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Amelioration of Oxidative Stress Through Supplementation of Self-formulated

Anti-oxidant Mixture for Early Recovery and Prophylaxis of Bovine Mastitis in
Kashmir Valley

Article in Indian Journal of Animal Research · April 2021

DOI: 10.18805/IJAR.B-4339

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 B-4339
[1-8]

RESEARCH ARTICLE Indian Journal of Animal Research

Amelioration of Oxidative Stress Through Supplementation of

Self- formulated Anti-oxidant Mixture for Early Recovery and
Prophylaxis of Bovine Mastitis in Kashmir Valley
A.Muhee, H.U. Malik, R.A. Bhat, Z.A. Akhoon, Adil Mehraj Khan 1,
S.U. Nabi, S.A. Hussain, S.A. Beigh, M. Shaheen 10.18805/IJAR.B-4339

ABSTRACT
Background: During lactation, mammary epithelial cells exhibit a high metabolic rate and thus produce large amounts of ROS and
lipid peroxides in vivo (Jin et al. 2014; Ganguly et al., 2016). A surplus of ROS and the absence of optimal amounts of antioxidants
(which neutralize these free radicals or ROS) results in oxidative stress (Lykkesfeldt and Svendsen, 2007). A clinical study was
undertaken on Bovine Mastitis in Kashmir valley to study the relation between oxidative stress and clinical mastitis. An attempt was
also made to see the effect of supplementation of self-formulated anti-oxidant trace mineral mixture on recovery and prophylaxis of
Bovine mastitis through amelioration of oxidative stress.
Methods: The oxidative stress was assessed through estimation of superoxide dismutase (SOD), catalase, reduced glutathione (GSH),
malondialdehyde (MDA) and nitric oxide (NO). In addition, blood trace mineral status for copper (Cu), zinc (Zn), manganese (Mn) and
selenium (Se) were also assessed in mastitic animals and compared with normal healthy lactating animals. The utility of anti-oxidants
in clinical management of mastitis was measured through response to treatment with trace minerals like Cu, Zn, Mn and Se in addition
to conventional antibiotic therapy. Two groups of mastitic animals received two therapeutic regimens. Group I animals received
antibiotics and self formulated anti-oxidant mixture at therapeutic doses while as Group II animals received only antibiotics (at same
dose rate and frequency). Clinical recovery was assessed on the basis of CMT point score, milk somatic cell count and milk biochemistry.
For prophylactic study, forty recently parturated lactating animals having susceptibility to occurrence of mastitis were divided into two
groups of twenty animals each. One group of animals was supplemented with self-formulated anti-oxidant mixture at prophylactic
doses for a period of thirty days so as to see the effect of supplementation on oxidative stress parameters and occurrence of clinical mastitis.
Result: A significant decrease was found in the values of SOD, catalase, GSH and Cu, Zn, Mn and Se but a significant increase was
found in the values of MDA and NO in clinical cases of mastitis as compared to healthy control group. Therapeutic regimen I proved
efficacious than the therapeutic regimen II in treatment of clinical mastitis with higher recovery rates and lesser number of mean days
required for recovery in group I than group II animals. The efficacy of prophylactic treatment was monitored by occurrence of mastitis
during the course of therapy and one month after therapy. Group I animals did not suffer from clinical mastitis and showed
considerable improvement in oxidative stress parameters, milk SCC and blood trace mineral status as compared to group II.

Key words: Antioxidants, Bovine mastitis, Catalase, GSH, MDA, NO, Oxidative stress, SOD, Trace minerals.

INTRODUCTION Division of Clinical Veterinary Medicine, Ethics and Jurisprudence,

Reactive oxygen species (ROS) are natural end products Faculty of Veterinary Sciences and Animal Husbandry, Shuhama,
of the intensive cellular metabolism. W hen the disturbance SKUAST-Kashmir, Shrinagar-190 025, Jammu and Kashmir, India.
of homeostasis occurs, oxidative processes lead to oxidative 1
Divis ion of Veterinary Pharmacology, Fac ulty of Veterinary
stress causing inflammation of the mammary gland (mastitis) Sciences and Animal Husbandry, Shuhama, SKUAST-Kashmir,
in high-yielding dairy cows. Mastitis is characterized by a Shrinagar-190 025, Jammu and Kashmir, India.
range of physical and chemical changes of the milk and Corresponding Author: Z.A. Akhoon, Faculty of Veterinary
pathological changes in the udder tissues (Constable et al, Sciences and Animal Husbandry, Shuhama, SKUAST-Kashmir,
2017). The importance of mastitis is emphasised by the fact Shrinagar-190 025, Jammu and Kashmir, India.
that it produces huge losses in cattle industry (Salman, 2009; Email: drzubair7866@gmail.com
Maćešić et al., 2016).The overall prevalence of bovine How to cite this article: Muhee, A., Malik, H.U., Bhat, R.A., Akhoon,
mastitis has been recorded to be 14.86 % in Kashmir valley Z.A., Mehraj Khan, A., Nabi, S.U., Hussain, S.A., Beigh, S.A. and
which highlights the economic importance of the disease Shaheen, M. (2021). Amelioration of Oxidative Stress Through
(Ifat et al.2017). Since mastitis is caused by different Supplementation of Self-formulated Anti-oxidant Mixture for Early
microorganisms, the prevention, treatment and control of Recovery and Prophylaxis of Bovine Mastitis in Kashmir Valley.
the disease is hardly feasible (Russell, 2011; Deb et al., Indian Journal of Animal Research. DOI: 10.18805/IJAR.B-4339.
2013). So far, conventional antibiotic therapy is the only Submitted: 22-10-2020 Accepted: 23-03-2021 Online: 28-04-2021
proven method for the prevention and control of mastitis,

