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Abstract In order to explore the resource utilization of the harmless treatment product （pork meat and bone meal， abbreviated PM） of pigs died of non-communicable diseases， the general nutritional components and amino acid composition of PM and fish meal were determined and compared. The results showed that the contents of moisture， crude protein， crude fat and ash in PM and fish meal were 3.25% and 8.92%， 66.65% and 66.67%， 13.52% and 8.23%， 18.25% and 21.50%， respectively. The contents of essential amino acids （EAA） in PM and fish meal were 19.94% and 22.35%， respectively. For PM and fish meal， the first limiting amino acid was Met （methionine） + Cys （cysteine）， and the second limiting amino acid was Lys （lysine）; their essential amino acid indexes （EAAI） were 66.60 and 77.04， respectively; and their delicious amino acid （DAA） contents were 26.89% and 23.15%， respectively. In summary， the meat and bone meal of pigs died of non-communicable diseases has the characteristics of high protein and low ash contents， and has certain development and utilization potential as a recycled waste resource， especially in aquatic feed to replace fish meal.
Key words Dead pigs from non-communicable diseases; Reuse of waste resources; Pork meat and bone meal; Fish meal; Nutritional content; Amino acid; Amino acid score （AAS）; Chemical score （CS）; Essential amino acid index （EAAI）; Delicious amino acids
Received： August 12， 2020 Accepted： September 29， 2020
Supported by Major Science and Technology Special Project in Hunan （2017NK1030）; Earmarked Fund for China Agriculture Research System （CARS-45-48）.
Xiaofei CHENG （1986-）， male， P. R. China， assistant research fellow， master， devoted to research about aquatic animal nutrition and feed.
*Corresponding author. E-mail： ryain1983@163.com; hnhhliliu@163.com.
With the rapid and intensive development of animal husbandry in China， the number of deaths of farmed animals due to diseases and other reasons during the breeding process is increasing. Taking the pig industry as an example， according to the statistics of the 2018 National Economic and Social Development Statistical Bulletin released by the National Bureau of Statistics， at the end of 2018， there were 428.17 million live pigs in China and 693.82 million live pigs for slaughter （except Hong Kong， Taiwan and Macau of China）[1]. The industry-recognized mortality rate of pig breeding is usually 3% to 12%[2-3]. According to the minimum mortality rate of 3%， more than 20 million pigs died in 2018. Therefore， the harmless treatment of animals died of illness in farms has become a hot research topic in related industries， and the research on resource utilization of the harmless treatment products of animals died of illness is an important part of the sustainable development of the industry. Chemical preparation method is one of the main methods of harmless treatment of dead animals. The product obtained from pigs died of non-communicable diseases through the chemical preparation method is sick pork meat and bone meal （referred to as "pork meat and bone meal"）， which has certain development and utilization potential in aquatic feed and special fur economic animal feed as an alternative protein source of fish meal. The harmless treatment and resource utilization of pigs died of illness can not only reduce environmental pollution caused by casually discarding sick and dead pigs， but also realize the reuse of waste resources， thereby reducing breeding costs and harmless treatment costs. At present， the research on meat and bone meal mainly focuses on the nutritional content[4-8]， safety[9-10] and the application in aquatic feed[11-13]， but there are few reports on the nutritional content of meat and bone meal from pigs died of illness[14].
In this study， with PM （pork meat and bone meal） and fish meal as the research objects， their general nutritional components and amino acid components were determined and compared， aiming to provide reference for the resource utilization of the pollution-free treatment products of sick and dead animals （pork meat and bone meal） and the development of fish meal alternative protein sources.
Materials and Methods
Sample source and pretreatment
PM， the harmless treatment product of pigs died of non-communicable diseases， was provided by a certain Ecological Environmental Protection Technology Co.， Ltd in Hunan Province. The fish meal used in the experiment was imported white fish meal from Peru， provided by a large feed company in Changde， Hunan. After being crushed by a small grinder， the materials were sieved with a 60-mesh sieve and stored at -20 ℃ for routine nutrient and amino acid composition determination and analysis.
