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20 April 2023  
EMA/CVMP/152100/2023   
Committee for Veterinary Medicinal Products  
Opinion of the Committee for Veterinary Medicinal 
Products on the establishment of maximum residue 
limits  
Procedure no: EMEA/V/MRL/003450/EXPL/0005  
Name of the substance: Rafoxanide (INN)  
Basis for the opinion  
Pursuant to Article 27(2) of Regulation (EC) No 470/2009 of 6 May 2009 , Ireland  submitted to the 
European Medicines Agency on 20 February 2023  a request  for the extrapolation  of maximum 
residue limits for rafoxanide to bovine and ovine milk. 
Recommendation  
The Committee, having considered the request , recommends by consensus  the extrapolation  of 
maximum residue limits for rafoxanide  to bovine and ovine milk . Furthermore, with reference to 
Article 5 of Regulation (EC) No 470/2009 and in line with the criteria laid down in Commission 
Regulation (EU) 2017/880, the Committee agreed to  extrapolate the maximum residue limits in 
bovine tissues and milk to all ruminants except ovine, for which (higher) MRLs are already 
established . Therefore, the Committee recommends the ext rapolation  of maximum residue limits for 
rafoxanide  in accordance with the following table:  
   
  
Opinion  of the Committee for Veterinary Medicinal Products on the establishment of maximum residue limits   
EMA/CVMP/152100/2023  Page 2/11  
Pharmaco -
logically 
active 
substance  Marker 
residue  Animal 
species  MRLs  Target tissues  Other 
provisions  Therapeutic 
classification  
Rafoxanide  Rafoxanide  All 
ruminants 
except 
ovine  30 μg/kg  
30 μg/kg  
10 μg/kg  
40 μg/kg  
10 μg/kg  Muscle  
Fat 
Liver 
Kidney  
Milk NO ENTRY  Antiparasitic 
agents/Agents 
(acting) 
against 
endoparasites  
Ovine  100 μg/kg  
250 μg/kg  
150 μg/kg  
150 μg/kg  
10 μg/kg  Muscle  
Fat 
Liver 
Kidney  
Milk 
 
The Norwegian  CVMP member agrees   with the a bove-mentioned recommendation of the 
Committee.  
The scientific conclusions of the Committee are presented in the European public MRL assessment 
report (EPMAR), provided in Annex I of this opinion.  
The analytical method for monitoring of residues is appende d to this opinion.  
The present opinion is forwarded to the European Commission and to Ireland together with its 
appendices.   
  
  
 Annex I   
 
 
European public MRL assessment report (EPMAR)  rafoxanide (bovine and ovine milk)   
EMA/CVMP/152100/2023   Page 4/11 
 European public MRL assessment report (EPMAR)  
Rafoxanide ( bovine and ovine  milk)  
Summary of the scientific discussion for the extrapolation of 
MRLs  
Substance name:  Rafoxanide  
Therapeutic class:  Antiparasitic agents/Agents against endoparasites  
Procedure number:  EMEA/V/MRL/00 3450/EXPL/0005 
Requesting Member State:  Ireland  
Target species:  Bovine and ovine milk  
Intended therapeutic indication:  Fasciolicide  and active against gastrointestinal nematodes and 
sheep nasal bot fly  
Route of administration:  Oral 
1.  Introduction  
Rafoxanide is a halogenated salicylanilide, [3' -chloro -4'-(p-chlorophenoxy) -3,5-diiodosalicyl -anilide] 
used as a fasciolicide in cattle and sheep. Rafoxanide is also active against gastrointestinal nematodes 
(Haemonchus , Bunostomum , Oesophagostomum , and Gaigeria  species) and against the sheep nasal 
bot fly ( Oestrus ovis ). Rafoxanide products are currently marketed in the EU for the treatment o f cattle 
and sheep. It is generally administered by the oral route at a dose of 11.25 mg/kg body weight.  
Rafoxanide  was previously assessed by the CVMP and a toxicological ADI of 2 µg/kg, i.e. 120  µg per 
person was established.  
Currently, r afoxanide  is included in Commission Regulation (EU) No 37/2010 in accordance with the 
following table:  
Pharmaco - 
logically active 
substance  Marker 
residue  Animal 
species  MRLs Target 
tissues  Other 
provisions  Therapeutic 
classification  
Rafoxanide  Rafoxanide  Bovine  30 μg/kg  
30 μg/kg  
10 μg/kg  
40 μg/kg  Muscle  
Fat 
Liver 
Kidney  NO ENTRY  Antiparasitic 
agents/Agents 
against 
endoparasites  
Ovine  100 μg/kg  
250 μg/kg  
150 μg/kg  
150 μg/kg  Muscle  
Fat 
Liver 
Kidney  
Bovine, 
ovine   10 μg/kg  Milk Provisional 
MRL shall 
expire on 31 
December 
2017  
 
