
3.3 Effects of Antibacterial Peptide and M on milk butter-fat rate
Table 4 Effect of Antibacterial Peptide and M on milk butter-fat rate
Groups |
Day 1st |
Day 15th |
Day 40th |
Day 60th |
Group 1 |
4.53 |
4.62 |
4.64 |
4.64 |
Control Group |
4.56 |
4.61 |
4.63 |
4.62 |
Group 3 |
4.57 |
4.67 |
4.73 |
4.76 |
P>F |
0.827 |
0.949 |
0.851 |
0.563 |
Milk butter-fat rate of each group were showed in table 4, milk butter-fat rate of each group had no significant difference (P>0.05),
all of them were between 4.53% and 4.76%, meanwhile, milk butter-fat rate of last sampling time was higher than the other 3 ones,
while milk butter-fat rate of group 3 was the highest (4.76%), and milk butter-fat rate gradually increased with time (P>0.05).
3.4 Effects of Antibacterial Peptide and M on somatic cell counts(SCCs)
Table 5 Effect of Antibacterial Peptide and M on SCCs(104/mL)
Groups |
Day 1st |
Day 15th |
Day 40th |
Day 60th |
Group 1 |
40 |
46 |
52ab |
68a |
Control Group |
42 |
54 |
67a |
82a |
Group 3 |
41 |
37 |
35b |
38b |
P>F |
0.559 |
0.074 |
0.030 |
0.013 |
SCCs of each group were showed in table 5, from which we could know that, SCCs of each group had no significant difference
(P>0.05) except third and fourth sampling time (P<0.05). SCCs in control group were higher than that in the other two groups,
SCCs of group 1 showed decreasing tendency and had no obvious difference with control (P>0.05) ; from data in control group,
we know that Fourth sampling result was the highest (82) and these data showed increasing tendency within time.
3.5 Effect of Antibacterial Peptide and M on urea nitrogen in milk
Table 6 Effect of Antibacterial Peptide and M on urea nitrogen in milk(mg/DL)
Groups |
Day 1st |
Day 15th |
Day 40th |
Group 1 |
13.1 |
13.59 |
13.07 |
Control Group |
13.5 |
13.18 |
13.21 |
Group 3 |
15.8 |
14.63 |
14.18 |
P>F |
0.599 |
0.329 |
0.453 |
Results of table 6 showed: milk urea nitrogen level of group 3 was the highest, next was group 1 and control group was the lowest.
Normal value of milk urea nitrogen level is 13 to 17, our data showed that milk urea nitrogen in the three groups were normal condition,
which indicated that cow’s intake and digestion of energy and protein were balanced and within the normal range.
3.6 Effects of Antibacterial Peptide and M on dry matter(DM)
Table 7 Effect of Antibacterial Peptide and M on dry matter(%)
Groups |
Day 1st |
Day 15th |
Day 40th |
Day 60th |
Group 1 |
14.28 |
14.71ab |
14.60b |
14.40b |
Control Group |
14.35 |
14.34b |
14.30b |
14.58b |
Group 3 |
14.96 |
15.45a |
15.49a |
15.67a |
P>F |
0.272 |
0.084 |
0.008 |
0.0004 |
Effects of Antibacterial Peptide and M on DM were showed in table 7: DM of first sampling time in three groups had no significant
difference (P>0.05); as for the second sampling results, DM in group 3 was significantly decreased compared with control group
(P<0.05); compared with group 1 and control group, the results of group 3 were significantly increased in 40th and 60th day (P<0.01).
From the whole experiment period, we could find that, DM in group 3 were significantly higher than the other two groups.
3.7 Effects of Antibacterial Peptide and M on lactose
Table 8 Effect of Antibacterial Peptide and M on lactose(%)
Groups |
Day 1st |
Day 15th |
Day 40th |
Day 60th |
Group 1 |
5.09 |
5.11 |
5.15 |
5.18 |
Control Group |
5.07 |
5.13 |
5.14 |
5.15 |
Group 3 |
5.05 |
5.15 |
5.15 |
5.16 |
P>F |
0.923 |
0.887 |
0.932 |
0.980 |
Effects of Antibacterial Peptide and M on lactose were showed in table 8: there was no significant difference of lactose content in
3 groups. Lactose contents were within 5.05 to 5.18 and lactose content of control group was less than it in group 1 and 3. With
the extension of trial period, lactose contents in each group were increased.
4. Analysis of blood parameters
4.1 Effects of Antibacterial Peptide and M on serum IgM(mg/mL)
Table 9 Effect of Antibacterial Peptide and M on serum IgM(mg/mL)
Groups |
Day 1st |
Day 15th |
Day 40th |
Day 60th |
Mean Value |
Group 1 |
12.15 |
13.49 |
13.81 |
14.15 |
13.40 |
Control Group |
12.80 |
11.12 |
10.45 |
11.79 |
11.54 |
Group 3 |
12.43 |
14.43 |
15.80 |
16.48 |
14.78 |
Effects of Antibacterial Peptide and M on IgM were showed in table 9: in early stage of the experiment, IgM of each group changed
little, about 12mg/ml, with the extension of the test period, IgM value in each group rised in different degree, the 3rd and 4th detecting
results of group 3 were 15.80 mg/ml and 16.48 mg/ml respectively, higher than the other two groups. The mean value of serum IgM
in group 3 was 14.78 mg/ml, which was a little higher than the other two test groups.
