Simultaneous determination of betaine and choline using derivatization by HPLC with UV detection

Abstract

Extraction of quaternary ammonium compounds (choline and betaine) from plant samples (spinach) using ion exchange resin (AG1, OH − form) is a very simple and inexpensive approach. However, it is very hard to determine amounts of choline and betaine simultaneously using high-performance liquid chromatographyultraviolet (HPLC-UV) detection. Unlike choline, betaine has low molar absorptivity in UV-visible (UV-Vis) region, which makes it difficult to carry out UV-Vis detection of betaine. The mixture of quaternary ammonium compounds (choline and betaine) was derivatized using 2-bromo acetophenone as a derivatizing agent. As a result, choline did not react with the derivatizing agent, whereas betaine formed a betaine derivative. This betaine derivative exhibited detectable UV absorption with baseline separation between choline and the betaine derivative. Thus, with this method, choline and betaine can be determined simultaneously by using the HPLC-UV method through one-step derivatization, which is an easy, sensitive, and reliable method.

Development and optimization of a method for determining betaine and trigonelline in the fruits of Lycium species by using solid‐phase extraction combined with high‐performance liquid chromatography–diode array detector

“…In order to obtain more better and efficient extraction of betaine from Lycium fruits, several published extraction methods such as different aqueous organic solvents, accelerated solvent extraction coupled with SPE, pre‐ and post‐derivatization [15,16,18–21], precipitation with reinecke salt solution [8] were investigated and compared. As shown in Supporting Information Figure S1A and B, complex peaks composition and bad separation were obtained by using the published derivatization methods [18,19], which were not suitable for betaine determination. Betaine was only partly extracted by using reflux extraction with water or methanol [15,16,21] as shown in Supporting Information Figure S1C and D, and the peak of betaine was not a pure single peak and bad separation with other peaks in the HPLC chromatograms.…”

“…Moreover, the existing published methods [8,15,16,19‐21] were simultaneously carried out and compared with the above method.…”

“…Because the special structure features of betaines consisting with a dipolar ion possessing a quaternary ammonium group and a carboxylic acid group, betaine and its analogues have a poor ultraviolet chromophore, and are highly soluble in water [15,16], which leads to a great challenge for their extraction and quantification. To date, several analytical approaches for determining betaine in Lycium fruits have been developed, such as HPLC–UV, HPLC–ELSD, GC–MS, TLC‐scanning, and LC–MS/MS [8,15–22]. In addition, accelerated solvent extraction coupled with SPE [10,18], pre‐ or post‐derivatization [18,19], and precipitation with reinecke salt solution have been employed for betaine extraction [8].…”

“…To date, several analytical approaches for determining betaine in Lycium fruits have been developed, such as HPLC–UV, HPLC–ELSD, GC–MS, TLC‐scanning, and LC–MS/MS [8,15–22]. In addition, accelerated solvent extraction coupled with SPE [10,18], pre‐ or post‐derivatization [18,19], and precipitation with reinecke salt solution have been employed for betaine extraction [8]. However, complex extraction procedures in ChP was hard to obtain good recovery, and TLC scanning with lower sensitivity and resolution degree was not suitable for the quantitative analysis of betaine [8].…”

“…However, complex extraction procedures in ChP was hard to obtain good recovery, and TLC scanning with lower sensitivity and resolution degree was not suitable for the quantitative analysis of betaine [8]. The published pre‐ or post‐derivatization extraction methods cannot be easily operated and repeated, and betaine and its analogs were detected and quantified as one chromatographic peak by HPLC–ELSD or LC–MS [18,19]. Moreover, the chemical characteristics and content distribution of betaine and its analogs in different Lycium species were still unclear.…”