METODE HPLC-DAD UNTUK ANALISIS GLUKOSAMIN PADA TERIPANG KERING DENGAN DERIVATISASI PREKOLOM (HPLC-DAD Method for Glucosamine Analysis in Dried Sea Cucumber with Precolumn Derivatization)

Dillani Putri Ramadhaningtyas; Andreas Andreas; Dian Muzdalifah; Hafiizh Prasetia; Hikmat Hikmat; Novita Ariani; Hendris Hendarsyah Kurniawan; Aprilia Nur Tasfiyati

doi:10.36974/jbi.v11i2.6394

METODE HPLC-DAD UNTUK ANALISIS GLUKOSAMIN PADA TERIPANG KERING DENGAN DERIVATISASI PREKOLOM (HPLC-DAD Method for Glucosamine Analysis in Dried Sea Cucumber with Precolumn Derivatization)

Dillani Putri Ramadhaningtyas, Andreas Andreas, Dian Muzdalifah, Hafiizh Prasetia, Hikmat Hikmat, Novita Ariani, Hendris Hendarsyah Kurniawan, Aprilia Nur Tasfiyati

Abstract

Glucosamine is known to be a supplement for treating osteoarthritis and joint pain. Recently, Indonesian coastal communities have developed sea cucumber enriched in glucosamine as an alternative to crustaceans, a widely consumed-glucosamine source. The aim of this study is to develop a high performance liquid chromatography tandem with diode array detector (HPLC-DAD) method for the determination of glucosamine in sea cucumber products was developed and validated. Glucosamine were extracted from dried sea cucumber samples through hydrolysis using hydrochloric acid 5 M, followed by derivatization using Fmoc-Cl (9-Fluorenylmethyl chloroformate) and borate buffer before subjected to HPLC-DAD. All preparation steps for samples and standards were performed using the gravimetric dilution method. The reversed-phase chromatographic separation was conducted in the C18 column by applying a gradient elution of water and acetonitrile. The results showed good linearity with coefficient determination at 0.999 in the range concentration of 1-220 mg/kg. The LOD and LOQ were found to be 0.16 and 0.53 mg/kg respectively, while the recoveries were in the range of 98-99%. The intraday precision values were lower than 2% for four different spiked concentrations (10, 60, 100, and 200 mg/kg). The method then successfully applied to analyze glucosamine in twenty local dried sea cucumber samples. The results showed that eleven of samples contain glucosamine in the range of 100 to 2700 mg/kg.

Keywords: fmoc-cl, glucosamine, hplc-dad, sea cucumber

ABSTRAK

Glukosamin dikenal sebagai suplemen untuk mengobati osteoartritis dan nyeri sendi. Baru-baru ini masyarakat pesisir Indonesia telah mengembangkan teripang yang diperkaya glukosamin sebagai alternatif pengganti krustasea yang merupakan sumber glukosamin yang banyak dikonsumsi. Tujuan dari penelitian ini adalah melakukan pengembangan dan validasi metode high performance liquid chromatography dengan diode array detector (HPLC-DAD) untuk penentuan glukosamin pada produk teripang. Glukosamin diekstraksi dari sampel teripang kering melalui hidrolisis menggunakan asam klorida 5 M, dilanjutkan dengan derivatisasi menggunakan larutan Fmoc-Cl (9-Fluorenylmethyl chloroformate) dan buffer borat sebelum dilakukan analisis menggunakan HPLC-DAD. Semua langkah preparasi sampel dan standar dilakukan dengan menggunakan metode pengenceran gravimetri. Pemisahan kromatografi fase terbalik dilakukan di kolom C18 dengan menerapkan elusi gradien menggunakan air dan asetonitril sebagai fasa gerak. Hasil penelitian menunjukkan linieritas yang baik dengan koefisien determinasi sebesar 0,999 pada rentang konsentrasi 1 sampai 220 mg/kg. Metode ini memiliki LOD dan LOQ masing-masing 0,16 dan 0,53 mg/kg, sedangkan nilai perolehan kembali berada di kisaran 98-99%. Nilai presisi intraday ditemukan lebih rendah dari 2% untuk empat konsentrasi spiking yang berbeda (10, 60, 100 dan 200 mg/kg). Metode ini kemudian berhasil diterapkan untuk menganalisis glukosamin pada dua puluh sampel teripang kering lokal. Hasilnya menunjukkan bahwa sebelas sampel mengandung glukosamin dalam rentang 100 hingga 2700 mg/kg.