Nutrient composition determination and amino acid evaluation
General nutrients determination methods[15]
① Moisture content， （105±2） ℃ atmospheric drying method （GB/T5009.3-2010 National Food Safety Standard： Determination of Moisture in Food）. ②Protein content， automatic Kjeldahl method （GB/ T5009.5-2010 National Food Safety Standard： Determination of Protein in Food）， using the Danish FOSS Kjeltec 8400 automatic nitrogen analyzer. ③ Crude fat content， Soxhlet extraction method （GB/T5009.6-2003 Determination of Fat in Food）， using the Danish FOSS Soxhlet extractor. ④ Ash content， 550 ℃ muffle furnace burning method （GB/T5009.4-2010 National Food Safety Standard： Determination of Ash in Food）. Amino acid determination method[16]
A certain amount of sample （0.5 g） was hydrolyzed with 6 mol/L hydrochloric acid at 110 ℃ for 22 h， followed by filtering and diluting to 50 ml. A certain amount of the filtrate （0.5 ml） was vacuum dried to a test sample， which was determined with an S-433D amino acid analyzer of Sykam company in Germany for the amino acid composition and content of the sample. According to the standard model of amino acid scoring recommended by the Food and Agriculture Organization of the United Nations/World Health Organization （FAO/WHO） in 1973 and the amino acid model of whole egg protein[17]， the amino acid score （AAS）， chemical score （ CS） and essential amino acid index （EAAI） were calculated according to following formulas[18-19]：
ASS=aaAAFAO/WHO; CS=aaAAEgg;
EAAI=n100AAE×100BBE×100CCE×…×100HHE.
Wherein aa is the amino acid content of the test sample， %; AAFAO/WHO is the content of the same amino acid in the FAO/WHO scoring standard model， mg/g （in N）; AAEgg is the content of the same amino acid in the whole egg protein， mg/g （in N）; n is the number of essential amino acids to be compared; A， B， C，…， H are the contents of essential amino acids in the sample， mg/g （in N， DM）; and AE， BE， CE，…， HE are the contents of the same essential amino acid in whole egg protein， mg/g （in N， DM）.
Amino acid content （mg/g， in N） = Amino acid content （%， DM）/Crude protein content （%， DM）×6.25×1 000
Data processing
The results were expressed as "mean ± standard deviation" （x±s）. The data was analyzed by sample t test in Excel 2007， and P<0.05 indicated a significant difference.
Results and Analysis
General nutrients of PM and fish meal
As can be seen from Table 1， the moisture contents of PM（pork meat and bone meal）and fish meal were 3.25% and 8.92%， respectively; the crude protein contents were 66.65% and 66.67%， respectively; the crude fat contents were 13.52% and 8.23%， respectively; and the ash contents were 18.25% and 21.58%， respectively. The moisture and ash contents of PM were significantly lower than fish meal （P<0.05）; the crude fat content was significantly higher than fish meal （P<0.05）; and there were no significant difference in crude protein content between the two group （P>0.05）. Therefore， PM in this study had the characteristics of high crude protein and crude fat contents， and low ash and moisture contents. Amino acid composition of PM and fish meal
It can be seen from Table 2 that 17 amino acids were detected in both PM and fish meal （tryptophan was not detected due to acid hydrolysis）. The total amino acid （TAA） contents were 60.60% and 57.82%， respectively; and the contents of 7 essential amino acids （EAA） were 19.94% and 22.35%， respectively; and the contents of 10 non-essential amino acids （NEAA） were 40.66% and 35.47%， respectively. Among the 17 amino acids of PM， glutamic acid had the highest content of 9.76%， followed by glycine （6.76%）， aspartic acid （5.49%）， leucine （5.30%）， alanine （4.88%）， proline （4.75%）， valine （3.56%）， arginine （3.01%）， lysine （2.70%）， phenylalanine （2.62%）， isoleucine （2.39%）， threonine （2.05%）， histidine （2.01%）， tyrosine （1.98%）， serine （1.72%） and methionine （1.32%）， and the cysteine content was the lowest at 0.30%. Among the 17 amino acids of fish meal， glutamic acid had the highest content of 8.92%， followed by aspartic acid （6.09%）， leucine （5.02%）， alanine （4.12%）， glycine （4.02%） ）， lysine （3.63%）， valine （3.47%）， isoleucine （2.88%）， arginine （2.87%）， threonine （2.77%）， proline （2.72%）， phenylalanine （2.62%）， serine （2.43%）， tyrosine （2.14%）， histidine （1.97%）， and methionine （1.93%）， and the cysteine content was the lowest at 0.19%. Thetotal delicious amino acid （DAA） contents in PM and fish meal were 26.89% and 23.15%， respectively; the EAA/TAA values were 32.90% and 38.65%， respectively; and the EAA/NEAA values were 49.04% and 63.01%， respectively.