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 Amelioration of Oxidative Stress Through Supplementation of Self-formulated Anti-oxidant Mixture for Early Recovery....

but several problems arise from the use of antibiotics like of oxidative stress biomarkers and anti-oxidants more
developing resistance to antibiotic, questionable drug accurately indicates the oxidative status of the animals
efficacy and presence of antibiotic residues in the milk (Cigliano et al., 2014, Costantini and Verhulst, 2009).
(Petrovski and Eats, 2014). These facts highlight the need Supplementation of antioxidant trace elements stabilize the
for completely newer moieties for treatment of mastitis. highly reactive free radicals generated as a result of oxidative
During lactation, mammary epithelial cells exhibit a high stress during mastitis, reduce the inflammatory response
metabolic rate and thus produce large amounts of ROS and and maintain the structural and functional integrity of cells
lipid peroxides in vivo (Jin et al. 2014; Ganguly et al., 2016). (Kushwaha and Mohan, 2019). The present study was
A surplus of ROS and the absence of optimal amounts of therefore undertaken to evaluate the possible oxidative
antioxidants (which neutralize these free radicals or ROS) stress and alterations in anti-oxidant and trace mineral
results in oxidative stress (Lykkesfeldt and Svendsen, 2007). profile during clinical mastitis and to investigate the possible
Oxidative stress in turn affects the cellular metabolism role of antioxidants in treatment and prophylaxis of this major
leading to deteriorated health in animals (Celi, 2011). It is economic disease of dairy animals.
the primary factor that leads to immune dysfunction and
impairs the inflammatory response which in turn is reflected MATERIALS AND METHODS
particularly as the inflammation of the udder (Abuelo et al., A total of 36 multiparous cows in the age group of 4-8 years
2013). The suppression of immune resistance due to with BCS 3.0 were included in the therapeutic study. The
oxidative stress can lead to the establishment of different study was undertaken on clinical cases of bovine mastitis
microbial pathogens in mammary gland leading to mastitis presented to Teaching Veterinary Clinical Complex (TVCC),
(Mir et al., 2017)). Mastitis could induce the increase of free Faculty of Veterinary Sciences and Animal Husbandry
radicals formation in milk and leading to oxidative stress (F.V.Sc and A.H), Shere Kashmir University of Agricultural
(Gu et al., 2009). Both CM and SCM are associated with Sciences and Technology of Kashmir (SKUAST-K) and local
release of free radicals, increased total oxidant capacity and animal husbandry dispensaries in Ganderbal, Manasbal,
decreased total antioxidants capacity in milk (Atakisi et al., Shuhama, Gulab bagh and Shalimar areas for treatment .
2010; Patnaik et al., 2014). It has been reported that significant 24 cases of clinical mastitis were studied and all the animals
decrease in blood superoxide dismutase (SOD) and catalase included were clinical cases. 12 apparently healthy lactating
activities, reduced glutathione (GSH) concentration and an animals served as control group. The cases of clinical
increase in erythrocytic lipid peroxides was observed in cows mastitis were diagnosed on the basis of clinical examination,
with clinical mastitis (Jhambh et al., 2013). case history and clinical findings. The severity of the cases
Trace minerals are critical for proper immune response was assessed on the basis of CMT score and SCC of milk
and play an important role in udder health (Ganda et al., samples. Cases with CMT score of 2 or 3 with somatic cell
2016). Trace minerals with an antioxidant function include counts more than 4×105 /mL of milk were considered positive
selenium (Se), copper (Cu), zinc (Zn), manganese (Mn) and for mastitis. For therapeutic trial, the clinical cases of mastitis
iron (Fe). W hile some nutrients have a role in directly were divided into two groups of twelve animals each. Group
quenching free radicals, these trace minerals have an I animals were given self formulated anti-oxidant trace
indirect role in which they are required as components of a mineral mixture at therapeutic doses as recommended by
variety of antioxidant enzymes (Marta Wolonciej et al; 2016). NRC, 2001 whose formulation is given in Appendix-I. All the
For example, enzymatic antioxidants such as superoxide four ingredients were mixed making a total weight of about
dismutase (SOD), glutathione peroxidase (GSH-Px) and 5.005 gms and stored in air tight zip lock polyethene
catalase are considered to be an important defense system pouches. Total 84 pouches were made and 7 pouches were
against free radical accumulation. The determination of given to individual animal owners for providing contents of
products of peroxidative damage to macromolecules and one pouch to the animal once daily for 7 days. Group II
antioxidant substances like reduced glutathione and animals were not given any anti-oxidant trace mineral
enzymes (e.g. SOD, and Catalase) are useful markers for mixture. Both the groups of animals were given antibiotic
the oxidative stress and antioxidant status respectively enrofloxacin at the dose rate of 6 mg/kg per animal twice
(Sharma et al. 2011b). Membrane phospholipids are prone daily for 5 days. The choice of the antibiotic was based on
to oxidation, and malondialdehyde (MDA) is generated as a antibiotic sensitivity test of milk which was performed in each
consequence of lipid peroxidation, and as such is assayed case. Animals which were not found sensitive to enrofloxacin
as a biomarker of oxidative stress (Abuelo et al. 2014). Nitric were excluded from the study.
oxide (NO) is one of the most important reactive nitrogen For prophylactic studies forty recently parturated
intermediates produced in a significant amount by epithelial lactating animals maintained at Mountain Livestock
cells and macrophages of mammary gland during mastitis Research Institute (MLRI), SKUAST-K, Manasbal under
(Bouchard et al. 1999). This excessive release of NO results similar managemental conditions but having susceptibility
in oxidative damage to mammary gland and thus estimation to occurrence of mastitis (as evidenced by precedence to
of NO is also assayed as a biomarker of oxidative stress occurrence of mastitis in previous lactations) were studied.
(Komine et al. 2004; Atakisi et al. 2010). So, joint evaluation One group of animals (Group I) comprising of twenty animals