 
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EMA/CVMP/152100/2023   Page 5/11 
 
 The provisional MRL for bovine and ovine milk expired on 13 December 2017. Subsequent to that, on 
the 21 February 2023, Ireland submitted to the European Medicines Agency , a request for an opinion 
on the extrapolation of maximum residue limits for rafoxanide to bovine and ovine milk, pursuant to 
the second paragraph of Article 27 of Regulation (EC) No 470/2009.  
The scientific assessment carried out by the Committee leading to the recommendation for the 
establishment of MRL s in bovine and ovine milk is reported in the paragraphs below. The 
considerations are largely identical to those that led to the establishment of a provisional MRL in 
bovine and ovine milk in 2014. 
2.  Scienti fic risk assessment  
2.1.  Safety assessment  
The CVMP has previously performed a consumer safety evaluation for rafoxanide and established a 
toxicological ADI of 2 µg/kg, i.e. 120  µg per person based on a NOEL of 400  µg/kg  bw/day from a 
thirteen week study in do gs and applying a safety factor of 200 . Therefore , no further assessment 
regarding the establishment of the ADI of the substance is required for the purpose of this request.  
2.2.  Residues assessment  
2.2.1.  Pharmacokinetics in target species  
No pharmacokinetic data for  milk were made available in sheep and cattle.  
The pharmacokinetic data previously assessed by the CVMP is reported in the paragraphs below.  
In sheep, after an oral administration of 14C-rafoxanide at a dose of 11.25  mg/kg  bw, about 0.12% of 
the administer ed dose was recovered in urine within 3 days. The major metabolite was  
3,5-diiodosalicylic acid, representing 91% of the excreted radioactivity; rafoxanide accounted for only 
9%. No information on the percentage of radioactivity excreted via faeces was gi ven. The maximum 
plasma concentration of radioactivity (19  750 µg equivalents rafoxanide/kg) occurred between 
1.4 to 1.8 days after administration. An apparent radioactive elimination half -life of 8.9 1.2 days was 
calculated . This persistence of rafoxanide residues was due to the fact that the compound was strongly 
bound to plasma proteins (greater than 99%). No information on the ratio of the parent compound to 
total radioactivity in plasma was available.  
In cattle, after an  oral administration of 14C-rafoxanide at a recommended therapeutic dose of 
11.25  mg/kg  bw, less than 0.6% of the administered dose was recovered in the urine within 6 days. 
Unchanged rafoxanide and 3,5 -diiodosalicylic acid were the 2 major substances dete cted. However, 
due to low concentrations of these compounds and as the different system of chromatographic analysis 
led to different separation, no figures can be given. No information on the percentage of radioactivity 
excreted via faeces was given. The m aximum plasma levels (close to 20  000 µg equivalent 
rafoxanide/kg) occurred at 1.8 days post dose. The apparent radioactive elimination half -life was 
3.87  0.59 days. No information on the ratio of the parent compound to tot al radioactivity in plasma 
was available.   
 