Amino acid scores， chemical scores and essential amino acid indexes of PM and fish meal
According to the amino acid score （AAS） and chemical score （CS） results of PM and fish meal （Table 3）， it can be seen that PM had the highest scores of Phe+Tyr and Leu， respectively， with values of 1.14 and 0.93， and the lowest Met+Cys scores， with values of 0.69 and 0.39， respectively， followed by Lys， with values of 0.74 and 0.57， respectively. The AAS of fish meal showed that Phe+Tyr was the highest at 1.17， and Met+Cys was the lowest at 0.92， followed by Lys at 1.00. The CS of fish meal showed that Thr was the highest at 0.89， and Met+Cys was the lowest at 0.52， followed by Lys at 0.77. The first and second limiting amino acids of meat and bone meal from pigs died of illness were Met+Cys and Lys， respectively， which were the same as the limiting amino acids of fish meal. The essential amino acid index （EAAI） values of PM and fish meal were 66.60 and 77.04， respectively. Discussion
Different countries have different policies and regulations on the application of animal protein raw materials in aquatic feeds. In the 1980s， because of the widespread epidemic of mad cow disease in the EU， the use of animal protein raw materials was banned， and the non-ruminant animal-derived feed proteins were allowed to be used in aquatic feeds until 2013， but animal protein sources are still prohibited from circulating in the same animal. In China， meat and bone meal is widely used in non-homogeneous animal feed， and there have been a lot of research and application in aquatic feeds especially[20]， but there has always been controversy regarding the safety of the application of meat and bone meal from pigs died of illness. Although Nong Yifa [2005]No.25 Measures for Disposal of Animals with Unknown Causes of Death and Diseases （Trial）， GB16548-2006 Regulations for the Biosafety Treatment of Diseased Animals and Diseased Animal Products and Nong Yifa[2013]No.34 Technical Specification for Harmless Treatment of Sick and Dead Animals put forward clear requirements for the reporting， diagnosis and harmless treatment of sick and dead animals and animal products， there are no specific regulations on the use of by-products after harmless treatment of sick and dead animals. Therefore， this study used pigs died of non-communicable diseases as raw materials for the determination and analysis of nutritional components， and scientifically explored the resource reuse of the harmless treatment products of pigs died of illness.
