![[Readers Insight] Triethylamine as a Mobile Phase Additive: What Does It Do?](http://www.welch-us.com/cdn/shop/articles/triethylamine-1080-1215_750x.jpg?v=1765790893)
[Readers Insight] Triethylamine as a Mobile Phase Additive: What Does It Do?
Author: Sepuxianyun
Previous article:
Heterocycles Structural Analysis in HPLC Method Development
Introduction
In HPLC method development, the choice of mobile phases is highly flexible. In an previous article, we briefed a general overview of mobile phase selection. (If you haven't read it, be sure to take a look.)
In this article, we will focus on a specific, familiar additive: Triethylamine (TEA), and discuss some of its unique roles in chromatography.
Role 1: Silanol Suppressor
This is the most well-known application of TEA.
Approximately 70% of pharmaceuticals are basic compounds, and over the years, it has become a common practice in method development to add triethylamine to mask secondary interactions of silanol groups (Si-OH) on the column packing and thereby resolve peak tailing issues of target analytes. (As noted in earlier articles, alternative chaotropic reagents can sometimes be more convenient; this will not be repeated here.)
Example: The mobile phase for Terbinafine Hydrochloride related substances in the Chinese Pharmacopoeia:
- Mobile Phase A: Triethylamine buffer (0.2% triethylamine solution, pH adjusted to 7.5 with glacial acetic acid) - Methanol - Acetonitrile (30:42:28, v/v/v).
- Mobile Phase B: Triethylamine buffer - Methanol - Acetonitrile (5:57:38, v/v/v).
Furthermore, in the analysis of basic compounds in normal-phase systems for isomer separation, triethylamine and diethylamine are often added to improve peak shape. In these cases, diethylamine is generally preferred over triethylamine for better effectiveness, as diethylamine has a slightly lower pKb, thus stronger in basicity than triethylamine. When the target analyte contains primary or secondary amines, its basicity is stronger than that of triethylamine, intensifying competitive interactions.
Example: The mobile phase for Butorphanol Tartrate chiral isomers:
- n-Hexane – Ethanol – Diethylamine (70:30:0.1, v/v/v).
Role 2: Ion-Pairing Reagent
Conventional IPRs (which modify the column)
Before we explain the role of triethylamine as an ion-pairing reagent (IPR), we first take a look at conventional IPRs. In the separation of highly polar, acidic compounds that are difficult to retain, IPRs are often added to provide ion-exchange interactions, increasing retention.
Tetrabutylammonium (TBA) salts such as TBAOH or TBAHS are the most common ion-pairing reagents. Newer IPRs, for example amylamine (a.k.a pentylamine) and N,N-dimethyloctylamine, also play a similar role and are more friendly to columns. But all those long-chain alkylamines modify the column, and thus their use should be on dedicated columns.
Alkyl sulfonic acid IPRs are organic acids, and are used primarily on retaining basic analytes, acting inversely to organic bases.
Example: The mobile phase for Cefixime related substances in the Chinese Pharmacopoeia:
- TBAOH solution (Measure 25 mL of 10% tetrabutylammonium hydroxide, dissolve in 1000 mL water, mix well, and adjust the pH to 7.0 with 1.5 mol/L phosphoric acid) - Acetonitrile (72:28, v/v).
Special IPRs (which do not modify the column)
Beyond acting as a silanol suppressor, triethylamine also possesses certain ion-exchange behavior, forming a triethylammonium acetate (TEAA) system when combined with acetic acid.
TEAA is frequently used for oligonucleotide analysis: oligonucleotides are highly polar and are poorly retained under conventional reversed-phase conditions, yet TEAA enables their retention on reversed-phase columns, illustrating the unique properties of this combination.
Similarly, TEAA can also be applied for small, highly polar molecules bearing multiple acidic groups, often producing unexpectedly good retention and separation.
Example: Mobile phase for Mipomersen (an oligonucleotide drug) related substances:
- Mobile Phase A: 0.1 mol/L TEAA solution (pH 7.0).
- Mobile Phase B: Acetonitrile - 0.1 mol/L TEAA solution (pH 7.0) (80:20, v/v).
It is also seen in some literatures that TEAA can have applications in certain chiral separations.
Role 3: Constructing Spatial Selectivity Systems
Some non-enantiomeric stereoisomers and constitutional isomers are difficult to resolve because of their structures being too similar. In such cases, adding triethylamine to the mobile phase can sometimes yield surprising improvements.
Why does triethylamine work here? While triethylamine binds to silanol groups ($Si-OH$) on C18 columns to mask secondary effects, its three "tails" (ethyl groups) extend outward. These ethyl groups are hydrophobic and, together with the surrounding C18 chains, construct a "stable" spatial environment. This creates a degree of spatial selectivity.
Therefore, when facing separations of closely related structural isomers, it is worthwhile to try triethylamine as an additive.
Conclusion
In HPLC method development, triethylamine is truly a "smelly treasure": while its distinct odor is notorious, its utility is undeniable. By mastering the strategic application of its unique characteristics, analysts can effectively troubleshoot and resolve a wide array of intractable separation challenges.