4.2 Effects of Antibacterial Peptide and M on serum IgG(mg/mL)
Table 10 Effects of Antibacterial Peptide and M on serum IgG(mg/mL)
Groups |
Day 1st |
Day 15th |
Day 40th |
Day 60th |
Mean Value |
Group 1 |
46.95 |
50.85 |
51.01a |
51.30a |
50.03a |
Control Group |
47.64 |
44.67 |
46.03a |
44.72a |
45.77a |
Group 3 |
46.95 |
53.68 |
56.67b |
59.02b |
54.08b |
Effects of Antibacterial Peptide and M on IgG were showed in table 10: in early stage of the experiment, IgG of each group changed
little, about 47 mg/ml, with the extension of the test period, IgG value in each group rised in different degree, the 3rd and 4th sampling
results of group 3 were 56.67 mg/ml and 59.02 mg/ml respectively, which were significantly higher than the other two groups at same
sampling time. The mean value of serum IgG content in group 3 was 54.08 mg/ml, which also was significantly higher than control
group and group 1 (P<0.05).
4.3 Effects of Antibacterial Peptide and M on serum calcium (Ca) (mmol/L)
Table 11 Effects of Antibacterial Peptide and M on serum Ca (mmol/l)
Groups |
Day 1st |
Day 15th |
Day 40th |
Day 60th |
Mean Value |
Group 1 |
2.29 |
2.32 |
2.34 |
2.34 |
2.32 |
Control Group |
2.30 |
2.33 |
2.37 |
2.48 |
2.39 |
Group 3 |
2.29 |
2.35 |
2.34 |
2.37 |
2.35 |
Effects of Antibacterial Peptide and M on Ca were showed in table 11: Antibacterial Peptide and M affected Ca concentration little
throughout the test period, there were no significant effects of Antibacterial Peptide and M on Ca concentration, and all of them were
within the nomal limits: 2.2 to 2.6 mmol/L.
4.4 Effects of Antibacterial Peptide and M on serum phosphorus (mmol/L)
Table 12 Effects of Antibacterial Peptide and M on serum phosphorus (mmol/l)
Groups |
Day 1st |
Day 15th |
Day 40th |
Day 60th |
Mean Value |
Group 1 |
2.48 |
2.49 |
2.48 |
2.48 |
2.48 |
Control Group |
2.47 |
2.45 |
2.45 |
2.46 |
2.46 |
Group 3 |
2.43 |
2.43 |
2.47 |
2.48 |
2.45 |
Effects of Antibacterial Peptide and M on serum phosphorus were showed in table 12: there were no significant effects of
Antibacterial Peptide and M on phosphorus concentration, all of them are within the normal limits: 1.4 to 2.48 mmol/L.
5. Index analysis of rumen liquid
5.1 Effects of Antibacterial Peptide and M on pH value
Table 13 Effects of Antibacterial Peptide and M on pH value
Groups |
Day 1st |
Day 15th |
Day 40th |
Day 60th |
Mean Value |
Group 1 |
6.13 |
5.94 |
6.08 |
6.19 |
6.09 |
Control Group |
5.80 |
5.94 |
6.24 |
6.43 |
6.10 |
Group 3 |
5.95 |
5.87 |
5.94 |
6.07 |
5.96 |
Effects of Antibacterial Peptide and M on pH value were showed in table 13: mean value of pH changed little and were within the
normal limits during the whole test period. Rumen pH of group 1 and 3 also changed little, pH of control group changed more, from
PH 5.80 to 6.43, all of these data were within normal limits: From PH5.5 to PH 6.8.
5.2 Effects of Antibacterial Peptide and M on DGGE
Fig.1 DDGE picture of August 15th
Fig.2 DDGE picture of August 31st
Fig.3 DDGE picture of September 15th
Fig.4 DDGE picture of September 28th
Description:
Fig 1: Banding “0” means blank control group; from banding “1” to “5” means treatment group 3, from banding “6” to “11” means
treatment group 1, banding “12” to “16” means control group Fig 2 to 4: Banding “0” means blank control group, from banding
“1” to “6” means treatment group 3, “7” to “12” means treatment group 1, “13” to “18” means control group. More bands means more
microfloras, but the specific diversity index were derived from the software analysis.
Table 14 Effects of Antibacterial Peptide and M on DDGE(mmol/l)
Groups |
Day 1st |
Day 15th |
Day 40th |
Day 60th |
Mean Value |
Group 1 |
1.18 |
1.11 |
1.23 |
1.17 |
1.17 |
Control Group |
1.14 |
1.08 |
1.17 |
1.15 |
1.13 |
Group 3 |
1.18 |
1.06 |
1.14 |
1.17 |
1.14 |
Microbial diversity index was derived by specific software on the basis of DDGE results, the bigger of the diversity index means the
richer microbial diversity, which indicating that the more stability of microbial flora in testing animals.