Kata kunci: fmoc-cl, glukosamin, hplc-dad, teripang

Keywords

Fmoc-Cl; Glucosamine; HPLC-DAD; Sea cucumber

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References

AOAC. (2016). Appendix F: Guidelines for Standard Method Performance Requirements. AOAC Official Methods of Analysis, 1–17.

Bhat, S., Curach, N., Mostyn, T., Bains, G. S., Griffiths, K. R., & Emslie, K. R. (2010). Comparison of Methods for Accurate Quantification of DNA Mass Concentration with Traceability to the International System of Units. Analytical Chemistry, 82(17), 7185–7192. https://doi.org/10.1021/ac100845m

Bordbar, S., Anwar, F., & Saari, N. (2011). High-Value Components and Bioactives from Sea Cucumbers for Functional Foods — A Review. Marine Drugs, 9, 1761–1805. https://doi.org/10.3390/md9101761

Catrinck, T. C. P. G., Dias, A., Aguiar, M. C. S., Silvério, F. O., Fidêncio, P. H., & Pinho, G. P. (2014). A Simple and Efficient Method for Derivatization of Glyphosate and AMPA Using 9-Fluorenylmethyl Chloroformate and Spectrophotometric Analysis. Journal of the Brazilian Chemical Society. https://doi.org/10.5935/0103-5053.20140096

Davis, C. M., Gupta, R. S., Aktas, O. N., Diaz, V., Kamath, S. D., & Lopata, A. L. (2020). Clinical Management of Seafood Allergy. The Journal of Allergy and Clinical Immunology: In Practice, 8(1), 37–44. https://doi.org/10.1016/j.jaip.2019.10.019

Du, J., White, N., & Eddington, N. D. (2004). The bioavailability and pharmacokinetics of glucosamine hydrochloride and chondroitin sulfate after oral and intravenous single dose administration in the horse. Biopharmaceutics & Drug Disposition, 25(3), 109–116. https://doi.org/10.1002/bdd.392

Hami, Z., Amini, M., Kiani, A., & Ghazi-Khansari, M. (2013). High Performance Liquid Chromatography Coupled with Pre-column Derivatization for Determination of Oxidized Glutathione Level in Rats Exposed to Paraquat. Iranian Journal of Pharmaceutical Research : IJPR, 12(4), 911–916.

Huang, T., Deng, C.-H., Chen, N., Liu, Z., & Duan, G.-L. (2006). High performance liquid chromatography for the determination of glucosamine sulfate in human plasma after derivatization with 9-fluorenylmethyl chloroformate. Journal of Separation Science, 29(15), 2296–2302. https://doi.org/10.1002/jssc.200600162

Huang, T. M., Deng, C. H., Chen, N. Z., Liu, Z., & Duan, G. L. (2006). High performance liquid chromatography for the determination of glucosamine sulfate in human plasma after derivatization with 9-fluorenylmethyl chloroformate. Journal of Separation Science, 29(15), 2296–2302. https://doi.org/10.1002/jssc.200600162

Ibrahim, A., & Jamali, F. (2010). Improved sensitive high performance liquid chromatography assay for glucosamine in human and rat biological samples with fluorescence detection. Journal of Pharmacy and Pharmaceutical Sciences, 13(2), 128–135. https://doi.org/10.18433/j3t01s

Kelly, W. R., MacDonald, B. S., & Guthrie, W. F. (2008). Gravimetric Approach to the Standard Addition Method in Instrumental Analysis. 1. Analytical Chemistry, 80(16), 6154–6158. https://doi.org/10.1021/ac702437f