 
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 2.2.2.  Residue depletion studies  
In a non -GLP study nine pregnant dairy cows were treated with a single dose (11.21 to 11.27 mg/kg 
bw) of a 3% oral suspension at the start of the dry period. Calving occurred between 26 and 62 days 
after treatment. Milk samples were analysed using UPLC -MS/MS.  
Highest concentrations of rafoxanide occur in cows that calve less than 35 days after treatment and in  
the milk produced immediately after calving . The concentrations  ranged from lower than 1.0 μg/kg to 
50 μg/kg. After the colostrum period, this is from about 3 days after calving, concentrations of 
rafoxanide in milk are typically low (<10 µg/kg) but persist at concentrations above the limit of 
quantification of the analytical method  (1.0 μg/kg) for between 1 0 and 38 days post -calving. Mean 
concentrations are lower than 10 µg/kg at 2 days post -calving.  
Administration to lactating cows by oral drench  of 11.75 mg/kg resulted in high concentrations in milk 
with maximum 3 to 4 days after treatment ( 516 ± 138 µg/kg ).  
There are no data available for the depletion of residues in ovine milk.  
Selection of marker residue and ratio of marker to total residues  
No radiolabelled data are available to confirm the identity of the marker residue and on which to base 
a ratio of  marker to total residues. The parent compound , rafoxanide , has been detected in milk but 
there are no data on metabolites in milk.   
Standard procedure is to determine the ratio of marker to total residues from radiolabelled depletion 
studies and the value  is chosen at a time point at which residue intake will be below the ADI.  
Since the metabolism of rafoxanide in milk has not been characterised and the ratio has not been 
determined, any approach to setting a ratio of marker to total residues should be sui tably 
conservative. In bovines, rafoxanide represented 50%, 50% and 30% of the total residues in muscle, 
fat and kidney, respectively, 14 days after treatment. In ovines rafoxanide represented 100%, 88%, 
50% and 50% of the total residues in muscle, fat, li ver and kidney, respectively, 30 days after 
treatment.  
Article 7(e) of Commission Regulation 2017/880 of 23 May 2017 indicates that, when extrapolating 
from tissues to milk, consideration shall be given to the physicochemical characteristics of the 
substan ce and whether these suggest accumulation in milk. From published information, it is noted 
that log Kow is about 7 -8, indicating that rafoxanide would accumulate in fat. Given the limited 
metabolic capacity of milk, it is considered reasonable, in this cas e, to establish a ratio of marker to 
total residues identical to that established for fat, as fat also has limited metabolic capacity. Therefore 
the worst -case value of 0.5, established for bovine fat, is retained as the ratio of marker to total 
residues i n bovine milk .  
2.2.3.  Monitoring or exposure data  
Results of the Irish national residue monitoring programme f or 2008 to October 2010 were provided. 
Rafoxanide was detected in one of 284 samples of tested bulk milk  in 2009 . The level of rafoxanide in 
the positive sample was 9.83 μg/kg but no information is available on when the animal(s) was treated 
with rafoxanide.  Rafoxanide was not detected in any samples in either 2008 or 2010.   
 
 
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 2.2.4.  Analytical method for monitoring of residu es 
A UPLC-MS/MS (ultra -performance liquid chromatography coupled to tandem mass spectrometry) 
method was developed by an Irish national reference laboratory for the purposes of residue 
surveillance in cow’s milk . However, the method is not fully validated  in line with the requirements of 
VICH GL 49 . In particular,  some information is missing on preparation of samples, accuracy, precision, 
stability. Also , the concentration range over which the method was validated did not include the MRL 
proposed. The limit  of quantification is 6.0 µg/kg.  
Concerning the applicability of the analytical method in sheep and goat milk, a similarity in the 
validation of the parameters  of accuracy and precision  is noticed when bovine, caprine and ovine milks 
are compared .   
The p roposed method has been reviewed by the relevant European Union Reference Laboratory, which 
confirmed the suitability of the method to control the proposed MRL with comparable performance for 
milk from the different animal species . 
Notwithstanding the defi ciencies identified, the data provided indicate that the analytical method has 
the potential to comply with the requirements of Regulation (EU) 2018/782. That is, the marker 
residue rafoxanide can be measured in bovine milk, in accordance with the guidance  on validation of 
the analytical method as detailed in VICH GL49.  Moreover , it is considered that the requirements for 
extrapolation of the Commission  Regulation  (EU) 2017/880 on extrapolation of MRLs are fulfilled, 
specifically Article 4,  that “a suitably  validated method is available for the reference species” . This is 
the case as a validated analytical method for bovine and ovine tissue is available.  
2.2.5.  Findings of EU or international scientific bodies  
No information was available on evaluation from other scientific bodies.  
3.  Risk management considerations  
3.1.  Potential effects on the microorganisms used for industrial food 
processing  
Microbiological effects are not expected for this substance therefore such data are not considered 
necessary.  
3.2.  Other relevant risk  management considerations for the establishment 
of maximum residue limits  
The data provided for the scientific evaluation of rafoxanide for the establishment of a maximum 
residue limit in milk were limited and do not fully meet the criteria described in C ommission Regulation 
(EU) 2018/782 of 29 May 2018 . In particular , the studies provided are not GLP compliant and no data 
on total residues in milk are available. However, the current evaluation relates to an extrapolation 
request in line with Regulation 20 17/880 and is considered to meet the relevant requirements.  
Recognising the deficiencies in the data presented the Committee took also into account the following:   
 
