iGEM - Champion Park

1.Introduction

Antibiotics are a type of product with antiviral potential or other activities which is produced by microorganism (bacteria, fungi and actinomycetes), higher animals and plants, or chemical synthesis methods. It can be classified as the following 8 types: (1)the β-lactams, such as penicillin, cephalosporin, etc.; (2) the aminoglycosides, such as streptomycin, gentamicin, etc.; (3) the tetracyclines, such as tetracycline, chlortetracycline, oxytetracycline, and etc.; (4) the large ring lactones, such as erythromycin, etc.; (5) the benzene hydrocarbon amines, such as chloramphenicol, etc.; (6) the sulfonamides, such as sulfamethoxazole, benzyl methyl oxygen organism; (7) the quinolones, such as ciprofloxacin and ofloxacin, norfloxacin and levofloxacin; (8) and the others, like polyene big ring, polypeptide, anthracycline-based, etc.

Antibiotics are widely used in in our daily life. In clinical, it is often used to treat the infections which are caused by bacteria, mycoplasma, chlamydia and other pathogens. Besides, it is also used in the research of science of life, agriculture, farming, animal husbandry and food industry. However, the abuse of antibiotics brings certain harms to both human health and the environment. The long-time intake of antibiotic (including clinical intake or food originated antibiotics residues) will lead to antibiotic accumulation in human body. When the concentration reaches to a certain value, it will result side effects to our hearing, kidney and immune system, cause allergy, and what's more, it even has the carcinogenic, teratogenic and mutagenic effects, it can also enhance the pathogens' drug resistance, and inhibit the growth rate of the intestinal flora.

In this assay, we did a policy research on antibiotic residues in animal origin food by analyzing the residue standards, banned substances and determine methods among CAC,EU, and American. Besides, we also made a comparison to the maximum residue limits of tetracycline between the standards in different countries and a prospect about food safety regulation system in China was also proposed.

2. Veterinary drug residue management system and related stan-dards

Since 1990, China government began to formulate technical regulations and standards related to veterinary drug residues, and set veterinary drug residues determination institutes. After the Melamine Milk Powder Scandal, China speeded up to build the food safety system and related standards of laws and regulations. Furthermore, it accelerated the issue process of the Food Safety Law.

2.1 Regulatory agency

Nowadays, China's food safety management system has been established and becomes to play more and more important roles. According to the Food Safety Law, the State Council established Food Safety Commission with the main responsibilities to analyze the situation of food in China, put forward important measures to food safety regulations and supervise the implementation of food safety regulatory. Ministry of Agriculture, National Department of Health and Family Planning Commission, State Administration for Quality Supervision and Inspection and Quarantine, State Food and Drug Administration, and Commerce Department are in charge of the central government level's food safety management.

2.2 Applicable laws and regulations

Now, an integrity food safety laws and regulations system was established based on 11 laws, including Food Safety Law of the People's Republic of China, a large number of technical standards related to animal source food safety demands and provincial and local food safety laws and regulations on animal source food safety regulation.

2.2.1 Law

At present, there are 11 laws which are suitable for our country's animal source food safety regulation, including the Food Safety Law of the People's Republic of China, Product Quality Law of the People's Republic of China, the Agriculture Law of the People's Republic of China, Standardization Law of the People's Republic of China, the Quarantine Law of the P.C.C., Concerning Import and Export Animals and Plants, Law of the People's Republic of China on Import and Export Commodity Inspection, the People's Republic of China on Animal Products, the Law of Produce Quality Safety of the People's Republic of China, Law of the People's Republic of China on Protection of Consumers' Rights and Interests, the Frontier Sanitation and Quarantine Law of the P.C.C., Fisheries Law of the Peoples Republic, and so on.

2.2.2 Regulations

The current existing laws and regulations related to the veterinary drug residue are mainly issued by the Ministry of Agriculture, including the followings: The prohibited catalog of the various kinds of drugs used in animal feed and drinking water, announced by Ministry of Agriculture, Ministry of Health, Food and Drug Administration. (People's Republic of China, Ministry of Agriculture Bulletin No. 176) Catalog of banned veterinary drugs and other compounds in food animals(People's Republic of China Ministry of Agriculture Bulletin No. 193) Local standards Abolished veterinary drugs catalog(People's Republic of China Ministry of Agriculture Bulletin No. 560) Prohibited substances used in animal feed and drinking water (People's Republic of China Ministry of Agriculture Bulletin No. 1519) Eliminated veterinary varieties catalog (People's Republic of China Ministry of Agriculture Bulletin No. 839) Animal maximum residue limits of veterinary drugs in food (People's Republic of China Ministry of Agriculture Bulletin No. 235) The detection method of Animal veterinary drug residues in food (People's Republic of China Ministry of Agriculture Bulletin No. 236)

2.3 banned substances

According to the Ministry of Agriculture No. 193, No. 235 and No. 560 announcement, veterinary drugs and other chemicals cannot be detected in food animal and animal source food.