Huang et al.[4] studied the nutritional components in 10 representative domestic meat and bone meal samples， and the results showed that the moisture， crude protein， crude fat and ash contents of domestic meat and bone meal were generally 6.09%-10.20%， 48.13%-54.93%， 4.99%-16.47%， and 17.76%-34.90%， respectively. The study on meat and bone meal produced in Australia and Uruguay by Zhang et al.[10] showed that the moisture， crude protein， crude fat and ash contents of imported meat and bone meal were generally 6.94%-7.24%， 48.16%-57.06%， 10.55%-13.93%， and 25.54%-38.57%， respectively. GB/T20193-2006 Bone Meal and Meat and Bone Meal for Feed stipulates that the moisture content of meat and bone meal is not higher than 10%; the crude fat content is not higher than 12%; and the crude protein contents of the first， second and third standards are lower than 50%， 45%， and 40%， respectively; and the ash contents of the first， second， and third standards are not higher than 33%， 38%， and 43%， respectively[21]. The moisture content of meat and bone meal affects its storage and quality to a certain extent， while in this study， the moisture and ash contents of the pork meat and bone meal from the harmless treatment of sick and dead pigs were 3.25% and 18.25%， respectively， which were significantly lower than those in fish meal， and are far lower than the moisture and ash contents of ordinary meat and bone meal. The crude protein content of pork bone meal in this study was equivalent to that of fish meal， and the values are much higher than that of ordinary meat and bone meal， and reach the national first-level meat and bone meal standard. The crude fat content of pork bone meal in this study was higher than that of fish meal， and is slightly higher than the upper limit （12%） specified in the national standard for meat and bone meal and is greatly different from the crude fat content of ordinary meat and bone meal. It may be related to the manufacturer， process， and raw material quality of the meat and bone meal， which makes the coefficient of variation of the crude fat content larger[4]. In this study， the meat and bone meal from harmless treatment of dead pig carcass that contained glutamic acid， glycine， and alanine more than fish meal， but its contents of as threonine， methionine， isoleucine and lysine， as well as its total content of essential amino acids， were lower than fish meal， which is basically consistent with the research results of Liu et al.[7]. According to the ideal model of FAO/WHO， the EAA/TAA of protein with good quality is about 40%， and the EAA/NEAA is above 60%[17]. The PM was slightly lower than the above index requirements， while the fish meal basically met the above index requirements， indicating that the amino acid nutritional value of meat and bone meal was slightly lower than that of fish meal. According to the China Feed Database （29th edition in 2018）[22]， the essential amino acid contents of soybean meal are： phenylalanine 2.21%， methionine 0.59%， lysine 2.68%， threonine 2.05%， and tryptophan 0.57%， leucine 3.35%， isoleucine 1.99%， and valine 2.09%. Compared with pork meat and bone meal in this study， the 7 essential amino acids in soybean meal are all lower than or equal to PM. It could be seen that the nutritional value of amino acids in pork meat and bone meal was not as good as fish meal， but it was better than soybean meal.
Conclusions
The harmless treatment product （pork meat and bone meal） of pigs died of non-communicable disease has the characteristics of high protein and low ash content， and has certain development and utilization potential as a recycled waste resource， especially in aquatic feed to replace fish meal.
References
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[2] HAN T， KANG WH， YUAN L， et al. Research progress in biosafety disposal of animals and resource utilization[J]. Chinese Journal of Animal Health Inspection， 2015， 32（2）： 47-49. （in Chinese）
[3] YANG JX， HUANG MM， QUAN Y， et al. Research and application of high-temperature biodegradation and harmless treatment technology for animals died of illness[J]. China Poultry， 2016， 38（8）： 1-4. （in Chinese）
[4] HUANG MQ， CHEN JW， SONG DJ. Comparative study on nutritional components of domestic meat and bone meal[J]. Feed Research， 2011（9）： 80-82. （in Chinese）
[5] XIAO X， JIN QZ. Research and application of meat and bone meal[J]. Cereal & Feed Industry， 2012（6）： 56-59. （in Chinese） [6] YU HR， ZHANG Q， TONG T， et al. Comparison of the nutritional components of three different sources of meat and bone meal [J]. Journal of Weifang University， 2014， 14（2）： 80-82. （in Chinese）
