İnsan İdrarında Bazı Tetrasiklinlerin Analizine Yönelik Yeşil RPLC Yaklaşımı

Öz

Gerçekleştirilen çalışmada geniş etki spektrumuna sahip tetrasiklin grubu bileşiklerden 4-epitetrasiklin, 4-epioksitetrasiklin ve tetrasiklinin katkılandırılmış insan idrarında tayini için organik çözücü olarak asetonitril, metanol gibi toksik organik çözücülerin yerine toksik olmayan etanolün kullanıldığı yeşil bir RPLC yöntem geliştirilmiştir. Sıvı kromatografik yöntem geliştirmede pH 2,5-7.0 aralığında bileşiklerin sıvı kromatografik davranışları sistematik olarak incelenmiş ve %10 (h/h) etanol içeren pH'sı 3,0 olan ve 37 oC kolon sıcaklığına sahip koşul optimum koşul olarak belirlenmiştir. Sonrasında geliştirilen yöntemin ICH yönergelerine göre validasyonu ve sağlamlık testleri gerçekleştirilmiştir. Validasyon ve sağlamlık testleri sonucunda geliştirilen yöntemin rutin analizlerinde kullanım için uygun olduğu belirlenmiştir. Geliştirilen yöntem 0,5-3,0 µg/mL aralığında doğrusaldır ve çalışılan bileşiklerin katkılandırılmış insan idrarından geri kazanım sonuçları %100'e yakın olarak bulunmuştur. Ayrıca bu çalışmada geliştirilen yöntemin yeşilliği Analitik Yeşillik Hesaplayıcı (AGREE) ve Yeşil Analitik Prosedür İndeksi (GAPI) programları kullanılarak değerlendirilmiştir.

Anahtar Kelimeler

• Tetrasiklinler
• yeşil RPLC
• metot geliştirme
• pKa

A Green RPLC Approach to Some Tetracyclines Analysis in Human Urine

Öz

In the study conducted, an environmentally friendly RPLC method using non-toxic ethanol instead of toxic organic solvents such as acetonitrile and methanol organic solvents was developed for the determination of 4-epitetracycline, 4-epioxytetracycline, and tetracycline, which are compounds of the tetracycline group with a broad spectrum of activity, in spiked human urine. In developing the liquid chromatographic method for the compounds studied, their liquid chromatographic behavior in the pH range of 2.5-7.0 was systematically investigated and the condition with 10% (v/v) ethanol in its structure, pH 3.0, and a column temperature of 37 oC was determined to be the optimum condition. Validation and robustness tests of the developed method were then carried out by the ICH guidelines. As a result of the validation and robustness tests, it was determined that the developed method is suitable for use in routine analyses. The developed method is linear in the range of 0.5-3.0 µg.mL-1 and the recovery results of the investigated compounds from spiked human urine were close to 100%. In addition, the greenness of the method developed in this study was evaluated using the Analytical Greenness Metric Approach (AGREE) and Green Analytical Procedures Index (GAPI) programs.

Anahtar Kelimeler

• Tetracyclines
• green RPLC
• method development
• pKa

Kaynakça

1. 1. Gabler WL. Fluxes and accumulation of tetracyclines by human blood cells. Res Commun Chem Pathol Pharmacol. 1991;72(1):39–51.
2. 2. Fritz JW, Zuo Y. Simultaneous determination of tetracycline, oxytetracycline, and 4-epitetracycline in milk by high-performance liquid chromatography. Food Chem. 2007;105:1297–301.
3. 3. Wua X, Wei Y, Zheng J, Zhao X, Zhong W. The behavior of tetracyclines and their degradation products during swine manure composting. Bioresour Technol. 2011;102:5924–31.
4. 4. Yu H, Tao Y, Chen D, Wang Y, Yuan Z. Development of an HPLC–UV method for the simultaneous determination of tetracyclines in muscle and liver of porcine, chicken and bovine with accelerated solvent extraction. Food Chem. 2011;124:1131–8.
5. 5. Anderson CR, Rupp HS, Wu WH. Complexities in tetracycline analysis-chemistry, matrix extraction, cleanup, and liquid chromatography. J Chromatogr A. 2005;1075:23–32.
6. 6. Halling-Sørensen B, Sengeløv BG, Tjørnelund J. Toxicity of tetracyclines and tetracycline degradation products to environmentally relevant bacteria, including selected tetracycline-resistant bacteria. Arch Environ Contam Toxicol. 2002;42:263–71.
7. 7. Gurgel JVM, Effting L, Galvan D, Nazário CED. QuEChERS method for simultaneous analysis of tetracyclines from chicken by HPLC–DAD. Food Anal Methods. 2024;17:1292–301.
8. 8. Butovskaya E, Carrillo Heredero AM, Segato G, Faggionato E, Borgia M, Marchis D, et al. Quantitative determination of tetracyclines in medicated feed for food-producing animals by HPLC–DAD. Food Addit Contam Part A. 2024;41(6):601–9.