2.3.1 Veterinary drug residue limits

In 2002, the Ministry of Agriculture's No. 235announcement published the "food animal veterinary drugs MRLs" standard. The main provisions was that 86 kinds of announcements do not need formulate pharmaceutical MRL standard, 99 species (including 551) approved animal foods , 9 kinds allow for therapeutic use. But veterinary drugs and 31 prohibited substances were banned and cannot be detected.

2.3.2 Determination methods

In 2003, the Ministry of Agriculture's No. 236 bulletin announced law of detection methods of animal veterinary drug residues in food. This bulletin provides the detection methods of 12 kinds of animal veterinary drug residues in food, including Carbadox label residue, nitroimidazole drug residues and et al., all of which are antibiotics (antibiotics), like carbadox label residue, nitroimidazoles, sulfamethazine, sodium lasalocid, enrofloxacin and ciprofloxacin, oxolinic acid and flumequine, penicillin, oxacillin, chloramphenicol. All the residues are detected by HPLC, with only one exception that penicillin residues in dairy is detected by microbial method.

2.4 Residue testing technology status and the relevant provisions

Now, China has more than 90 kinds of methods to determine the residue value of veterinary drugs which have limitation requirements, including using HPLC and vitamin method to determine Sulfonamides, tetracyclines, penicillins, instigate acid and trifluoromethanesulfonic chirp, enrofloxacin, ciprofloxacin and using LC -MS method to detect the Tetracycline drugs residue in pork and ELISA method to detect Tetracycline drugs residues in animal food.

3 EU

EU standards focus on the restrictions of veterinary drug residues and ensuring of food safety. Under the joint participation of Member States, (EU) No 37/2010 of veterinary drugs in animal foods MRLs Regulations were published and all member states should take this as the basis for regulations and better monitor the level of veterinary drug residues in foods of animal origin. The maximum residue limits (MRLs) set by (EU) No 37/2010 are the standard valve of controlling drug residues in animal origin foods. So far, the limitation set by (EU) No 37/2010 regulations is the alternative means and measures to control veterinary drugs residues.

3.1 Maximum residue limits of veterinary drugs

In January, 2010, EU published the Commission Regulation (EU) No 37/2010 ‘maximum residue limits and classification of pharmacologically active substance in animal origin food ', which collected the veterinary drug MRLs that EU has developed.

The regulations made a integration to the 4 appendix content in (EEC) No 2377/90 of. For easy use, all pharmacologically active substances are listed alphabetically in the same appendix, and the established two tables. The substances which are allowed to be used are in Table 1, with a total of 635 kinds; Table 2 includes banned substances with a total of 10 species.

So far (July 2015), the EU amended it 19 times, the amended content were pharmacologically active substances, including lasalocid, doxycycline and the others.

3.2 Prohibited substances regulations

In April 1996, EU issued Council Directive 96/22 / EC, this directive banned substances that cannot be used in animal husbandry, mainly with hormone, thyroid hormone action and β-doping substances.

In December 1999, the published Council resolution 1999/879 / EC prohibit the use of bovine growth hormone in dairy cows.

3.3 Determination methods

In order to reduce the mass sample analysis cost and improve efficiency of the analysis, the EU usually use both screening methods and confirmatory analysis method together to detect food safety which is the so called "two-step" strategy. At first, the large numbers of samples were rapidly analyzed by screening. If it was suspected as positive sample, then it will be confirmed by confirmation methods.

EU's main sample separation and purification techniques are solid phase extraction (SPE), supercritical fluid extraction, immune affinity chromatography (IAC) technology, matrix solid phase dispersion (MSPD) technology and so on. Veterinary drug residue detection methods are mainly divided into fast testing, routine quantitative detection and confirmation methods, including EIA, RIA, TLC, HPTLC, GC, HPLC, LC-MS, GC-MS and the like.