[7] LIU HY. Comparative analysis of nutritional components of fish meal and meat and bone meal[J]. Animals Breeding and Feed， 2014（5）： 34-36. （in Chinese）
[8] FENG Y， XU HJ， LI XG， et al. Quality control of meat meal and meat and bone meal and their application in fish production[J]. Siliao yu Xumu·Xinsiliao， 2014（3）： 25-27. （in Chinese）
[9] RAO ZH， WANG H， ZHANG S， et al. Detection of Salmonella in feed and its growth characteristics in meat and bone meal[J]. Quality And Safety Of Agro-Products， 2013（4）： 41-43. （in Chinese）
[10] ZHANG Q， YU HR， TONG T， et al. Comparative study on the safety of three different sources of meat and bone meal for aquatic products[J]. Feed Research， 2014（5）： 67-70. （in Chinese）
[11] LI H， FU WZ， SHAO QJ. The application of meat and bone meal in aquatic animal nutrition[J]. Hunan Siliao， 2012（3）： 26-28. （in Chinese）
[12] WANG YY， SHI Y， YANG YH， et al. Application of meat and bone meal in aquatic feed[J]. China Feed， 2012（2）： 32-35. （in Chinese）
[13] XU MS， LIANG YX， LIU ZP. The application status of meat and bone meal and the prospect of processing meat and bone meal from fur animal carcass[J]. Heilongjiang Animal Science and Veterinary Medicine， 2016（20）： 209-211. （in Chinese）
[14] YANG HJ， HAO LZ， LIU JX. The technology of biological degradation of case fatality pigs and application evaluation of its products[J]. Modern Journal of Animal Husbandry and Veterinary Medicine， 2015（8）： 57-60. （in Chinese）
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[17] FAO/WHO Ad Hoe Expert Committee. Energy and protein requirements[M]. Rome： FAO Nutrition Meeting Report Series， 1973： 52.
[18] PELLETT PL， YONG VR. Nutritional evaluation of protein foods[M]. Tokyo： The United National University Publishing Company， 1980： 26-29.
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[20] WANG YY， XU GC， NIE ZJ， et al. Research progress of animal protein sources in aquatic animal feed to replace fish meal[J]. Jiangsu Agricultural Sciences， 2019， 47（16）： 24-29. （in Chinese）
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Editor： Yingzhi GUANG Proofreader： Xinxiu ZHU
Key words Dead pigs from non-communicable diseases; Reuse of waste resources; Pork meat and bone meal; Fish meal; Nutritional content; Amino acid; Amino acid score （AAS）; Chemical score （CS）; Essential amino acid index （EAAI）; Delicious amino acids
Received： August 12， 2020 Accepted： September 29， 2020
Supported by Major Science and Technology Special Project in Hunan （2017NK1030）; Earmarked Fund for China Agriculture Research System （CARS-45-48）.
Xiaofei CHENG （1986-）， male， P. R. China， assistant research fellow， master， devoted to research about aquatic animal nutrition and feed.
*Corresponding author. E-mail： ryain1983@163.com; hnhhliliu@163.com.
With the rapid and intensive development of animal husbandry in China， the number of deaths of farmed animals due to diseases and other reasons during the breeding process is increasing. Taking the pig industry as an example， according to the statistics of the 2018 National Economic and Social Development Statistical Bulletin released by the National Bureau of Statistics， at the end of 2018， there were 428.17 million live pigs in China and 693.82 million live pigs for slaughter （except Hong Kong， Taiwan and Macau of China）[1]. The industry-recognized mortality rate of pig breeding is usually 3% to 12%[2-3]. According to the minimum mortality rate of 3%， more than 20 million pigs died in 2018. Therefore， the harmless treatment of animals died of illness in farms has become a hot research topic in related industries， and the research on resource utilization of the harmless treatment products of animals died of illness is an important part of the sustainable development of the industry. Chemical preparation method is one of the main methods of harmless treatment of dead animals. The product obtained from pigs died of non-communicable diseases through the chemical preparation method is sick pork meat and bone meal （referred to as "pork meat and bone meal"）， which has certain development and utilization potential in aquatic feed and special fur economic animal feed as an alternative protein source of fish meal. The harmless treatment and resource utilization of pigs died of illness can not only reduce environmental pollution caused by casually discarding sick and dead pigs， but also realize the reuse of waste resources， thereby reducing breeding costs and harmless treatment costs. At present， the research on meat and bone meal mainly focuses on the nutritional content[4-8]， safety[9-10] and the application in aquatic feed[11-13]， but there are few reports on the nutritional content of meat and bone meal from pigs died of illness[14].