Khotimchenko, Y. (2018). Pharmacological Potential of Sea Cucumbers. International Journal of Molecular Sciences, 19(1342), 1–42. https://doi.org/10.3390/ijms19051342

Lee, Y. H., Woo, J. H., Choi, S. J., Ji, J. D., & Song, G. G. (2010). Effect of glucosamine or chondroitin sulfate on the osteoarthritis progression: A meta-analysis. Rheumatology International, 30(3), 357–363. https://doi.org/10.1007/s00296-009-0969-5

Li, B., Zhang, J., Bu, F., & Xia, W. (2013). Determination of chitosan with a modified acid hydrolysis and HPLC method. Carbohydrate Research, 366, 50–54. https://doi.org/10.1016/j.carres.2012.11.005

Liu, Y., Xu, F., Zhang, Z., Yang, C., Song, R., & Tian, Y. (2009). Analysis of synephrine in da-cheng-qi decoction by HPLC employing precolumn derivatization with 9-fluorenylmethyl chloroformate. Journal of Chromatographic Science, 47(10), 925–930. https://doi.org/10.1093/chromsci/47.10.925

Lopez-Cervantes, J., Sonchez-Machado, D. I., & Delgado-Rosas, K. E. (2007). Quantitation of Glucosamine From Shrimp Waste Using HPLC. Journal of Chromatographic Science, 45(4), 195–199. https://doi.org/10.1093/chromsci/45.4.195

Magnusson, B., & Örnemark, U. (2014). Eurachem Guide: The Fitness for Purpose of Analytical Methods – A Laboratory Guide to Method Validation and Related Topics, (2nd ed.). In Eurachem Guide: The Fitness for Purpose of Analytical Methods – A Laboratory Guide to Method Validation and Related Topics, (2nd ed.).

Melton, J. R., Hoover, W. L., & Howard, P. A. (1973). Comparison of Gravimetric and Volumetric Sodium Tetraphenylboron Methods for Determining K2O in Fertilizers. Journal of AOAC INTERNATIONAL, 56(2), 375–377. https://doi.org/10.1093/jaoac/56.2.375

Mohammadi, B., Tammari, E., Fakhri, S., & Bahrami, G. (2013). Applicability of LC–MS/MS to optimize derivatization of topiramate with FMOC-Cl using reacted/intact drug ratio. Journal of Chromatography B, 928, 32–36. https://doi.org/10.1016/j.jchromb.2013.02.041

Morris, A. S., & Langari, R. (2016). Measurement Uncertainty. In Measurement and Instrumentation (pp. 45–73). https://doi.org/10.1016/B978-0-12-800884-3.00003-4

Pangestuti, R., & Arifin, Z. (2018). Medicinal and health benefit effects of functional sea cucumbers. Journal of Traditional and Complementary Medicine, 8(3), 341–351. https://doi.org/10.1016/j.jtcme.2017.06.007

Papich, M. G. (2016). Glucosamine Chondroitin Sulfate. In Saunders Handbook of Veterinary Drugs (pp. 357–358). https://doi.org/10.1016/B978-0-323-24485-5.00282-5

Rasyid, A. (2017). NUTRITIONAL VALUE AND HEAVY METALS CONTENTS OF THE DRIED SEA CUCUMBER Stichopus vastus FROM SALEMO ISLAND, INDONESIA. Jurnal Ilmu Dan Teknologi Kelautan Tropis, 9(2), 739–746. https://doi.org/http://dx.doi.org/10.28930/jitkt.v9i2

Researcher, V. B. (2010). HPLC Method Development for Glucosamine Sulphate and Diacerein Formulation. Journal of Pharmacy Research, 3(2), 361–363.