9. 9. Zeb S, Wong A, Khan S, Hussain S, Sotomayor MDPT. Synthesis and characterization of a magnetic molecular imprinted polymer for tetracycline quantification in food samples. Chem Select. 2024;9(19):e202400346.
10. 10. Nakhonchai N, Prompila N, Ponhong K, Siriangkhawut W, Vichapong J, Supharoek S. Green hairy basil seed mucilage biosorbent for dispersive solid-phase extraction enrichment of tetracyclines in bovine milk samples followed by HPLC analysis. Talanta. 2024;271:1–11.
11. 11. Uddin MN, Hasnaine A, Sajedul Haque M, Das S, Uddin M, Chakraborty D, et al. Development of an HPLC-PDA method for the simultaneous estimation of three antibiotics in pharmaceutical formulations and bovine milk and health risk assessment. J Food Prot. 2024;87:1–7.
12. 12. Demiralay EÇ. An experimental design approach to optimization of the liquid chromatographic separation conditions for the determination of metformin and glibenclamide in pharmaceutical formulation. Acta Chim Slov. 2012;59(2):307–14.
13. 13. Gündoğan B, Demiralay EÇ, Daldal YD, Üstün Z. Determination of optimum separation condition for some carbapenem antibiotics using RPLC with the aid of central composite design and desirability function. Curr Pharm Anal. 2017;13:100–9.
14. 14. Uysal R, Daldal YD, Üstün Z, Demiralay EÇ. Optimization of RPLC method for separation of some acetylcholinesterase inhibitors by using central composite design. Eurasian J Anal Chem. 2017;2(1):23–43.
15. 15. Daldal YD. İnsan idrarında valasiklovirin tayini için RPLC yöntemin geliştirilmesi ve validasyonu. Süleyman Demirel Üniversitesi Sağlık Bilimleri Dergisi. 2023;14(2):282–91.
16. 16. International Conference on Harmonization. ICH validation of analytical procedures, text and methodology Q2 (R1). 2005.
17. 17. Kazakevich Y, Lobrutto Y. HPLC for pharmaceutical scientists. 1st ed. Wiley-Interscience; 2007. p. 1135.
18. 18. Konçe İ, Demiralay Çubuk E, Yılmaz Ortak H. Chromatographic determination of thermodynamic acid dissociation constants of tetracycline antibiotics and their epimers. J Chromatogr Sci. 2019;57(8):745–50.
19. 19. Çubuk Demiralay E, Koç D, Daldal YD, Alsancak G, Ozkan SA. Determination of chromatographic dissociation constants of some carbapenem group antibiotics and quantification of these compounds in human urine. Biomed Chromatogr. 2014;28(5):660–6.
20. 20. Płotka-Wasylka J. A new tool for the evaluation of the analytical procedure: Green Analytical Procedure Index. Talanta. 2018;181:204–9.
21. 21. Pena-Pereira F, Wojnowski W, Tobiszewski M. AGREE-Analytical GREEnness Metric Approach and Software. Anal Chem. 2020;92:10076–82.
22. 22. Kuzucanli E, Çubuk Demiralay E, Daldal YD, Üstün Z, Konçe İ, Alsancak G. Development and validation of RPLC method for the simultaneous analysis of ACE inhibitors in tablet formulations. Turk J Anal Chem. 2022;4(2):103–10.