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class="content"><article class="post-4960 post type-post status-publish format-standard has-post-thumbnail category-ion-chromatography category-pharmaceutical tag-ion-chromatography tag-pharmaceutical entry" aria-label="Simple, Accurate and Robust Determination of Tromethamine in Pharmaceutical Formulations"><header class="entry-header"><h1 class="entry-title">Simple, Accurate and Robust Determination of Tromethamine in Pharmaceutical Formulations</h1> <span class="post-info-author"> By <a href="https://www.thermofisher.com/blog/analyteguru/tf-author/analyteguru-staff/" title="Posts by AnalyteGuru Staff" rel="author">AnalyteGuru Staff, </a> </span> <span class="post-info-date"> 11.28.2022 </span> </header><div class="entry-content"><div class="lia-message-body-content"> <p>Tromethamine is a useful compound in pharmaceutical formulations acting as a buffering agent, an alkalizer, an emulsifying agent, or a counterion for acidic drug substances. Although tromethamine brings a host of benefits to these preparations, current methods to determine it are expensive and suffer from lack of specificity. Now, with recent advances in ion chromatography (IC), newly designed methods can achieve increased separation of ions and improve determination.</p> <h3 id="toc-hId-1154439951">Solving the chromophore challenge</h3> <p><img decoding="async" class="alignright wp-image-4965 " src="https://www.thermofisher.com/blog/analyteguru/wp-content/uploads/sites/25/2023/06/19512-og-300x200.jpg" alt="" width="381" height="254" srcset="https://www.thermofisher.com/blog/analyteguru/wp-content/uploads/sites/25/2023/06/19512-og-300x200.jpg 300w, https://www.thermofisher.com/blog/analyteguru/wp-content/uploads/sites/25/2023/06/19512-og-768x512.jpg 768w, https://www.thermofisher.com/blog/analyteguru/wp-content/uploads/sites/25/2023/06/19512-og-96x65.jpg 96w, https://www.thermofisher.com/blog/analyteguru/wp-content/uploads/sites/25/2023/06/19512-og.jpg 998w" sizes="(max-width: 381px) 100vw, 381px" />As an alkanol amine, tromethamine has a host of interesting properties that make it a highly versatile compound in pharmaceutical preparations. Yet, its structure makes a determination of the compound challenging, most notably due to the lack of a chromophore. To overcome this, various derivatization reagents have been used to modify tromethamine. Adding a chromophore in this manner allows detection and thus determination by high-performance liquid chromatography (HPLC) and spectrophotometry<a href="#_edn1" target="_blank" rel="noopener nofollow noreferrer" name="_ednref1">[1]</a>.</p> <p>Alternatively, IC can be used for direct determination of tromethamine, as it does not require a chromophore for detection. IC offers additional advantages over existing assays, as it can simultaneously determine sodium, ammonium and other common cations<a href="#_edn2" target="_blank" rel="noopener nofollow noreferrer" name="_ednref2">[2]</a>. An IC method, first reported in 1995, uses manually prepared eluents and non-suppressed conductivity detection<a href="#_edn3" target="_blank" rel="noopener nofollow noreferrer" name="_ednref3">[3]</a>. Although the approach is effective, new advances in technology offer opportunities to modernize and improve the assay.</p> <h3 id="toc-hId-2041943632">Revolutionizing determination with a Dionex column</h3> <p>In a <a href="https://www.thermofisher.com/document-connect/document-connect.html?url=https%3A%2F%2Fassets.thermofisher.com%2FTFS-Assets%2FCMD%2FApplication-Notes%2Fan-000165-ic-ics-6000-vaccine-excipient-tromethamine-an000165-na-en.pdf&title=QXBwbGljYXRpb24gTm90ZTogQXNzYXkgb2YgVHJvbWV0aGFtaW5lIGluIFBoYXJtYWNldXRpY2FsIEZvcm11bGF0aW9ucw==" target="_blank" rel="nofollow noopener">recent application note, we designed a reagent-free IC (RFIC) method</a> using a Thermo Scientific™ <a href="https://www.thermofisher.com/order/catalog/product/302606" target="_blank" rel="nofollow noopener">Dionex™ IonPac™ CS20</a> cation exchange column to optimize separation and improve determination of tromethamine