Detection of cow milk in sheep and goat milk by TaqMan Real-Time PCR

Abstract

Milk and dairy products constitute an important part of the daily diet due to their high nutritional value. However, in addition to these important features, it is among the most adulterated foods. The most common adulteration in milk and dairy products is the mixing of cow's milk with sheep and goat milk and products. This situation causes socio-economic potential risks that are not preferred by consumers. In this study, it was aimed to determine the amount of cow's milk mixed with sheep and goat milk at different rates by TaqMan Real-Time PCR. For this purpose, 1%, 5%, 10%, 25%, 75% and 90% cow's milk was mixed with goat and sheep milk. In addition, the sensitivity of the PCR process was investigated by diluting the DNA obtained from pure cow's milk. As a result of this study, the presence of 1% cow's milk and 0.003 ng DNA mixed with sheep and goat milk was determined. As a result of the research, it is thought that TaqMan Real-Time PCR can be used as a reliable and sensitive method for detecting low level cow's milk mixed with sheep and goat milk.

Keywords

• cow milk
• goat milk
• sheep milk
• TaqMan Real-Time PCR
• authentication

Koyun ve keçi sütlerinde inek sütünün TaqMan Real-Time PCR ile tespit edilmesi

Öz

Süt ve süt ürünleri içerdikleri yüksek besin değeriyle günlük diyetin önemli bir parçasını oluşturmaktadır. Ancak bu önemli özelliklerinin yanı sıra en fazla hile yapılan gıdalar arasında yer almaktadır. Koyun-keçi sütü ve ürünlerine inek sütünün karıştırılması süt ve süt ürünlerinde en sık karşılaşılan hilelerin başında gelmektedir. Bu durum, tüketiciler tarafından tercih edilmeyen sosyo-ekonomik potansiyel risklere neden olmaktadır. Bu çalışmada koyun ve keçi sütlerine farklı oranlarda karıştırılan inek sütü miktarının TaqMan Real-Time PCR ile tespit edilmesi amaçlanmıştır. Bu amaçla %1, %5, %10, %25, %75 ve %90 oranlarında keçi ve koyun sütlerine inek sütü karıştırılmıştır. Ayrıca saf inek sütünden elde edilen DNA sulandırılarak PCR işleminin duyarlılığı araştırılmıştır. Bu çalışmanın sonucunda, koyun ve keçi sütlerine karıştırılan %1 inek sütü ve 0,003 ng DNA varlığı tespit edilmiştir. Araştırma sonucunda TaqMan Real-Time PCR’ın koyun ve keçi sütlerine karıştırılan düşük düzeydeki inek sütünün tespit edilmesinde güvenilir ve hassas bir yöntem olarak kullanılabileceği düşünülmektedir.

Anahtar Kelimeler

• inek sütü
• keçi sütü
• koyun sütü
• TaqMan Real-Time PCR
• tür tespiti

Kaynakça

1. Qin C, Liu L, Wang Y, Leng T, Zhu M, Gan B ve ark. Advancement of omics techniques for chemical profile analysis and authentication of milk. Trends Food Sci Technol 2022;127:114-128
2. Grant C, Rotherham B, Sharpe S, Scragg R, Thompson J, Andrews J ve ark. Randomized, double-blind comparison of growth in infants receiving goat milk formula versus cow milk infant formula. J Paediatr Child Health 2005;41(11):564–568.
3. Lad SS, Aparnathi K, Mehta B, Velpula S. Goat milk in human nutrition and health–a review. Int J Curr Microbiol Appl Sci 2017;6(5):1781–1792.
4. Guo MR, Dixon PH, Park YW, Gilmore JA, Kindstedt PS. Seasonal changes in the chemical composition of commingled goat milk. J Dairy Sci 2001;84:E79–E83.
5. Li Q, Zhao Y, Zhu D, Pang X, Liu Y, Frew R ve ark. Lipidomics profiling of goat milk, soymilk and bovine milk by UPLC-Q-exactive orbitrap mass spectrometry. Food Chem 2017;224:302–309.
6. Da Paixao Teixeira JL, Dos Santos Carames ET, Baptista DP, Gigante ML, Pallone JAL. Vibrational spectroscopy and chemometrics tools for authenticity and improvement the safety control in goat milk. Food Control 2020;112:107105.
7. Dabrowska A, Walecka E, Bania J, Zelazko M, Szoltysik M, Chrzanowska J. Quality of UHT goat’s milk in Poland evaluated by real-time PCR. Small Rumin Res 2010;94(1–3):32–37.
8. Azad T, Ahmed S. Common milk adulteration and their detection techniques. Int J Food Contam 2016;3(1): 22.