4. Horizontal comparison of tetracycline MRL

Tetracycline is a widely used broad-spectrum antibiotic and it is also the earliest antimicrobial drugs that attracted our attention in the food safety field. All countries have made relevant standards for tetracyclines residues.

Take the maximum residue levels of tetracycline in pork tissues as an example, in accordance with China's "veterinary drugs in animal foods MRLs", the maximum residue of single or composite tetracyclines in pork should not exceed 100mg / kg, namely 100000μg / kg. While The Codex Alimentarius Commission shall not exceed 200μg / kg, EU rules must not exceed 100μg / kg, and the FDA shall not exceed 2000μ / kg. The maximum residue limit of tetracycline on pork products in China is 50 times higher than the United States and is 500 times as CAC's requirement. what's more, it is 1,000 times higher as the EU countries limited. Then take the dairy oxytetracycline maximum residue levels as another example. In China, the maximum residue levels in dairy set by Ministry of

Agriculture was that a single or composite oxytetracycline drugs should not exceed 100mg / kg, namely 100000 μg / kg, while the Codex Alimentarius Commission is defined as not exceeding 100μg / l, EU rules should not exceed 100 μg / kg, and the US FDA shall not exceed 300μg / kg. In other words, the maximum dairy oxytetracycline residue limits in China is about 333 times the US FD regulations, and about 1000 times CAC and EU regulations.

5. Questions and suggestions

In China, the food safety management system is still bull management. When it needs to get to the certain place, there will be many other departments that will be involved in, even the business sector, public security, urban management and other regulatory authorities. The supervision department is not entitled to enforcement powers, the law enforcement is carried out by the police, urban management, business execution and the daily supervision and sample preparation is executed by the quality supervision department, the health sector which results in multiple prevarication.

Most of China's laws and regulations about veterinary drug residues are issued by the Ministry of Agriculture in the form of public announcement, instruction and the like. It is lack of unified planning and hard to conducive search. It suggested that unified management for veterinary drug residue limits should be executed as soon as possible, put all the files related in order and clean up all the old scattered regulations and standards to adapt to the China's existing laws and regulations, set the lateral MRLs of veterinary drugs in the form of national standards, so that the residue limits will be more unity, rationality and practicality.

The recent situation shows that parts of the standards' version are out of date with slower update time, less coverage and irrational maximum residue limits which differs from that of CAC and FDA to a great extent. So it proposed to speed up the standard scientific research, develop reasonable standards and improve the detection technology to catch up with the international standards as soon as possible.

The limited public profile is another problem. Nowadays, only the national compulsory standards are available for free download on the relevant state ministries and commissions' website and a large number of mandatory industry standards and local standards which are not covered by national standards can still not be obtained for free. It suggested that the related department shall refer to the data base of maximum residue limits of veterinary drugs which is established by Codex Alimentarius Commission, make it available to inquiries the substance by drug name, drug classes, animal species and target tissues and other to facilitate public use.

MRLs of veterinary drugs in different countries and international organizations have both similarities and its own characteristics. Companies or regulatory authorities should carry out comparative analysis based on the products situation and the requirements of the output or input country. For the veterinary that China has laxer limits, the regulation should be strengthened to ensure that export products meet the requirements of the importing countries and regions. For the veterinary drug that does not allow the use of, or the absence of residues in Chin's limited requirements which is striker than that of foreign country, we should focus on monitoring implementation in imported animal origin foods.

6. Appendix

Table.1 The approved prescribed MRLs of tetracycline in animal foods of China
Ministry of Agriculture Bulletin No. 235, "veterinary drugs in animal foods MRLs"

Drug Name	Residue marker	animal species	target tissue	MRL mg/kg
Oxytetracycline/Chlortetracycline/ Tetracycline ADI : 0-30	Parent drug, single or compound	All kinds of food animals Cattle/sheep Poultry Fish/shrimp	Muscle Liver Kidney Dairy Egg Meat	100 300 600 100 200 100