in pharmaceutical formulations. To electrolytically generate methanesulfonic acid (MSA) eluent, we used a Thermo Scientific™ <a href="https://www.thermofisher.com/us/en/home/industrial/chromatography/ion-chromatography-ic/ic-consumables/eluent-generator-cartridges.html?gclid=Cj0KCQiAg_KbBhDLARIsANx7wAxe-5wXvl2yEORg6fUtObr6J8xa44yrHAktAGul8s2-gHmdyDZiC20aAgzAEALw_wcB&cid=E.22CMD.DL104.05702.01&ef_id=Cj0KCQiAg_KbBhDLARIsANx7wAxe-5wXvl2yEORg6fUtObr6J8xa44yrHAktAGul8s2-gHmdyDZiC20aAgzAEALw_wcB:G:s&s_kwcid=AL!3652!3!625132766067!p!!g!!eluent%20generation" target="_blank" rel="nofollow noopener">Dionex™ EGC 500 MSA</a> eluent generator cartridge with a Thermo Scientific™ <a href="https://www.thermofisher.com/order/catalog/product/040192" target="_blank" rel="nofollow noopener">Dionex™ CR-CTC 500</a> (continuously regenerated cation trap column). Additionally, the method features suppressed conductivity detection with a Thermo Scientific™ <a href="https://www.thermofisher.com/order/catalog/product/088670CMD?SID=srch-srp-088670CMD" target="_blank" rel="nofollow noopener">Dionex™ CDRS 600</a> (cation electrolytically regenerated suppressor).</p> <p>A challenge in the accurate determination of tromethamine is interference from matrix ions. However, the Dionex IonPac CS20 column overcomes this by providing a large separation window between monovalent and divalent cations with most amines eluting after the alkali metals, but before the alkaline earth metals — providing a well-resolved chromatogram.</p> <h3 id="toc-hId--1365519983">Accurate, robust and sensitive tromethamine determination</h3> <p>To demonstrate the benefits of this new assay, we highlight the effective separation of tromethamine. Using the Dionex IonPac CS20 column under isocratic elution, good separation of tromethamine from the nearest cations (sodium and ammonium) was achieved in just 30 minutes (Figure 1).</p> <p><img decoding="async" class="aligncenter size-full wp-image-12296" src="https://www.thermofisher.com/blog/analyteguru/wp-content/uploads/sites/25/2023/06/112822-Figure-1-tromethamine-1.png" alt="" width="479" height="319" srcset="https://www.thermofisher.com/blog/analyteguru/wp-content/uploads/sites/25/2023/06/112822-Figure-1-tromethamine-1.png 479w, https://www.thermofisher.com/blog/analyteguru/wp-content/uploads/sites/25/2023/06/112822-Figure-1-tromethamine-1-300x200.png 300w, https://www.thermofisher.com/blog/analyteguru/wp-content/uploads/sites/25/2023/06/112822-Figure-1-tromethamine-1-96x65.png 96w" sizes="(max-width: 479px) 100vw, 479px" /></p> <p>We also validated the method to determine tromethamine per the guidelines outlined in USP General Chapter <1225>, Validation of Compendial Procedures<a href="#_edn1" target="_blank" rel="noopener nofollow noreferrer" name="_ednref1">[4]</a>. The assay was studied for robustness, linearity and precision, sensitivity and accuracy. Overall, we found the method was:</p> <ul> <li><strong>Robust: </strong>It is resistant to 10% flow rate and eluent concentration changes.</li> <li><strong>Linear: </strong>A coefficient of determination value determined was 1 for a quadratic fit.</li> <li><strong>Precise:</strong> All retention time relative standard deviations (RSD) were ≤0.83%.</li> <li><strong>Sensitive:</strong> The limit of detection (LOD) and limit of quantitation (LOQ) for tromethamine were 0.05 ppm and 0.15 ppm, respectively.</li> <li><strong>Accurate:</strong> Recovery of tromethamine was good, between 95.4 and 112%.</li> </ul> <h3 id="toc-hId--478016302">Transforming determination in pharmaceuticals</h3> <p>To investigate a real-world application, a simulated matrix sample was prepared containing tromethamine, sucrose, acetic acid, and sodium acetate, based on a COVID-19 vaccine excipient composition listed by John Hopkins’ Institute of Vaccine Safety<a href="#_edn1" target="_blank" rel="noopener nofollow noreferrer" name="_ednref1">[5]</a>. As can be seen in Figure 2, the tromethamine peak was well resolved from the largest component— sodium — in the simulated matrix sample. The suitability of this assay for the determination of tromethamine in a simulated matrix shows great promise for pharmaceutical formulation analysis.