2 Indian Journal of Animal Research

 Amelioration of Oxidative Stress Through Supplementation of Self- formulated Anti-oxidant Mixture for Early Recovery....

Appendix- I: Therapeutic dose of mineral mixture. digested sample was diluted to 10 mL using double glass
Copper sulphate : 1 gm distilled water for mineral estimation. The samples were
Zinc sulphate : 2 gm
analysed at Kashmir University, Hazratbal in the deparment
of University Scientific Instrumentation Centre (USIC) using
Manganese sulphate : 2 gm
Polarised Zeeman Atomic Absorption Spectrophotometer (Z-
Selenium : 5 mg
2300, Hitachi).
Total dose : 5.005g/animal/day for 7 days
Evaluation of treatment efficacy
Appendix- II: Prophylactic dose of mineral mixture.
The efficacy of therapeutic regimes in clinical cases of
Copper sulphate : 500 mg mastitis was evaluated on the basis of clinical recovery,
Zinc sulphate : 500 mg mean days required for recovery, CMT score, milk SCC,
Manganese sulphate : 400 mg blood oxidative stress parameters on day 0, day 5 and day
Selenium : 2 mg 10 of treatment and trace mineral status on day 0 and day
Total dose : 1.042 g/animal/day for 30 days

received self-formulated anti-oxidant trace mineral mixture

for a period of thirty days while as the other group (Group II)
did not receive any anti-oxidants. The anti-oxidant trace
minerals were given at prophylactic doses (Appendix-II) as
recommended by NRC, 2001. The total dose per animal
was 1.402 grams. Six hundred pouches were made and
one pouch per animal per day was given to selected twenty
animals (Group I) for 30 days.
Blood sampling
Blood samples were collected from healthy lactating animals
to obtain baseline values of oxidative stress parameters and
trace minerals. Approximately 10 mL of blood was collected Fig 1: Std. Curve of Catalase
in mineral free heparinised vials under aseptic conditions
from the jugular vein of healthy as well as mastitic cows.
Blood was immediately centrifuged at 3000 rpm for 20
minutes to separate plasma. 3 mL of plasma was stored at
-80o C for estimation of oxidative stress parameters (MDA,
GSH and NO) and trace minerals. The RBC’s were washed
thrice with phosphate buffer saline in the ratio of 1:1 on v/v
basis and centrifuged at 2000 rpm for 10 minutes and 1 %
erythrocyte lysate was prepared in normal saline (100µL
washed RBC+9.9 mL normal saline) for estimation of rest
of oxidative stress parameters (SOD and catalase). MDA,
SOD, catalase and GSH were estimated using commercial
kits supplied by Sigma Aldrich (USA) whereas nitric oxide
was quantified using modified Greiss reaction as described
Fig 2: Std. Curve for GSH.
by Miranda et al. (2001). To determine superoxide dismutase
activity in erythrocytes, the lysate was diluted 50-fold with
0.01 mmol/L phosphate buffer, pH 7.0, and then SOD was
measured by a kinetic method using a commercial kit by
Sigma Aldrich (USA) and results were expressed in
international units per milliliter. For estimation of catalase,
1% hemolysate was used. The procedures for estimation of
catalase, reduced glutathione, malondialdehyde and nitric
oxide were first standardised to obtain standard curves and
regression equations after which the values of the test
samples were estimated using regression equation. The
standard curves for catalase, reduced glutathione,
malondialdehyde and nitric oxide are given in Fig 1, 2, 3
and 4 respectively. The plasma samples were digested using
distilled concentrated nitric acid (AR, 15 mL) and each Fig 3: Std. Curve for MDA.

