Designation of Benzoic acid  as a feed additive  
 
Ministry of Agriculture, Forestry and Fisheries (MAFF) will designate Benzoic  acid as a  
feed additive and establish its standards and specifications in the ministerial ordinance.  
Outline of standards and specifications is as follows.  
 
Benzoic acid  
 
Compositional specifications in general  
The content of Benzoic acid in feed must be no more than 0. 5%. 
 
Specifications for method  of manufacture  of feed in general  
Benzoic acid  is allowed to be used as feed additives for pigs (no more than 70 kg  of body weight)  
only.  
 
Specifications for feed additives  
Active Substance  
Compositional specifications  
Content:  When this dried product is determined, it contains more than 99.5 % of 
Benzoic acid (C 7H6O2). 
Physical and chemical properties  
(1) It comes in white crystals or crystalline powder.  
(2) Melting point: 121 -123 °C  
Confirmation test:  
(1) Dissolve 1.0 g (0.95 -1.04 g) of this product with  8 mL of 1 mol /L sodium 
hydroxide  solution and add water to make 100 mL. When ferric chloride solution  
is added t o this solution , light yellowish red precipitate is generated , and when 
dilute hydrochloric acid  is added to this solution,  white precipitate is generated.  
Purity test:  
(1) Lead:  
2.0 g (1.95 -2.04 g)  of this product is w eighed into a pot (platinum , quartz, or 
porcelain) , and heat gradually until  the sample begins  to carboni ze. Stop to heat, 
add 1 mL of sulfuric acid and heat gradually until the sample carbonizes and no 
white smoke of sulfuric acid is generated.  If necessary, a dd more sulfuric acid 
and heat until the sample almost carbonizes.  Cover the pot loosely , heat 
gradually and ignite at 450-600°C  until the sample is incinerated. If any 
carbonized material remains, crush with a glass rod, moisten with 1 mL of 
sulfuric acid (1  in 4) and 1 mL of nitric acid, heat until no white smoke is  generated, and ignite to incinerate completely.  To the residue add 10 mL of 
hydrochloric acid ( 1 in 4), heat on a water bath,  and evaporate to dryness.  To the 
residue add a small amount of nitric acid (1  in 100), and dissolve by warming.  
After cooling, add nitric acid (1  in 100) to make exactly 10 mL, and use this 
solution as the Sample solution.  Pipet 0.4 mL of Standard Lead So lution into a 
10-mL flask, add nitric acid ( 1 in 100) to the mark to make 10 mL, and use this 
solution as the Standard solution.  Perform the test with the Sample solution and 
Standard solution as directed under Lead Tests (Atomic Absorption 
Spectrophotomet ry Method 1): the amount of lead shall be not more than 2 μg/g.  
(2) Arsenic:  
0.5 g(0.45 -0.54 g) of this product is w eighed, prepare the sample solution 
according to Method 3 of the Arsenic Test, and perform the test for arsenic 
according to the method usin g Equipment A: the color of the absorbing solution 
shall not be more intense than the standard color (not more than 3 μg/g).  
(3) Phthalic acid : 
To 0.10g (0.095 -0.104 g) of this product, add 1 mL of water and 1 mL of 
resorcinol -sulfuric acid solution , heat in oil bath at 120 -125°C to evaporate the 
water, and heat for 90 minutes.  After cooling, dissolve the residue in 5 mL of 
water, to 1 mL of this solution,  add 10 mL of sodium hydroxide  solution (43 in 
500) shake, and use this solution as the sample solution.  Separately, dissolve 61 
mg of potassium hydrogen phthalate (standard reagent) in water to make 
exactly 1000 mL.  Pipet 1 mL of th is solution, add 1 mL of resorcinol -sulfuric acid 
soluti on, proceed as directed for the sample solution, and use this solution as the 
control solution.  When  the sample solution and the control solution is irradiated 
with  light of 470 -490 nm: the green fluorescence of the sample solution shall not 
be more intens e than that of the control solution (not more than 500 μg/g).  
(4) Related substances:  
Transfer 5.0g (4.95 -5.04 g) of this product to a 25 -mL volumetric flask , add 1 mL 
of propyl benzoate -dimethylformamide solution  to dissolve, then add 
dimethylformamide to make 25 mL, and use this solution as the sample solution.  
Separately, weigh 40 mg each of propyl benzoate (NPB) , biphenyl (BP), 2-
methylbiphenyl (2MBP) , 3-methylbiphenyl (3MBP) , 4-methylbiphenyl (4MBP)  and 
benzyl benz oate(BB) , and 20 g of benzoic acid, transfer to a 100 -mL volumetric 
flask , dissolve in about 50 mL of dimethylformamide, add dimethylformamide to 
make 100 mL, and use this solution as the standard solution.  Perform the test 
with 1 μL each of the sample sol ution and standard solution as directed under 
Gas Chromatography according to the following conditions.  
In the chromatogram of sample solution, i dentify the peaks of biphenyl (BP), 2-methylbiphenyl (2MBP) , 3-methylbiphenyl (3MBP) , 4-methylbiphenyl (4MBP)  and 
benzyl benzoate (BB) from the chromatogram of standard  solution, and calculate 
the peak area Ac 1. For unknown  peak s appearing between the peaks of 2 -
methylbiphenyl and benzoic acid , calculate  sum of the peak areas  Ac1 as 
dimethylbiphenyl isomer . From Ac1 and the peak area Ai 1 of propyl 
benzoate (NBP)  calculated  from the chromatogram of sample solution, the 
content of the related substances ( biphenyl, 2 -methylbiphenyl, 3 -methylbiphenyl, 
4-methylbiphenyl, benzyl benzoate and dimethylbiphenyl isomers) in the sample 
solution shall not exceed 100 μg/g when calculated by the following equation.  
Content  of each  component =𝐴𝑐1×𝑀𝑖1×1000
𝑅𝐹𝑐×𝐴𝑖1×𝑀𝑠 1 
Ai1：peak area of NPB  in the sample solution  
Ac1：peak area of each component  in the sample solution  
Ms1：weight of sample in the sample  solution（g） 
Mi1：weight of NPB in the sample solution（mg） 
RFc：Response factor  of each component  
 