9. Teixeira J, Carames E, Baptista DP, Gigante ML, Pallone J. Rapid adulteration detection of yogurt and cheese made from goat milk by vibrational spectroscopy and chemometric tools. J Food Compos Anal 2021;96:103712.
10. Pinto PA, Anconi ACSA, de Abreu LR, Magalhães EJ, Nunes CA. Strategies to determine lactose in cow milk by mid infrared spectroscopy. J Food Compos Anal 2021;104:104176.
11. Mafra I, Roxo Á, Ferreira IM, Oliveira MBP. A duplex polymerase chain reaction for the quantitative detection of cows’ milk in goats’ milk cheese. Int Dairy J 2007;17(9):1132-1138.
12. Silanikove N, Leitner G, Merin U, Prosser CG. Recent advances in exploiting goat's milk: quality, safety and production aspects. Small Rumin Res 2010;89(2-3):110-124.
13. García V, Rovira S, Boutoial K, López MB. Improvements in goat milk quality: A review. Small Rumin Res 2014;121(1):51–57.
14. Di Pinto A, Terio V, Marchetti P, Bottaro M, Mottola A, Bozzo G ve ark. DNA-based approach for species identification of goat-milk products. Food Chem 2017;229:93-97.
15. Li Q, Yu Z, Zhu D, Meng X, Pang X, Liu Y ve ark. The application of NMR‐based milk metabolite analysis in milk authenticity identification. J Sci Food Agric 2017;97(9):2875-2882.
16. Balthazar CF, Pimentel TC, Ferrão LL, Almada CN, Santillo A, Albenzio M ve ark. Sheep milk: physicochemical characteristics and relevance for functional food development. Compr Rev Food Sci Food Saf 2017;16(2):247– 262.
17. Tribst AAL, Falcade LTP, Leite Júnior BRDC, De Oliveira MM. Why are most physicochemical parameters not useful in predicting the quality of sheep milk? Int J Dairy Technol 2020;73(1):292– 295.
18. Valizadeh Yonjalli R, Mirzaei Aghjehgheshlagh F, Mahdavi A, Navidshad B, Staji H. The effects of tannin extract and linseed oil on yield, physicochemical characteristics and fatty acid profile of ewe milk. Int J Dairy Technol 2020;73(4):656-666.
19. Wang Z, Li T, Yu W, Qiao L, Liu R, Li S ve ark. Determination of content of camel milk in adulterated milk samples by normalized real-time polymerase chain reaction system based on single-copy nuclear genes. J Sci Food Agric 2020;100(8):3465–3470.
20. Murphy AM, Shariflou MR, Moran C. High quality genomic DNA extraction from large milk samples. J Dairy Res 2002;69(4):645– 649.
21. Souza SS, Cruz AG, Walter EHM, Faria JAF, Celeghini RMS, Ferreira MMC ve ark. Monitoring the authenticity of Brazilian UHT milk: a chemometric approach. Food Chem 2011;124(2):692e695.
22. Moore JC, Spink J, Lipp M. Development and application of a database of food ingredient fraud and economically motivated adulteration from 1980 to 2010. J Food Sci 2012;77(4):R118-R126.
23. Kamal M, Karoui R. Analytical methods coupled with chemometric tools for determining the authenticity and detecting the adulteration of dairy products: A review. Trends Food Sci Technol 2015;46(1):27-48.
24. Golinelli LP, Carvalho AC, Casaes RS, Lopes CSC, Deliza R, Paschoalin VMF ve ark. Sensory analysis and species-specific PCR detect bovine milk adulteration of frescal (fresh) goat cheese. J Dairy Sci 2014;97(11):6693-6699.
25. Keyvan E, İplikçioğlu Çil G, Çınar Kul B, Bilgen N, Şireli UT. Identification of meat species in different types of meat products by PCR. Ankara Univ Vet Fak Derg 2017;64(4):261-266.
26. Agrimonti C, Pirondini A, Marmiroli M, Marmiroli N. A quadruplex PCR (qxPCR) assay for adulteration in dairy products. Food Chem 2015;187:58-64.
27. Guo L, Qian JP, Guo YS, Hai X, Liu GQ, Luo JX ve ark. Simultaneous identification of bovine and equine DNA in milks and dairy products inferred from triplex TaqMan real-time PCR technique. J Dairy Sci 2018;101(8):6776-6786.
28. Guo L, Ya M, Hai X, Guo YS, Li CD, Xu WL ve ark. A simultaneous triplex TaqMan real-time PCR approach for authentication of caprine and bovine meat, milk and cheese. Int Dairy J 2019;95:58-64.
29. Snirc M, Fekete T, Belej L, Židek R, Golian J, Haščík P ve ark. Detection of ovine milk adulteration using TaqMan real-time PCR assay. Potr S J F Sci 2017;11(1):338-343.
30. Fekete T, Šnirc M, Belej Ľ, Židek R, Golian J, Haščík P ve ark. Authentication of caprine milk and cheese by commercial qPCR assay. Potr S J F Sci 2017;11(1):580-586.
31. Deng L, Li A, Gao Y, Shen T, Yue H, Miao J ve ark. Detection of the bovine milk adulterated in camel, horse, and goat milk using duplex PCR. Food Anal Methods 2020;13(2):560-567.
32. Tortorici L, Di Gerlando R, Tolone M, Mastrangelo S, Sardina MT. 12S rRNA mitochondrial gene as marker to trace Sicilian mono-species dairy products. Livest Sci 2016;193:39-44.
33. Mayer HK, Bürger J, Kaar N. Quantification of cow’s milk percentage in dairy products–A myth? Anal Bioanal Chem 2012;403(10):3031–3040.