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 Amelioration of Oxidative Stress Through Supplementation of Self- formulated Anti-oxidant Mixture for Early Recovery....

Prophylactic study
Group I animals receiving anti-oxidant minerals showed a
considerable improvement in the oxidative stress
parameters, trace mineral status and milk SCC as compared
to group II on day 15 and day 30 post treatment. The animals
in the group II showed consistently increasing oxidative
stress parameters, lower levels of trace mineral status and
increased milk SCC. Clinical mastitis was not detected in
any of the animals in both the groups during the course of
the therapy and one month after the study. However, the
udder health status of group I animals was found better than
group II as evidenced by significant decrease in milk SCC
Fig 4: Standard curves of NO.
counts 15 days after stopping the treatment (Table 5 and 6).
Our findings of increased oxidative stress markers and
10 of treatment in both the groups of animals. The efficacy decreased anti-oxidants in clinical mastitis are in agreement
of prophylactic treatment was evaluated by occurrence of with Ibrahim et al, 2016 who reported that in dairy cows with
mastitis during the course of therapy and one month after acute clinical mastitis, there is a significant (p<0.05)
therapy, estimation of SCC in milk and blood oxidative stress
decrease in the TAC (total antioxidant capacity), GSH and
parameters on day 0, day 15 and day 30 of treatment in
CAT as well as in the level of zinc and iron but there is a
both the groups of animals. Mineral estimation for copper,
significant (p<0.05) increase in the level of MDA, NO and
zinc, manganese and Selenium was also done on day 0,
the OSI (oxidative stress index).Since the levels of trace
day 15 and day 30 to determine the effect of anti-oxidant
minerals were also found to be decreased in clinical mastitis
trace mineral supplementation on their blood values. After
in the present study, their supplementation resulted in better
fifteen days of stopping the treatment, the udder health status
and earlier recovery rates in mastitic animals. Our findings
of both the groups of animals was assessed through are in agreement with Khushwaha and Mohan,2019 who
estimation of SCC of milk and CMT. reported that deficiency of vitamins and minerals as
Statistical analysis antioxidants particularly vitamin E, vitamin D, selenium, zinc
The data was analyzed using statistical tools (SPSS version and copper leads to increased incidence of mastitis with
20). ANOVA followed by Duncan’s muliple range test was infection of longer duration and more severe clinical signs
and supplementation of antioxidant vitamins and trace
used for multiple comparisons. Paired ‘t’ test was used for
elements stabilizes the highly reactive free radicals
pre and post treatment comparisons within each treatment
generated as a result of oxidative stress, reduces
group. Repeated measures ANOVA was used for pre and
inflammatory response and maintains the structural and
post treatment multiple comparisons. Statistical differences
functional integrity of cells during mastitis. Yang and Li(2015)
were determined at the 5% level of significance.
also reported that supplementation of mastitic dairy cows