 
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EMA/CVMP/152100/2023   Page 8/11 
 
  Although other flukicidal substances exist for which MRLs in ruminant milk have been establis hed, 
these substances are not approved for the treatment of immature fluke, and consequently it is 
recognised that at present there is a lack of available products, authorised for the treatment of 
immature fluke in animals producing milk for human consumpt ion; 
 Liver fluke is a highly debilitating disease leading to loss of condition and ultimately cachexia and 
potentially death  and therefore the availability of  an adequate range of  products for the treatment 
of immature  fluke is essential in order to avoid unnecessary suffering of the animals;  
 The establishment of a maximum residue limit is essential to provide the reference level for control 
purposes and to enable the use of the substance;  
 The lack of available products coupled with welfare issues may lead to increased use of products 
under non -authorised conditions.  
3.3.  Extrapolation of MRLs  
The extrapolation to milk has been assessed in line with the requirements laid down in Commission 
Regulation 2017/880 of 23 May 2017, particularly articles 4 and 7 of that regulation.  
Based on the existing MRL values, the daily intake of residues from ovine tissues, which represents a 
worst case calculation, equates to 89.3 µg (equivalent to 74.3% of t he ADI), leaving 30.8 µg 
(equivalent to 25.7% of the ADI) for the establishment of a MRL for milk.  
Given the available information relating to residues in other tissues, rafoxanide was retained as marker 
residue in bovine milk and  a ratio of marker to tota l residues of 0.5 was estimated  from bovine and ovine 
meat depletion profile . 
Considering that the standard food basket indicates a consumption for milk of 1.5 kg per consumer per 
day, and in order to ensure that consumer exposure total residues of rafoxan ide remains below the ADI, 
the maximum allowable total residues in milk would be 10 µg/kg . 
In view of the information available and the risk management considerations , the CVMP recommends 
the extrapolation of the existing MRLs for rafoxanide in cattle to c attle milk. The proposed MRL is  
10 µg/kg.  
Although data were not provided to demonstrate the presence of the marker residue rafoxanide in 
sheep milk, metabolism in plasma and tissues is known to be limited, and therefore rafoxanide can be 
accepted as the marker residue  in sheep milk . Although the ratios of marker to total residues 
established in sheep tissues are not the same as those established for bovine tissues, for the purpose 
of establishing an MRL in milk and in view of the fact that  ovine animals producing milk for human 
consumption are considered as a minor species , the marker to total residue of 0.5 agreed for bovine 
milk is also considered to be acceptable for ovine milk.  
Therefore , the proposed MRL of 10  μg/kg  for cattle can also  be accepted for sheep milk.  
The analytical method for milk was not fully validated. However , requirements of Commission  
Regulation (EU) 2017/880 on extrapolation of MRLs have been fulfilled, notably a validated analytical 
method for monitoring residues in bovine and ovine tissues is available . Moreover , in any future 
application for marketing authorisation for rafoxanide in milk, a fully validated analytical method in 
support of residue depletion studies will be required.  
Available data indicate that follo wing the oral administration of 1 1.2 mg/kg bw to cows with a dry 
period of at least 54 days residue values in milk were below 10 μg/l from 3  days after calving  while  
 
