Glüten Analizinde HPLC, LC-MS/MS Yöntemlerinin ELISA ile Karşılaştırılması

Year 2020, Issue: 24, 30 - 54, 30.08.2020

Ali Özcan İsmail Azar Arzu Yavuz Hakan Yavaş Emre Tokat Vesile Çetin

Abstract

Öz

Amaç: Tahıl ve ürünleri içerdikleri Glüten nedeni ile bazı insanlarda rahatsızlıklara neden olabilmektedirler. Bu hastalıklardan biri olan çölyak, buğday, çavdar, arpa ve bazen de yulaf ürünlerinin tüketimi sonucu bağırsakta ortaya çıkan bir hastalıktır. Hastalığın nedenini oluşturan temel etken glüten proteininin gliadin adlı alt
fraksiyonudur. Hastalığa sebep olan glüten fraksiyonları glütenin ve gliadinlerdir. Glütenin kendi içerisinde ω5, LMW ve HMW olmak üzere üç alt fraksiyona ayrılmaktadır. Gliadinler ise α(Alfa), ω(Omega), β(Beta) ve γ(Gamma) olmak üzere dört fraksiyondan oluşmaktadır.

Materyal ve Yöntem: Bu çalışmada çölyak hastalarına özel olarak üretilen gıdaların analizlerinde kullanılabilecek ve yasal limitler olan 0-20 mg/kg ile 20-80 mg/kg aralıklarında LOD değerine sahip kromatografik bir yöntem geliştirilmeye çalışılmıştır.

Bulgular ve Sonuç: Yapılan HPLC çalışmalarında LOD değeri 31,94mg/kg, LOQ değeri ise 106,47mg/kg düzeylerinde metot performansları elde edilmiştir. LC MS/MS çalışmalarında elde edilen LOD değeri 19,24 mg/kg, LOQ değeri ise 64,13 mg/kg olarak tespit edilmiştir.

Keywords

Çölyak Hastalığı,, ELISA,, HPLC, Glüten, LC-MS/MS, Deliac Disase,

Comparison of HPLC, LC-MS / MS Methods with ELISA in Gluten Analysis

Abstract

Abstract

Objective: Cereals and products may cause disorder in some people due to the Gluten they contain. Celiac, one of these diseases, is a disease that occurs in the intestine as a result of consumption of wheat, rye, barley and sometimes oat products. The main cause of the disease is the sub-fraction of gluten protein called gliadin. The gluten fractions that cause disease are glüten and gliadins. Gluten is divided into three sub-fractions: ω5, LMW and HMW. Gliadins consist of four fractions: α (Alpha), ω (Omega), β (Beta) and γ (Gamma).

Marerials and Methods: In this study, it was tried to develop a chromatographic method that can be used in the analysis of foods specially produced for celiac patients and has an LOD value between 0-20 mg / kg and 20-80 mg / kg, which are legal limits.

Result and Conclusion: In the HPLC studies, the method performances were obtained at the LOD value of 31,94 mg/kg and the LOQ value at 106,47 mg/kg. LOD value obtained in LC MS/MS studies was determined 19,24mg / kg and LOQ value 64,13 mg / kg.

Keywords

Celiac disase,, Gluten, HPLC, ELISA, LC-MS/MS

References

• Anonim, 2012a.EURACHEM / CITAC Guide CG 4 Quantifying Uncertainty in Analytical Measurement, QUAM:2012.P1
• Anonim, 2012b. FAPAS, (27/12/2012). Available Quality Control Materials <http://www.fapas. com/quality-control-materials/Available-qualitycontrol- materials.cfm>
• Anonim, 2017. ISO 21748:2017 Guidance for the Use of Repeatability, Reproducibility and Trueness Estimates in Measurement Uncertainty Evaluation
• Anonim, 2019. ISO 5725-2:2019 Accuracy (Trueness and Precision) of Measurement Methods and Results—Part 2: Basic Method for the Determination of Repeatability and Reproducibility of a Standard Measurement Method
• Battais, F., Courcoux, P., Popineau, Y., Kanny, G., Moneret-Vautrin, D. A. and Denery-Papini, S., 2005. Food Allergy to Wheat: Differences in Immunoglobulin E-Binding Proteins as a Function of Age or Symptoms. Journal of Cereal Science, 42(1), 109-117.
• Ciclitira, P. J., Ellis, H. J. and Lundin, K. E., 2005. Glüten-Free Diet—What is Toxic?. Best Practice and Research Clinical Gastroenterology, 19(3), 359-371.
• Farrell, R. J. and Kelly, C.P., 2002. Celiac Sprue. New England Journal of Medicine, 346(3), 180-188.
• Goesaert, H., Brijs, K., Veraverbeke, W.S., Courtin, C.M., Gebruers, K. and Delcour, J.A., 2005. Wheat Flour Constituents: How They Impact Bread Quality, and How to Impact Their Functionality. Trends in food science & technology, 16(1-3), 12-30.
• Gómez, A.V., Ferrer, E., Añón, M.C. and Puppo, M.C., 2012. Analysis of Soluble Proteins/Aggregates Derived From Glüten-Emulsifiers Systems. Food Research International, 46(1), 62-68.
• Hopper, A., Hadjivassiliou, M., Butt, S. and Sanders Ds., 2007. Adult Coeliac Disease. BMJ; 335: 558- 562. L5-29
• Kieffer, R., Schurer, F., Köhler, P. and Wieser, H., 2007. Effect of Hydrostatic Pressure and Temperature on the chemical and Functional Properties of wheat Glüten: Studies on Glüten, Gliadin and Glütenin. Journal of Cereal Science, 45(3), 285-292.
• Maki, M, and Lohi, O., 2004. Celiac Disease. In: Walker WA, Goulet O, Kleinman RE, Sherman PM, Shneider BL, Sanderson IR (eds). Pediatric Gastrointestinal Disease. 4th ed. Ontario: B.C. Decker,: 932-43.
• Mowat, A.M., 2003. Coeliac Disease – a Meeting Point for Genetics, Immunology and Protein Chemistry. The Lancet; 361:1290-1292
• Niewinski, M.M., 2008. Advances in Celiac Disease and Glüten-Free Diet. Journal of the American Dietetic Association, 108(4), 661-672.
• Özkaya, B., 1999. Tahılların Neden Olduğu Alerjiler Ve Önemi-2. Food Hi-Tech, Mar. 82-88.
• Sealey-Voyksner, J. A., Khosla, C., Voyksner, R.D. and Jorgenson, J.W., 2010. Novel Aspects of Quantitation of Immunogenic Wheat Gluten Peptides by Liquid Chromatography – Mass Spectrometry/Mass Spectrometry. Journal of Chromatography A, 1217(25), 4167-4183.
• Sollid, L.M. and Jabri, B., 2005. Is Celiac Disease an Autoimmune Disorder?. Current opinion in immunology, 17(6), 595-600.
• Urgancı, N., 2005. Çölyak Hastalarına Ekmek Zehir Oluyor . http://212.174.4 6 .1 4 9 / w /dergi/basinpdf/kasim2004/18_19_20.pdf,
• Van Eckert, R., Berghofer, E., Ciclitira, P.J., Chirdo, F., Denery-Papini, S., Ellis, H. J. and Mendez, E., 2 0 0 6 . To wards a New Gliadin Reference Material–Isolation and Characterisation. Journal of cereal science, 43(3), 331-341.