RESULTS AND DISCUSSION

A statistically significant decrease was found in the values Table 1: Comparison of blood oxidative stress parameters, blood
of anti-oxidant enzymes like SOD, catalase and GSH and trace mineral prof ile and milk SCC between healthy
a statistically significant increase was found in the values lactatating (control group ) and mastitic animals (Mean±S.E).
of oxidative stress markers like MDA (µmoles/L) and Control group Mastitic group
NO(µmoles/L) in mastitic animals (mastitic group) than Parameters
(N=12) (N=24)
normal healthy animals (control group). A statistically
a
significant decrease was found in the values of Cu, Zn, SOD (U/mL) 151.47±1.69 100.67±1.39b
Mn and Se in mastitic group as compared to control group Catalase (nmoles/min/mL) 36.08±1.16 a
20.24 ±.67b
(Table 1). Therapeutic regimen I for Group I animals proved GSH (nmoles/mL) 59.09±1.57 a
38.66±1.08b
efficacious than the therapeutic regimen II for group II MDA (µmoles/L) 30.37±1.10 a
54.69±1.45b
animals in treatment of clinical mastitis with recovery rates NO (µmoles/L) 33.02±1.29 a
58.48± 1.09b
of 73.33 and 66.66% , respectively in the two groups. Group Copper (µmol/L) 8.25±.07 a
7.01 ± 0.06b
I animals showed early recovery with lesser number of Zinc (µmol/L) 14.47±.31 a
11.71±0.16b
mean days required for recovery (4±0.5 days) than group Mn (µmol/L) 3.78±.11 a
2.94±0.04b
II animals (4.5±0.8 days).The early and efficacious clinical a
Se (ng/mL) 38.47±1.67 31.29±1.14b
recovery in group I animals receiving therapeutic regimen 5 a
SCC (×10 /mL) 2.83±.05 9.94±0.22b
I was also confirmed by the return of oxidative stress
parameters, milk SCC, and trace mineral status to normal Values within a row having superscript (a,b,) with atleast one
or near normal values on day 10 of treatment than group II common alphabet do not differ significantly at 5% level (p<0.05)
animals receiving therapeutic regimen II (Table2, 3 and 4). from each other.

4 Indian Journal of Animal Research

 Amelioration of Oxidative Stress Through Supplementation of Self- formulated Anti-oxidant Mixture for Early Recovery.....

Table 2: Effect of treatment on blood oxidative stress parameters and milk SCC in animals with clinical mastitis (Mean±S.E).
Group I Group II
Parameters Days of treatment
(Antibiotic+Antioxidant trace minerals) (Antibiotic only)
SOD (U/mL) Day 0 100.55±1.47a1 100.81±2.61a1
Day 5 118.00±1.52b1 110.44±2.47b2
Day 10 140.00±2.19c1 116.88±2.12b2
Catalase Day 0 20.69±.98a1 19.67±.89a1
(nmoles/min/mL) Day 5 25.67±1.08b1 22.25±.81b2
Day 10 31.23±1.00c1 23.60±.71b2
GSH Day 0 38.88±1.53a1 38.39±1.55a1
( nmoles/mL) Day 5 49.91±1.60b1 43.29±1.35b2
Day 10 60.57±.98c1 48.25±1.37c2
MDA Day 0 57.73±1.69a1 50.91±2.08a2
(µmoles/L) Day 5 44.14±1.38b1 44.98±2.26ab1
Day 10 30.52±.85c1 40.37±2.23b2
NO Day 0 59.95±1.33a1 56.65±1.72a1
(µmoles/L) Day 5 37.97±1.26b1 46.78±1.66b2
Day 10 30.33±.78c1 40.80±1.19c2
SCC Day 0 9.88±.31a1 10.03±.33a1
(×105/mL) Day 5 6.00±.36b1 7.63±.17b2
Day 10 3.19±.09c1 4.43±.13c2
Values within a row having superscript (1, 2, 3) with atleast one common numerical do not differ significantly at 5% level (p<0.05) from
each other. Values within a columnhaving superscript (a,b,c) with at least one common alphabet do not differ significantly at 5% level
(p<0.05) from each other.

Table 3: Effect of therapeutic regimens on trace mineral profile of animals with clinical mastitis (Mean±S.E).
Group I Group II
Parameters Days of treatment
(Antibiotic+Antioxidant trace minerals) (Antibiotic only)
Copper (µmol/L) Day 0 6.88±.06a1 7.18±.10a1
b1
Day 10 7.65±.06 7.12±.13 a2
a1
Zinc (µmol/L) Day 0 11.86±.22 11.52±.22 a1
b1
Day 10 13.85±.21 11.13±.16 b2
a1
Mn (µmol/L) Day 0 2.94±.06 2.94±.04 a1
b1
Day 10 3.30±.03 2.93±.05 a2
a1
Se (ng/mL) Day 0 29.94±1.80 32.98±1.16 a2
b1
Day 10 35.39±1.46 31.67±1.08 b2
Values within a column having superscript (a,b) with atleast one common alphabet do not differ significantly at 5% level (p<0.05) from
each other. Values within a row having superscript (1, 2, 3) with atleast one common numericaldo not differ significantly at 5% level
(p<0.05) from each other.