 
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 median residue values were below 10 μg/l from two days after calving . It is also noted tha t rafoxanide -
containing products are authorised for administration by other routes (subcutaneous and topical) and 
this may impact on the time required for residues in milk to deplete to the level of the recommended 
MRL.  
Calculation of theoretical daily in take of residues  
Details used in the calculation of theoretical daily intake of residues from ovine tissues and milk  (worse 
case scenario):  
Edible tissue or 
products  Daily 
consumption 
(kg) MRL proposal 
(µg/kg)  Ratio of the 
marker/total 
residue  Amount per edible 
tissue or product  
Muscle  0.30 100 1.00 30 µg  
Fat  
  0.05 
 250 0.88 14.2 µg  
Liver 0.10 150 0.50 30 µg  
Kidney  
  0.05 
 150 0.50 15 µg  
Milk 1.50 10 0.5 30 µg 
Total    119.2  µg 
(99.33 % of the 
ADI) 
4.  Considerations on possible extrapolation of MRLs  
In line with Article 5 of Regulation (EC) No 470/2009, the CVMP considered the possibility of 
extrapolating the maximum residue limits recommended for rafoxanide to other food producing species 
and commodities. Taking into account the  provisions laid down in Commission Regulation (EU) 
2017/880, the recommendations on extrapolation are justified as follows:  
Animal species/ 
food commodities  Extrapolation 
possible 
(Yes/No)  Justification  
All ruminants, 
including milk  Yes Pharmacokinetics and residue data in ruminants other than 
cattle and sheep are not available. However, as other 
ruminants are considered minor species, in line with 
Commission Regulation (EU) 2017/880, extrapolation of the 
MRLs recommended for bovine (lowe r than for sheep meat) 
can be accepted.  
Other species than 
ruminants  No Pharmacokinetic and residue data were only available in 
bovine and ovine. In the absence of supporting data 
demonstrating similarity of metabolism, and in line with 
Commission Regulation (EU) 2017/880, extrapolation to 
unrelated species is not recommended.   
 
 
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 5.  Conclusions and recommendation for the establishment of 
maximum residue limits  
Whereas:  
 the toxicological ADI of 2 μg/kg bw (i.e 120 μg/person) was established as  the overall ADI for 
rafoxanide;  
 rafoxanide  has been accepted as the marker residue in  cattle and sheep  milk; 
 the marker to total residues ratio of 0.5 established for bovine fat was considered a suitabl y 
conservative value to be considered in relation to milk given that milk has limited metabolic 
capacity ; 
 there is a lack of available products for the treatment of immature fluke in animals produ cing milk 
for human consumption;  
 there is a need for a reference level for control purposes and to enable the use  of the substance ; 
and having considered that:  
 an analytical method for the monitoring of residues of rafoxanide in milk is availab le and 
considered fit for purpose while not fully validated;  
 a validated analytical method for monitoring residues in bovine and ovine tissues is available;  
 
the Committee, having considered the request from Ireland, recommends the extrapolation of 
maximum residue limits for rafoxanide to milk in bovine and ovine and the amendment of the entry in 
Table 1 of the Annex to Commission Regulation (EU) No 37/2010 in accordance with the following 
table:   
Pharmaco - 
logically 
active 
substance  Marker 
residue  Animal 
species  MRLs Target 
tissues  Other 
provisions  Therapeutic 
classification  
Rafoxanide  Rafoxanide  All 
ruminants 
except 
ovine   30 μg/kg  
30 μg/kg  
10 μg/kg  
40 μg/kg  
10 μg/kg  Muscle  
Fat 
Liver 
Kidney  
Milk NO ENTRY  Antiparasitic 
agents/Agents 
(acting) 
against 
endoparasites  
Ovine  100 μg/kg  
250 μg/kg  
150 μg/kg  
150 μg/kg  
10 μg/kg  Muscle  
Fat 
Liver 
Kidney  
Milk 
Based  on these values, the theoretical maximum daily intake from ovine tissues and milk is 119.2  µg, 
which corresponds to 99.3% of the  ADI.  
 
 
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 6.  Background information on the procedure  
Submission of the dossier       20 February 2023  
Steps taken for assessment of the  substance  
 Clock started:    21 February 2023  
 CVMP opinion on MRLs adopted :   20 April 2023