In this study， with PM （pork meat and bone meal） and fish meal as the research objects， their general nutritional components and amino acid components were determined and compared， aiming to provide reference for the resource utilization of the pollution-free treatment products of sick and dead animals （pork meat and bone meal） and the development of fish meal alternative protein sources.
Materials and Methods
Sample source and pretreatment
PM， the harmless treatment product of pigs died of non-communicable diseases， was provided by a certain Ecological Environmental Protection Technology Co.， Ltd in Hunan Province. The fish meal used in the experiment was imported white fish meal from Peru， provided by a large feed company in Changde， Hunan. After being crushed by a small grinder， the materials were sieved with a 60-mesh sieve and stored at -20 ℃ for routine nutrient and amino acid composition determination and analysis.
Nutrient composition determination and amino acid evaluation
General nutrients determination methods[15]
① Moisture content， （105±2） ℃ atmospheric drying method （GB/T5009.3-2010 National Food Safety Standard： Determination of Moisture in Food）. ②Protein content， automatic Kjeldahl method （GB/ T5009.5-2010 National Food Safety Standard： Determination of Protein in Food）， using the Danish FOSS Kjeltec 8400 automatic nitrogen analyzer. ③ Crude fat content， Soxhlet extraction method （GB/T5009.6-2003 Determination of Fat in Food）， using the Danish FOSS Soxhlet extractor. ④ Ash content， 550 ℃ muffle furnace burning method （GB/T5009.4-2010 National Food Safety Standard： Determination of Ash in Food）. Amino acid determination method[16]
A certain amount of sample （0.5 g） was hydrolyzed with 6 mol/L hydrochloric acid at 110 ℃ for 22 h， followed by filtering and diluting to 50 ml. A certain amount of the filtrate （0.5 ml） was vacuum dried to a test sample， which was determined with an S-433D amino acid analyzer of Sykam company in Germany for the amino acid composition and content of the sample. According to the standard model of amino acid scoring recommended by the Food and Agriculture Organization of the United Nations/World Health Organization （FAO/WHO） in 1973 and the amino acid model of whole egg protein[17]， the amino acid score （AAS）， chemical score （ CS） and essential amino acid index （EAAI） were calculated according to following formulas[18-19]：
ASS=aaAAFAO/WHO; CS=aaAAEgg;
EAAI=n100AAE×100BBE×100CCE×…×100HHE.
Wherein aa is the amino acid content of the test sample， %; AAFAO/WHO is the content of the same amino acid in the FAO/WHO scoring standard model， mg/g （in N）; AAEgg is the content of the same amino acid in the whole egg protein， mg/g （in N）; n is the number of essential amino acids to be compared; A， B， C，…， H are the contents of essential amino acids in the sample， mg/g （in N， DM）; and AE， BE， CE，…， HE are the contents of the same essential amino acid in whole egg protein， mg/g （in N， DM）.
Amino acid content （mg/g， in N） = Amino acid content （%， DM）/Crude protein content （%， DM）×6.25×1 000
Data processing
The results were expressed as "mean ± standard deviation" （x±s）. The data was analyzed by sample t test in Excel 2007， and P<0.05 indicated a significant difference.