Table 4: Therapeutic efficacy of treatment regimes in animals with a considerable improvement in the oxidative stress
clinical mastitis. parameters, trace mineral status and milk SCC as well as
udder health status as compared to group II indicating
Group II Group III
increased capability of udder to defend against the attack of
Animals treated 12 12 pathogenic microorganisms responsible for causing mastitis
Animals Recovered 10 8 and hence decreased susceptibility to the occurrence of
% Recovery 83.33% 66.66% mastitis in this group. Our findings of improving udder health
Recovery time (Days±S.E) 4±0.5 4.5±0.8 status through feeding of anti-oxidant formulation are in
agreement with Machado et al. (2013),who reported that dairy
with antioxidant vitamins as vitamin A, C, E and β-carotene, cows supplemented with a combination of different trace
and antioxidant minerals as selenium, zinc and copper is elements (Se, Cu, Zn and Mn) showed lower SCC levels
very important to help the animal recover early. (decreased incidence of mastitis) in comparison to the control
In the prophylactic trial, Group I animals supplemented cows. Sordillo and Mavangira (2014) also advocated
with anti-oxidant trace minerals at prophylactic doses showed controlling mastitis through appropriate antioxidant

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 Amelioration of Oxidative Stress Through Supplementation of Self- formulated Anti-oxidant Mixture for Early Recovery....

Table 5: Impact of prophylactic anti-oxidant trace mineral therapy on blood oxidative stress parameters and milk SCC of healthy
actating animals (Mean±S.E).
Group I Group II
Parameters Days of treatment
(With anti-oxidants) (Without anti-oxidants)
SOD (U/mL) Day 0 126.33±1.89a1 125.09±1.74a1
Day 15 135.33±1.90a2 126.65±1.57b1
Day 30 137.33±1.89a2 127.49±1.57b1
Catalase(nmoles/min/mL) Day 0 28.65±.77a1 27.98±1.35a1
Day 15 38.86±.22a2 29.14±1.27b1
Day 30 40.76±.26a3 29.81±1.30b1
GSH(nmoles/mL) Day 0 55.86±1.52a1 57.75±1.20a1
Day 15 59.78±1.19a1 58.81±1.14a1
Day 30 65.23±1.33a2 59.70±1.15b1
MDA(µmoles/L) Day 0 34.09±.91a1 36.74±1.31a1
Day 15 31.56±.78a2 35.52±1.32b1
Day 30 23.89±.63a3 34.80±1.36b1
NO(µmoles/L) Day 0 36.95±1.20a1 36.63±1.21a1
Day 15 34.17±1.07a1 35.66±1.20a1
Day 30 28.01±.89a2 34.68±1.19b1
Day 0 2.81±0.07a1 2.81±0.06a1
Day 15 2.65±0.06a1 2.77±0.06 a1,2
SCC(×105/mL) Day 30 2.48±0.03a2 2.97±0.07b2
15 days after 2.72±0.09a1 3.07±0.06b3
stopping therapy
Values within a row having superscript (a,b) with atleast one common alphabet do not differ significantly at 5% level (p<0.05) from each
other. Values within a column having superscript (1,2,3) with atleast one common numerical do not differ significantly at 5% level
(p<0.05) from each other.

Table 6: Impact of prophylactic anti-oxidant trace mineral therapy on trace mineral profile of healthy lactating animals (Mean±S.E).
Group I Group II
Parameters Days of treatment
(With anti-oxidants) (Without anti-oxidants)
Copper (µmol/L) Day 0 8.08±.08a1 8.09±.05a1
Day 15 8.26±.06a1,2 7.99±.05b1,2
Day 30 8.43±.05a2 7.88±.04b2
Zinc (µmol/L) Day 0 13.87±.17a1 13.83±.12a1
Day 15 14.13±.16a1,2 13.67±.13b1
Day 30 14.56±.14a2 13.54±.14b1
Manganese (µmol/L) Day 0 3.65±.08a1 3.36±.11b1
Day 15 3.95±.08a2 3.32±.10b1
Day 30 4.07±.07a2 3.32±.09b1
Se (ng/mL) Day 0 38.81±1.56a1 36.87±.89a1
Day 15 40.78±1.46a1,2 36.75±.54b1
Day 30 43.58±.99a2 36.47±.46b1
Values within a row having superscript (a,b) with atleast one common alphabet do not differ significantly at 5% level (p<0.05) from each other.
Values within a column having superscript (1,2,3) with atleast one common numerical do not differ significantly at 5% level (p<0.05) from
each other.

supplementation in terms of trace mineral supplementation that CONCLUSION

could potentially boost the animals’ health status and Oxidative stress plays a significant role in the pathogenesis
performance. Ganda et al.,2016 also reported that a single of mastitis as evident by increase in biomarkers of oxidative
subcutaneous injection containing zinc, manganese, selenium, stress like MDA and NO and decreased anti-oxidant
and copper administered to dairy cows with elevated SCC enzymes like SOD, Catalase and GSH in mastitic animals.
reduces the incidence of chronic clinical mastitis, particularly The requirements of trace minerals like Cu, Zn, Mn and Se
in primiparous cows, and tends to increase subclinical mastitis are increased in mastitis as evidenced by their decreased
cure in cows with 3 or more lactations. levels in clinical mastitis and improved clinical response on