</p> <p><img loading="lazy" decoding="async" class="aligncenter size-full wp-image-12297" src="https://www.thermofisher.com/blog/analyteguru/wp-content/uploads/sites/25/2023/06/112822-Figure-2-tromethamine-1.png" alt="" width="487" height="345" srcset="https://www.thermofisher.com/blog/analyteguru/wp-content/uploads/sites/25/2023/06/112822-Figure-2-tromethamine-1.png 487w, https://www.thermofisher.com/blog/analyteguru/wp-content/uploads/sites/25/2023/06/112822-Figure-2-tromethamine-1-300x213.png 300w" sizes="(max-width: 487px) 100vw, 487px" /></p> <p>To summarize, the IC-based assay described here is an accurate, sensitive and robust approach to determine tromethamine. As the method requires no prior derivatization, it is convenient and cost-effective compared to other assays. We also envisage that it should be applicable to other pharmaceutical formulations.</p> <p>For <a href="https://www.thermofisher.com/document-connect/document-connect.html?url=https%3A%2F%2Fassets.thermofisher.com%2FTFS-Assets%2FCMD%2FApplication-Notes%2Fan-000165-ic-ics-6000-vaccine-excipient-tromethamine-an000165-na-en.pdf&title=QXBwbGljYXRpb24gTm90ZTogQXNzYXkgb2YgVHJvbWV0aGFtaW5lIGluIFBoYXJtYWNldXRpY2FsIEZvcm11bGF0aW9ucw==" target="_blank" rel="nofollow noopener">more details on the IC method for determination of tromethamine in pharmaceutical formulations, read…</a>, and be sure to check out an upcoming blog post to see how high-performance anion-exchange chromatography with pulsed amperometric detection (HPAE-PAD) coupled to a mass spectrometer (MS) can be used for structural characterization of label-free O-glycans.</p> <p>[1] Bubois, P.; Lacroix, J.; Lacroix, R.; Levillain, E.; Viel. C. Application of Z α,β-dinitrostilbene to aminic drugs assay. J. Pharm. Belg. 1981, 36, 203.</p> <p>[2] Fritz, J.S.; Gjerde, D.T.; Becker, R.M. Cation Chromatography with a Conductivity Detector. Anal. Chem. 1980, 52, 1519–1522.</p> <p>[3] Hall, R.E.; Havner, G.D.; Good, R.; Dunn, D. Ion chromatographic method for rapid and quantitative determination of tromethamine, J. Chromatogr. A. 1995, 718(2), 305–308.</p> <p>[4] Validation of Compendial Methods, General Chapter <1225>. U.S. Pharmacopeia/National Formulary: Rockville, MD, USP36-NF31; p 983.</p> <p>[5] Johns Hopkins Institute of Vaccine Safety. [Online] <a href="https://www.vaccinesafety.edu/components-Excipients.htm" target="_blank" rel="noopener nofollow noreferrer">https://www.vaccinesafety.edu/components-Excipients.htm</a> (Accessed May 16, 2022)</p> <p><a href="#_ednref1" target="_blank" rel="noopener nofollow noreferrer" name="_edn1"></a></p> </div> </div></article><section class="author-box"><h4 class="author-box-title">AnalyteGuru Staff</h4><div class="author-box-content" itemprop="description"></div></section><div class="adjacent-entry-pagination pagination"><div class="pagination-previous alignleft"><i class="fa fa-icon fa-chevron-left"></i> <a href="https://www.thermofisher.com/blog/analyteguru/discovery-of-mosh-moah-in-german-butter-may-spoil-your-taste-for/" rel="prev">MOSH/MOAH Discovery in German Butter May Spoil Your Taste for Holiday Treats</a></div><div class="pagination-next alignright"><a href="https://www.thermofisher.com/blog/analyteguru/helping-laboratories-work-toward-sustainability/" rel="next">Helping Laboratories Work Toward Sustainability</a> <i class="fa fa-icon fa-chevron-right"></i></div></div></main></div></div><footer class="site-footer"><div class="wrap"><div class="creds"><p class="footer-links"><a href="https://corporate.thermofisher.com/en/footer/privacy-statement.html" target="_blank" rel="noopener noreferrer">Privacy Statement</a><a href="https://corporate.thermofisher.com/en/footer/terms-and-conditions.html" target="_blank" rel="noopener noreferrer">Terms & Conditions</a><a href="/blog/locations/">Locations</a><a href="/blog/sitemap/">Sitemap</a></p><p>© 2024 <a href="https://www.thermofisher.com">Thermo Fisher Scientific</a>. 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