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Ethnoveterinary formulation for treatment of bovine mastitis

Article · March 2017

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Research & Reviews: Journal of Veterinary Sciences

Ethno-veterinary Formulation for Treatment of Bovine Mastitis

Balakrishnan Nair MN1*, Punniamurthy N2, Mekala P2, Ramakrishnan N2 and Kumar SK1
1
TransDisciplinary University, School of Health Sciences, Veterinary Ayurveda group, Bangalore-560106,
Karnataka, India
2
EVM Herbal Training and Research Centre, Tamil Nadu Veterinary and Animal Sciences
University, Thanjavur-613403, India

Research Article

Received date: 16/05/2017 ABSTRACT

Accepted date: 16/06/2017
Introduction: The objectives of the study were to carry out clinical
Published date: 23/06/2017
trial of ethno-veterinary medicine for mastitis, to conduct an intervention
impact analysis on the reduction of antibiotic residue in the milk when
*For Correspondence herbal formulation is used and to evaluate in vitro the pharmacological
understanding of the ingredients in the formulation based on the
Balakrishnan Nair MN, TransDisciplinary published literature and phytochemical assessment.
University, University School of Health Sciences,
Veterinary Ayurveda Group, Methods: The herbal formulation for mastitis was documented
74/2 Jarakabandekaval, Attur Post, from a healer and the safety and efficacy were confirmed using the
Yelahanka, Bangalore, Karnataka, India. methodology suggested earlier [3] . Clinical mastitis was identified by
California Mastitis Test (CMT) method. The pH, Somatic Cell Count (SCC)
E-mail: nair.mnb@tdu.edu.in; and Electric Conductivity (EC) were monitored and documented before and
nairunni2003@yahoo.co.uk post treatment with Ethno-Veterinary Medicine (EVM) in the field studies
involving 27 animals. Freshly prepared fine paste of herbal formulation
Keywords: Ethno-veterinary, Natural/herbal was applied externally over the udder 10 times per day for 7 days. Before
remedies, Livestock health, Medicinal plants, each application, the udder was washed well with water and the milk
Antibiotic residue, Antibiotic resistance from all quarters was removed completely. The affected cows were also
fed with 2 lemon fruits twice daily. Bulk milk samples were tested for the
presence of antibiotic residue(s) in milk before and after intervention. The
pharmacological understanding of the ingredients in the formulation was
evaluated based on the literature and phytochemical assessment.
Results: The select parameters (pH, SCC, EC) in the animals with
mastitis became normal within 6-7 days of treatment. The milk production
returned to near normal to the pre-mastitis level. Hence, the mastitis can
be efficiently managed with this formulation. The intervention impact
analysis showed 18 to 49% reduction in antibiotic residues in the milk.
The combination [Aloe vera (L.) Burm. F, Curcuma longa L. and calcium
hydroxide] acts synergistically and provides effective cure in clinical
mastitis owing to its broad spectrum antimicrobial, anti-inflammatory and
immunomodulatory activities.
Conclusion: Mastitis can be efficiently managed with this
formulation. The intervention impact analysis showed 18-49% reduction
in antibiotic residues in the milk. The combination [Aloe vera (L.) Burm.
F, Curcuma longa L. and calcium hydroxide] acts synergistically and
provides an effective cure in clinical mastitis owing to its broad spectrum
antimicrobial, anti-inflammatory and immunomodulatory activities.

INTRODUCTION
In the last 50 years, mastitis in India reportedly increased 115 times [1]. The incidence of resistant mastitis in dairy cows in
Tamil Nadu, India is 56.01% -Escherichia coli 50.64%, Staphylococcus aureus 44.25% and Methicillin-resistant Staphylococcus
aureus 5.11% [2]. Hence, an alternate approach was attempted.

RRJVS | Journal of Veterinary Sciences-S1, March, 2017 25

Research & Reviews: Journal of Veterinary Sciences

MATERIALS AND METHODS

The safety and efficacy of the herbal formulation for mastitis were confirmed using the methodology suggested earlier [3].
Clinical mastitis was identified by CMT method. The pH, (SCC) and (EC) before and 6 days post treatment with EVM involving 27
animals were reported. Freshly prepared fine paste of Aloe vera leaves, Curcuma longa rhizome and calcium hydroxide was applied
externally over the udder 10 times per day for 7 days. The udder was washed well with water and the milk from all quarters was
removed completely before each application. The affected cows were also fed with 2 lemon fruits twice daily. Bulk milk samples
collected from the farmers before and after intervention and tested for the presence of antibiotic residue(s) in milk [4,5]. The
formulation was evaluated in vitro for the pharmacological understanding based on the literature and phytochemical assessment.