Results and Analysis
General nutrients of PM and fish meal
As can be seen from Table 1， the moisture contents of PM（pork meat and bone meal）and fish meal were 3.25% and 8.92%， respectively; the crude protein contents were 66.65% and 66.67%， respectively; the crude fat contents were 13.52% and 8.23%， respectively; and the ash contents were 18.25% and 21.58%， respectively. The moisture and ash contents of PM were significantly lower than fish meal （P<0.05）; the crude fat content was significantly higher than fish meal （P<0.05）; and there were no significant difference in crude protein content between the two group （P>0.05）. Therefore， PM in this study had the characteristics of high crude protein and crude fat contents， and low ash and moisture contents. Amino acid composition of PM and fish meal
It can be seen from Table 2 that 17 amino acids were detected in both PM and fish meal （tryptophan was not detected due to acid hydrolysis）. The total amino acid （TAA） contents were 60.60% and 57.82%， respectively; and the contents of 7 essential amino acids （EAA） were 19.94% and 22.35%， respectively; and the contents of 10 non-essential amino acids （NEAA） were 40.66% and 35.47%， respectively. Among the 17 amino acids of PM， glutamic acid had the highest content of 9.76%， followed by glycine （6.76%）， aspartic acid （5.49%）， leucine （5.30%）， alanine （4.88%）， proline （4.75%）， valine （3.56%）， arginine （3.01%）， lysine （2.70%）， phenylalanine （2.62%）， isoleucine （2.39%）， threonine （2.05%）， histidine （2.01%）， tyrosine （1.98%）， serine （1.72%） and methionine （1.32%）， and the cysteine content was the lowest at 0.30%. Among the 17 amino acids of fish meal， glutamic acid had the highest content of 8.92%， followed by aspartic acid （6.09%）， leucine （5.02%）， alanine （4.12%）， glycine （4.02%） ）， lysine （3.63%）， valine （3.47%）， isoleucine （2.88%）， arginine （2.87%）， threonine （2.77%）， proline （2.72%）， phenylalanine （2.62%）， serine （2.43%）， tyrosine （2.14%）， histidine （1.97%）， and methionine （1.93%）， and the cysteine content was the lowest at 0.19%. Thetotal delicious amino acid （DAA） contents in PM and fish meal were 26.89% and 23.15%， respectively; the EAA/TAA values were 32.90% and 38.65%， respectively; and the EAA/NEAA values were 49.04% and 63.01%， respectively.
Amino acid scores， chemical scores and essential amino acid indexes of PM and fish meal
According to the amino acid score （AAS） and chemical score （CS） results of PM and fish meal （Table 3）， it can be seen that PM had the highest scores of Phe+Tyr and Leu， respectively， with values of 1.14 and 0.93， and the lowest Met+Cys scores， with values of 0.69 and 0.39， respectively， followed by Lys， with values of 0.74 and 0.57， respectively. The AAS of fish meal showed that Phe+Tyr was the highest at 1.17， and Met+Cys was the lowest at 0.92， followed by Lys at 1.00. The CS of fish meal showed that Thr was the highest at 0.89， and Met+Cys was the lowest at 0.52， followed by Lys at 0.77. The first and second limiting amino acids of meat and bone meal from pigs died of illness were Met+Cys and Lys， respectively， which were the same as the limiting amino acids of fish meal. The essential amino acid index （EAAI） values of PM and fish meal were 66.60 and 77.04， respectively. Discussion
Different countries have different policies and regulations on the application of animal protein raw materials in aquatic feeds. In the 1980s， because of the widespread epidemic of mad cow disease in the EU， the use of animal protein raw materials was banned， and the non-ruminant animal-derived feed proteins were allowed to be used in aquatic feeds until 2013， but animal protein sources are still prohibited from circulating in the same animal. In China， meat and bone meal is widely used in non-homogeneous animal feed， and there have been a lot of research and application in aquatic feeds especially[20]， but there has always been controversy regarding the safety of the application of meat and bone meal from pigs died of illness. Although Nong Yifa [2005]No.25 Measures for Disposal of Animals with Unknown Causes of Death and Diseases （Trial）， GB16548-2006 Regulations for the Biosafety Treatment of Diseased Animals and Diseased Animal Products and Nong Yifa[2013]No.34 Technical Specification for Harmless Treatment of Sick and Dead Animals put forward clear requirements for the reporting， diagnosis and harmless treatment of sick and dead animals and animal products， there are no specific regulations on the use of by-products after harmless treatment of sick and dead animals. Therefore， this study used pigs died of non-communicable diseases as raw materials for the determination and analysis of nutritional components， and scientifically explored the resource reuse of the harmless treatment products of pigs died of illness.