From table 14 we could know that, DGGE value of three group in different time point had no significant difference, group 1 had a
slightly higher DGGE content, group 3 was in second place while value of control group was lowest.
6. Economic and efficiency analysis
Table 15 Economic and efficiency analysis of each group during whole trial period
Groups |
Dosage |
Price |
Cost |
Icreased milk production |
Mean milk price |
Benefit |
|
g/cow.day |
RMB/kg |
RMB/cow.day |
Kg/day |
RMB/kg |
RMB |
Group 1 |
30 |
30 |
0.90 |
2.58 |
4 |
9.42 |
Control Group |
- |
- |
- |
- |
- |
- |
Group 3 |
15 |
200 |
3 |
4.16 |
4 |
13.64 |
From table 15 we could know that, dosage of different products in experimental group were different, costs were 0.9 and 3 RMB/cow
per day in group 1 and 3, respectively, while their economic benefits were 9.42 and 13.64 RMB, thus we know that, economic benefit
of Antibacterial Peptide group was 4.22 RMB/cow per day higher compared with group 1.
7. Discussions
The experimental period was 60 days from August 2nd to October 1st, using total mixed ration method feeding cows. Group 3 had
added 1 kg/t Antibacterial Peptide, which could effectively resist potential mold or other harmful bacterium, reduce effect of heat stress
and mycotoxins, improve healthy condition of cows and ensure milk production of them.
Group 3 was the best effective group for milk production, cow’s body condition and feed intake according to the charging man of pasture.
The second better effective group for milk production one was group. It suggested that the effect of Antibacterial Peptide on milk production
was significant.
7.1 Discussion of milk indexes results
From results of milk indexes we could know that, in the whole experimental period, milk production of control group and group 1 were
significantly lower than group 3, urea content of control group was less than the other two groups, while somatic cell counts in control
group were significantly higher than group 3. Milk protein, fat and lactose content of control group had no markedly difference compared
with the other two groups. The possible reasons may be that cows in Maheli pasture were imported from New Zealand which had a
relatively high milk composition, milk protein, fat and lactose value than China Holstein cows, so the microecologics feed additive had
no significant effects on milk composition. However, microecologics products could regulate cow's ruminal microbial environment,
restore normal type and amount of ruminal microbes rapidly, supplement decomposition of cellulose, hemicellulose and other plant
nutrients, thereby regulating ruminal environment better, improving production performance on the premise of ensuring the health of cows,
increasing milk production. DM content of milk in group 3 was higher than the control group (P<0.05), and the somatic cell counts were
significantly decreased compared with control group(P<0.05).
7.2 Discussion of serum indexes results
Results of serum calcium and serum phosphorus levels showed that, each test group had no significant effects on serum calcium
and phosphorus. The serum Ca and phosphorus levels are within the normal limits, thus, we could conclude that cows are in healthy
condition.
When B cells stimulated by antigen, it will proliferate and differentiate into plasmocyte which could generate one kind of glycoprotein
named specific immunoglobulins. The Specific immunoglobulins are mainly exist in serum and other body fluid, and its immunologic
function showed through specificity combination with corresponding antigens and playing an important role in protecting body from
bacteria, viruses and toxins infection. Serum IgG, IgM levels reflect the state of humoral immunity, the IgG is the most abundant
immunoglobulin in serum, it is a primary antibody generated by animal and human after they receive positive immunization. The IgG
content directly reflects animal immunity levels. From data in table 9 and 10, we know that serum IgG content in group 3 were
significantly higher than it in group 1 and control group, while IgM content was a little higher compared with the other two groups,
which indicated that Antibacterial Peptide could improve immunologic function, IgG value of control group were relatively lower,
so immunity of cows in control group were decreased, thus affecting milk production and body health condition.
7.3 Discussion of ruminal fluid results
Normal pH value of rumen fluid is between 5.5 and 6.8, best pH value is between 6.0 and 6.3, ciliate and protozoa can grow rapidly
in the ideal pH value 6.0, thus produce large amount of volatile fatty acid(VFA), VFA are the main source of energy required by cows.
From the whole experimental period, we know that pH value of group 1 and 3 were in the ideal range, only pH value of control group
was higher than 6.3, but it also in the normal range. We concluded that microecologics could regulate ruminal fluid pH value in a normal
range.
8. Conclusion
There is no significant difference Milk fat, protein, lactose content, serum Ca, serum phosphorus and ruminal pH value for each groups.
Compared with control group, milk production in treatment group 1 and 3 were increased significantly. Meanwhile, Antibacterial Peptide
ould significantly raise the DM content and reduce somatic cell counts, it indicate that Antibacterial Peptide could able to increase
serum IgG and IgM concentration levels, improving cow’s healthy condition, maintaining stable ruminal pH value, and finally it can
create more economic benefits. In conclusion, effects of group 3 were better than group 1, which means Antibacterial Peptide
could provide more economic benefits.