RESULTS
Figure 1. A-C shows cow treated with clinical mastitis using the herbal formulation has got cured within 6 to 7 days of
treatment. The pH, SCC, and EC of milk became normal within 6 days of treatment (Figure 2). The combination provides broad
spectrum antimicrobial, anti-inflammatory, and immunomodulatory activities. Post-intervention impact analysis after one year
showed up to 18-49% reduction in antibiotic residues (Figures 3-7) in the milk suggestive of the efficacy of the herbal combination
in mastitis and in reducing the use of antibiotics. There was an outbreak of foot and mouth disease in certain areas in 2014
and therefore, there is an increase of tetracycline, Ceftriaxone and Ciprofloxacin in the milk certain centres (Figure 5-7). The
phytochemical screening of the formulation revealed the presence of alkaloids, saponins, steroids, carbohydrates, flavonoids,
terpenoids, triterpenoids, sterols, emodin and diterpenes. TLC analysis revealed the presence of flavonoids, phenols and glycosides
and they exhibited antioxidant activity [6,7]. The molecular docking study revealed that the active ingredients from Aloe vera and
Curcuma longa interact with the proteins that play crucial role in Staphylococcus aureus.

DISCUSSION
As per the texts of Ayurveda, mastitis is known as Sthanavidhradi, a disease of pitta origin, the drugs used in this formulation
(Aloe vera, Curcuma longa and Calcium hydroxide) is potent pitta shamaka (Pacifies Pitta humour). The formulation possesses
Krimighna (antimicrobial), Vranashodaka (wound cleanser), Vranaropaka (wound healing), Shothahara (anti-inflammatory) and

(A) (B) (C)

Figures 1. A cross bred cow treated with clinical mastitis using the herbal formulation.

5 Before treatment

After treatment
4
Normal Milk
3

0
pH EC SCC

Figure 2. The figure shows reduction of pH, Electrical Conductivity (EC) and Somatic Cell Count (SCC) before and after treatment with herbal
formula in comparison with normal values.
RRJVS | Journal of Veterinary Sciences-S1, March, 2017 26
Research & Reviews: Journal of Veterinary Sciences

500

450

400

350

300

250 2013 (µ/l)

200 2014 (µ/l)

150

100

50

0
Concentration of Concentration of Concentration of Concentration of
Gentamicin Tetracycline Ceftriaxone Ciprofloxacin
Figure 3. Mean antibiotic residue with Standard error (Gentamicin, Tetracycline, Ceftriaxone and Ciprofloxacin µg/L) in the milk before (2013)
and after intervention (2014).

700

600

500

400
Gentamicin 2013 (µ/l)
300 Gentamicin 2014 (µ/l)

200

100

0
1 2 3 4 5 6 7 8 9 10 11 12 13

Figure 4. Gentamicin in the milk from 13 milk collection centres before (2013) and after intervention (2014).

600

500

400

300 Tetracycline 2013 (µ/l)

Tetracycline 2014 (µ/l)

200

100

0
1 2 3 4 5 6 7 8 9 10 11 12 13
Figure 5. Tetracycline in the milk from 13 milk collection centres before (2013) and after intervention (2014).

RRJVS | Journal of Veterinary Sciences-S1, March, 2017 27

Research & Reviews: Journal of Veterinary Sciences

140

120

100

80
Ceftriaxone 2013 (µ/l)
60 Ceftriaxone 2014 (µ/l)

40

20

0
1 2 3 4 5 6 7 8 9 10 11 12 13

Figure 6. Ceftriaxone in the milk from 13 collection centres before (2013) and after intervention (2014).

25

20

15
Ciprofloxacin 2013 (µ/l)
Ciprofloxacin 2014 (µ/l)
10

0
1 2 3 4 5 6 7 8 9 10 11 12 13
Figure 7. Ciprofloxacin in the milk from 13 milk collection centres before (2013) and after intervention (2014).