Calculate the response factor (RFc) for each compound using the equation . For 
the calculation of dime thylbiphenyl isomers , use response factors of 3MBP . 
 
Response  factor  (𝑅𝐹𝑐)=𝐴𝑐2×𝑀𝑖2
𝐴𝑖2×𝑀𝑐 2 
Ai2：the peak area of NPB in  the standard solution  
Ac2：the peak area of each component  in the standard solution  
Mc2：amount of each component in the standard solution （mg） 
Mi2：amount of  NPB in the standard solution （mg） 
 
Operating condition:  
Detector:  
Hydrogen flame -ionization detector  
Column:  
A fused silica tube  (inner diameter of 0.32 mm and length of 15 to 30 m ) 
coated with polyethylene glycol modified with nitroterephthalic acid to a 
thickness of 1 μ m  
Column temperature:  
Hold at 100°C for 1 minute, then warm to 210°C at 3°C per minute.  
Inlet temperature:  
240°C (If the temperature programme can be set, the temp erature rises to 270°C at 12°C per second at an initial temperature of 80°C.)  
Detector temperature :  
240-250°C 
Carrier gas:  
Helium  
Flow rate:  
Approx. 3 mL/min. (If pressure control is possible, hold at 47 kPa for 4.35 
minutes, then boost to 58 kPa at 0.28 kPa/min.)  
Injection method:  
Split  
Split ratio:  
1:10 
Loss on drying:  ≤ 0.5 % (1 g, Drying with silica gel, 3 hours)  
Ignition residue: ≤ 0.05 % (1 g)  
Assay:   
This product is dried and approximately 0.5 g of it is weighed to the digits of 0.001 g 
and the value is recorded.  It is dissolve d with  25 mL of neutralized ethanol and 25 
mL of water, and titrate with 0.1 mol/L sodium hydroxide solution (indicator: 3 drops 
of indicator phenolphthalein test solution ). 
0.1 mol/L sodium hydroxide solution 1 mL = 12.21 mg C 7H6O2 
 
Standard  for method  of manufacture  
The products must be  manufactured by air -oxidizing toluene.  
 
Standard  for method  of storage  
The products  must be stored in airtight  containers.  
 
Product  
Compositional specifications  
Same as the Compositional specifications by Active Substance of Benzoic acid . 
 
Standard for method of storage   
Same as the standard for method of storage  by Active Substance  of Benzoic acid .