Table.2 EU regulations on animal foods tetracyclines residues (EU)No 37/2010

Pharmacologically active Substance	Marker residue	Animal Species	MRL	Target Tissues	Other Provisions	Therapeutic Classification
Pharmacologically active Substance	Marker residue	Animal Species	MRL	Target Tissues	(according to Article 14(7) of Regulation (EC) No 470/2009)	Therapeutic Classification
Chlortetracycline aureomycin	Sum of parent drug and its 4- epimer	All food-producing species	100 μg/kg	Muscle	For fin fish the muscle MRL relates to 'muscle and skin in natural proportions'. MRLs for liver and kidney do not apply to fin fish.	Anti-infectious agents/Antibiotics
			300 μg/kg	Liver
			600 μg/kg	Kidney
			100 μg/kg	Dairy
			200 μg/kg	Eggs
Oxytetracycline	Sum of parent drug and its 4- epimer	All food-producing species	100 μg/kg	Muscle	For fin fish the muscle MRL relates to 'muscle and skin in natural proportions'. [C1] and kidney do not apply to fin fish.	Anti-infectious agents/Antibiotics
			300 μg/kg	Liver
			600 μg/kg	Kidney
			100 μg/kg	Dairy
			200 μg/kg	Eggs
Tetracycline	Sum of parent drug and its 4- epimer	All food-producing species	100 μg/kg	Muscle	For fin fish the muscle MRL relates to 'muscle and skin in natural proportions'. MRLs for liver and kidney do not apply to fin fish.	Anti-infectious agents/Antibiotics
			300 μg/kg	Liver
			600 μg/kg	Kidney
			100 μg/kg	Dairy
			200 μg/kg	Eggs

Table.3 The relevant provisions of the United States on the residues of tetracycline residues in animal food 21 CFR part556

Pharmacologically active Substance	ADI	Marker residue	Animal Species	Toleran- ces	Target Tissues	21 CFR part 556 Number
Chlortetracycline	25 mg per kilogram of body weight per day.	sum of residues of the tetracyclines including chlortetracycline, oxytetracycline, and tetracycline	beef cattle, nonlactating dairy cows, calves, swine, sheep, chickens, turkeys, ducks	2 ppm	Muscle	§556.150
				6 ppm	Liver
				12 ppm	Kidney
				12 ppm	Fat
				0.4 ppm	Eggs
Oxytetracycline	25 mg per kilogram of body weight per day.	sum of residues of the tetracyclines including chlortetracycline, oxytetracycline, and tetracycline	beef cattle, dairy cattle, calves, swine, sheep, chickens, turkeys, finfish, and lobster	2 ppm	Muscle	§556.500
				6 ppm	Liver
				12 ppm	Kidney
				12 ppm	Fat
				0.3 ppm	Dairy
Tetracycline	25 mg per kilogram of body weight per day.	sum of residues of the tetracyclines including chlortetracycline, oxytetracycline, and tetracycline	calves, swine, sheep, chickens, and turkeys	2 ppm	Muscle	§556.720
				6 ppm	Liver
				12 ppm	Fat
				12 ppm	Kidney

Table.4 The tetracycline max residue in animal food by Codex Alimentarius Commission (CAC) CACMRL 2-2015

Species	Tissue	MRL (μg/kg)	CAC	Notes
CHLORTETRACYCLINE/OXYTETRACYCLINE/TETRACYCLINE (antimicrobial agent) JECFA Evaluation: 45 (1995); 47 (1996); 50 (1998); 58 (2002) Acceptable Daily Intake : 0-30 μg/kg body weight (50th JECFA, 1998). Group ADI for chlortetracycline, oxytetracycline and tetracycline. Residue Definition : Parent drugs, singly or in combination.
Cattle	Muscle	200	26th (2003)
Cattle	Liver	600	26th (2003)
Cattle	Kidney	1200	26th (2003)
Cattle	Dairy (μg/l)	100	26th (2003)
Fish	Muscle	200	26th (2003)	Applies only to oxytetracycline.
Giant prawn (Paeneus monodon)	Muscle	200	26th (2003)	Applies only to oxytetracycline.
Pig	Muscle	200	26th (2003)
Pig	Liver	600	26th (2003)
Pig	Kidney	1200	26th (2003)
Poultry	Muscle	200	26th (2003)
Poultry	Liver	600	26th (2003)
Poultry	Kidney	1200	26th (2003)
Poultry	Eggs	400	26th (2003)
Sheep	Muscle	200	26th (2003)
Sheep	Liver	600	26th (2003)
Sheep	Kidney	1200	26th (2003)
Sheep	Dairy (μg/l)	100	26th (2003)

Team:Scut-Champion-Park/Practices/Policy