Rienitz, O., Schiel, D., Güttler, B., Koch, M., & Borchers, U. (2007). A convenient and economic approach to achieve SI-traceable reference values to be used in drinking-water interlaboratory comparisons. Accreditation and Quality Assurance, 12(12), 615–622. https://doi.org/10.1007/s00769-007-0320-3

Ross, M. P., Ferguson, M., Street, D., Klontz, K., Schroeder, T., & Luccioli, S. (2008). Analysis of food-allergic and anaphylactic events in the National Electronic Injury Surveillance System. Journal of Allergy and Clinical Immunology, 121(1), 166–171. https://doi.org/10.1016/j.jaci.2007.10.012

Schneider, K., Silverman, J., Woolsey, E., Eriksson, H., Byrne, M., & Caldeira, K. (2011). Potential influence of sea cucumbers on coral reef CaCO 3 budget: A case study at One Tree Reef. Journal of Geophysical Research, 116(G4), G04032. https://doi.org/10.1029/2011JG001755

Song, M., Hang, T. J., Wang, C., Yang, L., & Wen, A. D. (2012). Precolumn derivatization LCMS/MS method for the determination and pharmacokinetic study of glucosamine in human plasma and urine. Journal of Pharmaceutical Analysis, 2(1), 19–28. https://doi.org/10.1016/j.jpha.2011.08.003

Tekko, I. A., Bonner, M. C., & Williams, A. C. (2006). An optimized reverse-phase high performance liquid chromatographic method for evaluating percutaneous absorption of glucosamine hydrochloride. Journal of Pharmaceutical and Biomedical Analysis, 41(2), 385–392. https://doi.org/10.1016/j.jpba.2005.11.044

Towo, A. (2004). Status of sea cucumber fisheries and farming in Indonesia. 49–60.

Turner, P. J., Jerschow, E., Umasunthar, T., Lin, R., Campbell, D. E., & Boyle, R. J. (2017). Fatal Anaphylaxis: Mortality Rate and Risk Factors. The Journal of Allergy and Clinical Immunology: In Practice, 5(5), 1169–1178. https://doi.org/10.1016/j.jaip.2017.06.031

Vanatta, L. E., & Coleman, D. E. (2007). Calibration, uncertainty, and recovery in the chromatographic sciences. Journal of Chromatography A, 1158(1–2), 47–60. https://doi.org/10.1016/j.chroma.2007.02.040

Wang, X., Chen, X., Chen, L., Wang, B., Peng, C., He, C., … Wei, Y. (2008). Optimizing high-performance liquid chromatography method for quantification of glucosamine using 6-aminoquinolyl- N -hydroxysuccinimidyl carbamate derivatization in rat plasma: application to a pharmacokinetic study. Biomedical Chromatography, 22(11), 1265–1271. https://doi.org/10.1002/bmc.1056

Wilson, R. E., Groskreutz, S. R., & Weber, S. G. (2016). Improving the Sensitivity, Resolution, and Peak Capacity of Gradient Elution in Capillary Liquid Chromatography with Large-Volume Injections by Using Temperature-Assisted On-Column Solute Focusing. Analytical Chemistry, 88(10), 5112–5121. https://doi.org/10.1021/acs.analchem.5b04793

Zhang, Z., Renen, Z., & Liu, G. (1996). High-performance liquid chromatographic analysis of hexosamines, hexosaminitols, N-acetylhexosamines and Nacetylhexosaminitols by ultraviolet and fluorescence detection at picomole levels. Journal of Chromatography A, 730(1–2), 107–114. https://doi.org/10.1016/0021-9673(95)01236-2

Zhou, J. Z. Q., Waszkuc, T., & Mohammed, F. (2004). Single laboratory validation of a method for determination of glucosamine in raw materials and dietary supplements containing glucosamine sulfate and/or glucosamine hydrochloride by high-performance liquid chromatography with FMOC-Su derivatization. Journal of AOAC International, 87(5), 1083–1092. https://doi.org/10.1093/jaoac/87.5.1083

Zhu, X., Cai, J., Yang, J., & Su, Q. (2005). Determination of glucosamine in impure chitin samples by high-performance liquid chromatography. Carbohydrate Research, 340(10), 1732–1738. https://doi.org/10.1016/j.carres.2005.01.045

DOI: http://dx.doi.org/10.36974/jbi.v11i2.6394