6 Indian Journal of Animal Research

 Amelioration of Oxidative Stress Through Supplementation of Self- formulated Anti-oxidant Mixture for Early Recovery....

their supplementation. Anti-oxidant trace minerals like Cu, Ibrahim, H.M.M., El-seedy, Y.Y. and Gomaa, N.A. (2016). Cytokine
Zn, Mn and Se significantly reduce oxidative stress and response and oxidative stress status in dairy cows with
thereby aid recovery in bovine mastitis and play a significant acute clinical mastitis. Journal of Dairy, Veterinary and
role in prophylaxis of mastitis in lactating animals. Thus, Animal Research. 3(1): 2-6.
standardization of oxidative status assessment in dairy cattle If at Ashraf, H.U., Malik, O.S. Shah, S.U. Nabi, A. Muhee, Hilaltantray
is vital as a first step remedy for establishing critical and Muheet Mir. (2017). Retrospective study on prevalence
thresholds that could aid in deûning appropriate protective of mastitis in dairy cattle in Ganderbal district. Journal
in dairy cattle on the basis of antioxidant supplementation. of Entomology and Zoology studies. 5(6): 2567-2568.
Jhambh, R., Dimri, U., Gupta, V.K. and Rathore, R. (2013). Blood
antioxidant profile and lipid peroxides in dairy cows with
REFERENCES clinical mastitis. Veterinary World. 6(5): 271-273.
Abuelo, A., Hernandez, J., Benedito, J.L., Castillo, C. (2013). Oxidative Jin L., Yan, S.M., Shi, B.L., Bao, H.Y., Gong, J. (2014). Effects of
Stress Index (OSI) as a new tool to assess redox status vitamin A on the milk performance, antioxidant functions
in dairy cattle during the transition period. Animal. 7: 1374- and immune functions of dairy cows. Animal Feed Science
1378. and Technology. 192: 15-23.8.
Abuelo, A., Hernandez, J., Benedito, J.L., Castillo. C. (2014). The Kiro R. Petrovski, Patricia T. Eats (2014). Best Practice: W hy
importance of the oxidative status of dairy cattle in the Mastitis Treatment is not Successful? Proceedings of the
periparturient period: revisiting antioxidant supplementation. AVA Annual Conference. pp. F4. 1.1
Journal of Animal Physiology and Animal Nutrition. 99(6): Komine, K., Kuroishi, T., Komine, Y., Watanabe, K., Kobayashi, J.,
1003-1016. Yamaguchi, T., Kamata, S. and Kumagai, K. (2004).
Atakisi, O., Oral, H., Atakisi, E., Merhan, O., Meti n, P.S., Ozcan, Induction of nitric oxide production mediated by tumor
A., Marasli, S., Polat, B., Colak, A. and Kaya, S. (2010). necrosis factor alpha on Staphylococcal enterotoxin C-
Subclinical mastitis causes alterations in nitric oxide, total stimulated bovine mammary gland cells. Clinical and
oxidant and antioxidant capacity in cow milk. Research Diagnostic Laboratory Immunology. 11: 203-210.
in Veterinary Science. 89: 10-13. Khushwaha, N. and Mohan, A. (2019). Role of antioxidants in
Bouchard, L., Blais, S., Desrosiers, C., Zhao, X. and Lacasse, P. management of mastitis in dairy cows. Adv. Complement
(1999). Nitric oxide productionduring endotoxin-induced Alt Med. 3(3): 236-37.
mastitis in the cow. Journal of Dairy Science. 82: 2574-2581. Lykkesfeldt, J. and Svendsen, O. (2007). Oxidants and antioxidants
Celi, P. (2011). Biomarkers of oxidative stress in ruminant medicine. in disease: oxidative stress in farm animals. The Veterinary
Immunopharmacol. Immunotoxicol. 33: 233-240. Journal. 173: 502-511.
Constable, P.D., Radostits, O.M., Gay, C.C., Hinchcliff, K.W. (2017). Maćešić, N., Bačić, G., Božičević, K., Benić, M., Karadjole, T.,
Veterinary Medicine: A Text Book of the Disease of Cattle, Prvanović Babić, N., Lojkić, M., Efendić, M., Baèić, I., Pavlak,
Horses, Sheep, Pigs and Goats. 11th ed. London: Elsevier Ltd. M. (2016). Assessment of the Zagreb mastitis test in
Cigliano, L., Strazzullo, M., Rossetti, C., Grazioli, G., Auriemma, diagnosis of mastitis in dairy cattle. Veterinarski Arhiv. 86:
G., Sarubbi, F., Iannuzzi, C., Iannuzzi, L. and Spagnuolo, 475-485.
M.S. (2014). Characterization of blood redox status of Machado, V.S., Bicalho, M.L.S., Pereira, R.V., Caixeta, L.S., Knauer,
early and mid-late lactating dairy cows. Czech Journal W.A., Oikonomou, G., Gilbert, R.O. and Bicalho, R.C. (2013).
of Animal Science. 59: 170-181. Effect of an injectable trace mineral supplement containing
Costantini, D. and Verhulst, S. (2009). Does high antioxidant capacity selenium, copper, zinc, and manganese on the health
indicate low oxidative stress. Functional Ecology. 23: 506-509. and production of lactating Holstein cows. Veterinary
Deb, R. Kumar, A., Chakraborty, S., Verma A.K., Tiwari, R., Journal. 197: 451-456.
Dhama, K., Singh, U., Kumar, S. (2013). Trends in Marta, W., Elzbieta, M. and Jakimiec, W.R. (2016). Trace elements
diagnosis and control of Bovine Mastitis: a review. Pak. as an activator of antioxidant enzymes. Postepy Hig.
J. Biol. Sci. 16(23): 1653-61. Med. Dosw (online). 70(0): 1483-1498.
Ganda, E.K., Bisinotto, R.S., Vasquez, A.K., Teixeira, A.G.V., Mir, B.A., Razak, R., Ali, A., Muzamil, S., Mir, M.R., Khaliq, T., Baba,
Machado, V.S., Foditsch, C., Bicalho, M., Lima, F.S., O.K., Rehman, M.-U., Bilal, S., Hussain, I. and Rashid,
Stephens, L., Gomes, M.S., Dias, J.M. and Bicalho, R.C. S.M. (2017). Assessment of antioxidant profile in subclinical
(2016). Effects of injectable trace mineral supplementation and clinical mastitis in dairy cattle Journal of Entomology
in lactating dairy cows with elevated somatic cell counts. and Zoology Studies. 5(6): 1022-1025.
J. Dairy Sci. 99: 7319-7329. Miranda, K.M., Espey, M.G.and Wink, D.A. (2001). A rapid, simple
Ganguly, S., Padhy, A., Sahoo, S., Garg, S.L., Wakchaure, R., Praveen, spectroscopic method for simultaneous detection of nitrrate
P.K., Para, P.A., Mahajan, T., Qadri, K. and Sharma, R. and nitrire. Nitric Oxide:Biology and Chemistry. 5(1):
(2016). Antibiogram of milk sample of a farm maintained 62-71.
dairy cow suffering from mastitis followed by its clinical NRC (2001). Nutrient Requirements of Dairy Cattle. 7 th Rev. Ed.
recovery. Int. J. Sci. Environ. Technol. 5(1): 148-151. Natl. Acad. Press, Washington, DC.
Gu, B., Zhu, Y., Zhu, W., Miao, J., Deng, Y.E. and Zou, S. (2009). Patnaik, S., Prasad, A. and Ganguly, S. (2014). Biochemical characteri-
Retinoid protects rats against neutrophil-induced oxidative zation and antibiogram of Staphylococcal microorganisms
stress in acute experimental mastitis. International associated with subclinical mastitis in lactating crossbred
Immunopharmacology. 9: 223-229. cows. Animal Science Reporter. 8(4): 123-129.

