An investigation into sources of contamination of cattle with the veterinary drug phenylbutazone - Steven Crooks, Veterinary Sciences Division ...
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An investigation into sources of
contamination of cattle with the veterinary
drug phenylbutazone
Steven Crooks,
Veterinary Sciences Division,
Agri-Food and Biosciences Institute,
Belfast, UK
What is Phenylbutazone? N
O
N
CH3
O
• A non-steroidal anti-inflammatory drug (NSAID)
• Abbreviated to PBZ; often called “Bute”
• Introduced to human medicine in 1949 for the treatment of
rheumatoid and arthritic disorders
• Found to induce disorders of the blood including aplastic
anaemia, leukopenia, granulocytosis and thrombocytopenia, in
some cases leading to death
• Consequently, its use in human medicine became limited with
the licence for use in man revoked in the U.K. in 1984
Phenylbutazone ………………. • The principal metabolites are oxyphenylbutazone and γ- hydroxyphenylbutazone • They possess analgesic/anti-inflammatory properties and thus contribute (probably small-moderate degree) to the pharmacological actions of phenylbutazone • For both metabolites, urine concentrations are much higher than those in plasma • Suxibuzone is a pro-drug of PBZ, designed to reduce gastro- intestinal disturbances. Also banned in food producing animals • Alternative NSAIDs are available which do not have the safety issues associated with PBZ i.e. they are MRL compounds
Use in horses
• Choice of NSAID for equines since its introduction to veterinary
medicine in the 1950s
• Substantial clinical history of efficacy and safety in horses
accumulated over both short and long treatment periods
• Problem is that horse meat is consumed in many countries
• The CVMP assessed PBZ in 1997:
• Health risks are blood dyscrasias and the genotoxic & carcinogenic potential
• No thresholds identified so maximum residue limits could not be established
• Therefore PBZ is not permitted for use in any food producing animals
• Findings confirmed by EFSA and EMA 2013
So residues of phenylbutazone are a horse problem?? From 2005-2013, 1.6% of equines tested in EU were non-compliant
However………
EFSA report 2012 and 2013 indicate 0.1% non compliance in bovines in UK
In Northern Ireland, 2008 – present, 0.77% of bovines tested were non
compliant
EFSA 2013 report highlights 41 suspect bovine samples non compliant in
Belgium/Germany
Represents significant non compliance for an unauthorised drug
So ……..it’s a horse and cow problem!!
On farm investigation of two non compliant samples Farm 1 • Plasma sample taken from single animal at slaughter • PBZ confirmed by LC-MS/MS at 0.4 ng/ml • Intentionally fed to bulls due to issues with lameness • Animal sampled was a cow! Farm 2 • Plasma sample taken from single animal at slaughter • PBZ confirmed by LC-MS/MS at 0.32ng/ml • No intentional misuse • Horse on farm fed PBZ in diet – however housed separately from cattle • The dirty bucket theory …………. Is it possible that the bovine problem could be largely due to cross contamination from misuse or from horses?
Investigation of potential contamination sources 1. The dirty bucket – could sharing of feeding vessels provide a bovine with sufficient PBZ to give rise to detectable residues of the drug? 2. Treated animals – could detectable residues be due to contamination from an animal (horse/cow) which had been treated 3. Contaminated pasture – could pasture be sufficiently contaminated to give rise to detectable PBZ concentrations
So how much is a detectable concentration of PBZ? Plasma sample analysed (0.5ml) Supported Liquid Extraction columns Elution in hexane:dichloromethane Evaporate to dryness & reconstitute Analysis using UHPLC-MS/MS CCα phenylbutazone 0.28 µg/L-1
Study 1 – The dirty bucket
Can bovine plasma contain detectable PBZ if fed from a
contaminated bucket?
• A bovine (T1) was treated with PBZ orally, via meal in a bucket
(Pro-Dynam® Oral Powder)
• Dosage regime was that recommended for treatment of a horse
(similar to that being utilized on the treated bulls)
Day 1: 4.4 mg/kg bw twice (two sachets twice daily)
Days 2-4: 2.2 mg/kg bw twice (one sachet twice daily)
Day 5: 2.2 mg/kg bw (one sachet)
• A second bovine (B1) was fed non-medicated meal from the
same bucket each timeStudy 2 – Treated animal contamination • When treatment was complete, T1 was moved into a house with three other steers (H1, H2 & H3)
Study 3 – Pasture contamination • A second bovine (T2) was treated with PBZ in the same way as T1 • After treatment T2 was moved onto a paddock to graze for four days • A group of steers (P1, P2 & P3) were allowed to graze on the paddock for three days, after removal of T2 • A second group (E1, E2 & E3) were allowed to graze on new grass in the same paddock 20 days after removal of P1-P3
The dirty bucket………..
Days after last feeding from Plasma (B1)
contaminated vessel (µg/L-1)
(pre-bleed)Risk by association with treated animals
Days after T1 was introduced to Plasma
the pen µg/L-1 T1
Containing H1-H3 H1 H2 H3
0T2
The Risk from Contaminated Pasture
Plasma
Days after P1-P3 were introduced T2
µg/L-1
to the grass paddock
P1 P2 P3
0Is contaminated pasture really a risk……………?? • Studies unusual in that untreated animals very quickly follow treated onto pasture • Study more closely mimicking normal farming practice • 5 animals housed over the winter period • Extended treatment with PBZ • Manure stored as per normal • Spread onto field mid February • Field remained empty for 70 days • Five animals allowed onto grass
Is contaminated pasture really a risk……………??
Plasma
Days after introduction
µg/L-1
to field
W1 W2 W3 W4 W5
0Is contaminated pasture really a risk……………?? • Trial stopped after 27 days due to lack of grass • Field remained empty for 23 days to permit re-growth • Introduced five new cattle • Sampled at various intervals over 44 days • Only one bovine sample showed detectable concentrations • 0.29 µg L-1 PBZ
Contamination shown to be a risk…………………
• Studies clearly demonstrated the risks of contamination
• How much PBZ is required to provide a detectable concentration?
• Normal dose is ~ 2.2mg/kg bodyweight
• Four bovines given a single dose of PBZ
• Plasma sample taken 1 day after treatment
PBZ
PBZ Conc. in
incorporated in
plasma (µg/L-1)
1 kg of feed (µg)
Therapeutic 1,100,000 --------
C1 100,000 1279
C2 1000 13.80
C3 30 0.290
C4 20Conclusions • Residues are a significant possibility as a result of contamination from legal use of PBZ • PBZ is used therapeutically at high concentrations yet extremely small amounts can give rise to detectable concentrations in plasma • Environmental contamination is clearly an issue • Is PBZ contamination surface contamination only? • Difficult to limit contamination risk • Alternative NSAIDs available for use • Is the ongoing authorisation for use of PBZ in horses justified?
Thanks • Colleagues from AFBI: Terence Fodey, Wesley Smyth, Paul Barnes, Imelda Traynor & Glenn Kennedy for their assistance in this study • Organising Committee and Scientific Committee for providing the opportunity to present this work • You for your attention
Contamination Investigation Fodey, T. L., Smyth, W.G., Barnes, P., Traynor, I.M., Kennedy, D.G., Crooks, S.R.H. (2015) Investigation into sources of contamination of cattle with phenylbutazone. Veterinary Record 176(3) 74-76
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