Huang et al.[4] studied the nutritional components in 10 representative domestic meat and bone meal samples， and the results showed that the moisture， crude protein， crude fat and ash contents of domestic meat and bone meal were generally 6.09%-10.20%， 48.13%-54.93%， 4.99%-16.47%， and 17.76%-34.90%， respectively. The study on meat and bone meal produced in Australia and Uruguay by Zhang et al.[10] showed that the moisture， crude protein， crude fat and ash contents of imported meat and bone meal were generally 6.94%-7.24%， 48.16%-57.06%， 10.55%-13.93%， and 25.54%-38.57%， respectively. GB/T20193-2006 Bone Meal and Meat and Bone Meal for Feed stipulates that the moisture content of meat and bone meal is not higher than 10%; the crude fat content is not higher than 12%; and the crude protein contents of the first， second and third standards are lower than 50%， 45%， and 40%， respectively; and the ash contents of the first， second， and third standards are not higher than 33%， 38%， and 43%， respectively[21]. The moisture content of meat and bone meal affects its storage and quality to a certain extent， while in this study， the moisture and ash contents of the pork meat and bone meal from the harmless treatment of sick and dead pigs were 3.25% and 18.25%， respectively， which were significantly lower than those in fish meal， and are far lower than the moisture and ash contents of ordinary meat and bone meal. The crude protein content of pork bone meal in this study was equivalent to that of fish meal， and the values are much higher than that of ordinary meat and bone meal， and reach the national first-level meat and bone meal standard. The crude fat content of pork bone meal in this study was higher than that of fish meal， and is slightly higher than the upper limit （12%） specified in the national standard for meat and bone meal and is greatly different from the crude fat content of ordinary meat and bone meal. It may be related to the manufacturer， process， and raw material quality of the meat and bone meal， which makes the coefficient of variation of the crude fat content larger[4]. In this study， the meat and bone meal from harmless treatment of dead pig carcass that contained glutamic acid， glycine， and alanine more than fish meal， but its contents of as threonine， methionine， isoleucine and lysine， as well as its total content of essential amino acids， were lower than fish meal， which is basically consistent with the research results of Liu et al.[7]. According to the ideal model of FAO/WHO， the EAA/TAA of protein with good quality is about 40%， and the EAA/NEAA is above 60%[17]. The PM was slightly lower than the above index requirements， while the fish meal basically met the above index requirements， indicating that the amino acid nutritional value of meat and bone meal was slightly lower than that of fish meal. According to the China Feed Database （29th edition in 2018）[22]， the essential amino acid contents of soybean meal are： phenylalanine 2.21%， methionine 0.59%， lysine 2.68%， threonine 2.05%， and tryptophan 0.57%， leucine 3.35%， isoleucine 1.99%， and valine 2.09%. Compared with pork meat and bone meal in this study， the 7 essential amino acids in soybean meal are all lower than or equal to PM. It could be seen that the nutritional value of amino acids in pork meat and bone meal was not as good as fish meal， but it was better than soybean meal.
Conclusions
The harmless treatment product （pork meat and bone meal） of pigs died of non-communicable disease has the characteristics of high protein and low ash content， and has certain development and utilization potential as a recycled waste resource， especially in aquatic feed to replace fish meal.
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Editor： Yingzhi GUANG Proofreader： Xinxiu ZHU