Volume Issue 7
 Amelioration of Oxidative Stress Through Supplementation of Self- formulated Anti-oxidant Mixture for Early Recovery....

Russell, 2011; Russell, R.J. (2011). Bovine mastitis. In: Production Transition Period in Dairy Cows. Asian-Australasian
diseases of dairy animals. Satish Serial Publishing House. Journal of Animal Sciences. 24(4): 479-484.
pp. 351-390. Sordillo, L.M. and Mavangira, V. (2014). The nexus between nutrient
Salman, S., Khol-Parisini, A., Schafft, H., LahrssenW iederholt, M., metabolism, oxidative stress and inûammation in transition
Hulan, H.W., Dinse, D., Zentek, J. (2009): The role of cows. Animal Production Science. 54: 1204-1214.
dietary selenium in bovine mammary gland health and Yang, F.L. and Li, X.S. (2015). Role of antioxidant vitamins and
immune function. Animal Health Research Reviews. 10: trace elements in mastitis in dairy cows. Journal of
21-34. https://doi.org/10.1017/S1466252308001588.
Advanced Veterinary Animal Research. 2(1): 1-9.
Sharma, N., Singh, N.K., Singh, O.P., Pandey, V. and Verma, P.K.
(2011). Oxidative Stress and Antioxidant Status during

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