Srotoshodaka (channel cleanser) properties. Hence, mastitis can be efficiently managed with this formulation. The antimicrobial
activity of Aloe vera is attributed to the anthraquinones (aloin and emodin), flavonoids, tannins (active against MRSA), saponins,
p-coumaric acid, ascorbic acid, pyrocatechol and cinnamic acid. Alkaloids, tannins, phenolics, terpenoids, phytosterols, saponins,
flavonoids, glycosides, fatty acids such as palmitoleic acid and α-turmerone in fixed oils of Curcuma longa also possess antimicrobial
activity against wide range of bacteria. The anti-inflammatory activity of Aloe vera is reported to be due to Brady kinase which
decreases vascular permeability, neutrophil migration, and leukocyte adhesion and reduces edema formation. It is also found to
decrease the production of TNFα, inhibit PGF2α and TB4. Curcumin, the active principle of Curcuma longa is reported to inhibit
NF-κB which in turn decreases TNF-α, superoxides, COX-2, iNOS and NO. It inhibits LOX pathway and decreases the formation of
leukotriene. Calcium hydroxide is known to possess anti-inflammatory action and reduces edema formation. Thus all the three
ingredients in the formulation act at various steps in the inflammatory pathway and synergistically produce anti-inflammatory
effect [6-9].
Polysaccharides present in Aloe vera are rich in mannose and act as biological response modifier by targeting antigen
presenting cell and cytokine cascade. Acemannan increases TNFα, IL-1B, IFN γ, IL-2 and IL-6; aleoride increases NFκB activation
and stimulate macrophages which in turn increase nitric oxide production and other cytokines responsible for immunomodulation.
Curcumin also possesses immunomodulatory and antioxidant activity[9]. The urgent need of dissemination of this traditional
veterinary practice is highlighted in the light of the benefits it provides.

CONCLUSION

This traditional formulation is very effective in the cure of mastitis. The intervention impact analysis showed 18 to 49%
reduction in antibiotic residues in the milk which indicates EVM based natural products is an effective alternative to synthetic
chemicals in treating mastitis in cattle.
RRJVS | Journal of Veterinary Sciences-S1, March, 2017 28
Research & Reviews: Journal of Veterinary Sciences

ACKNOWLEDGEMENT

The authors thank Indian Council of Agriculture Research and ETC Foundation the Netherlands for financial support. We also
thank Dr. P. M. Unnikrishnan for his suggestions during the preparation of the paper and the local healers and farmers who have
shared their traditional knowledge.

COMPETING INTERESTS

The authors declare that they have no competing interest.

REFERENCES
1. Anon. NDRI News. 2012;17:1.
2. Chandrasekaran D, Nambil AP, Thirunavukkarasu PS, et al. The incidence of resistant mastitis in dairy cows in Tamil Nadu,
India. J Appl Nat Sci. 2015;7:304-308.
3. Raneesh S, et al. Documentation and participatory rapid assessment of ethnoveterinary practices. Indian J Tradit Knowl.
2008;7:360-364.
4. Kaya SE and Filazi A. Determination of antibiotic residues in milk samples. Kafkas Univ Vet Fak Derg. 2010;16:31-35.
5. Nagwa NHS, El-Hasheme E, El-Enany N, et al. Kinetic spectrophotometric method for the determination of tetracycline
hydrochloride in pharmaceutical formulations. Arc Apl Sci Res. 2009;1:1-11.
6. Bellik Y, et al. Molecular mechanism underlying anti-inflammatory and anti-allergic activities of phytochemicals: An update.
Molecules 2013;18:322-353.
7. Liju VB, et al. An evaluation of antioxidant, anti-inflammatory and antinociceptive activities of essential oil from Curcuma
longa. L. Indian J Pharmacol. 2011;43:526-531.
8. Pandey GS (2006) Commentary of Dr. K. C. Chunekar, Bhavaprakasha Nighantu of Sri Bhavamishra, Hareethakyadi varga
verse no. 195, Varanasi, Chowkamba Bharathi Academy.
9. Park MY, et al. Evaluation of aloe and aloe-emodin as anti-inflammatory agents in aloe by using murine macrophages.
Biosci Biotechnol Biochem. 2009;73:828-832.

This article was published under the special issue, The Lost Love: “ONE HEALTH” in the era of Antimicrobial Resistance
handled by Editor. Vikas Saxena, Center for Vascular and Inflammatory Diseases, School of Medicine, University of
Maryland, USA

RRJVS | Journal of Veterinary Sciences-S1, March, 2017 29

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