Determination of Lactate with Liquid Chromatography/Electrochemistry Coupled with a Lactate Oxidase IMER

published for the determination of lactate using liquid chromatography. One method (1) involves reverse-phase separation with a phosphate buffer at pH 3 as the mobile phase. Following the separation, the lactate is converted to hydrogen peroxide by a lactate oxidase IMER and then detected amperometrically. The optimal pH for the enzyme lactate oxidase is around 7, and at pH 3 the enzyme is basically not functioning. Therefore, for this system, a second pump and an online mixer have to be used to increase the pH of the mobile phase after the separation column to keep the enzyme in the IMER active. This makes the system rather complicated. Another method (2) also involving reverse-phase separation with low pH phosphate buffer uses UV detection instead of the electrochemical detection coupled with a lactate oxidase IMER. Lactate does not absorb in the usual UV-VIS region, and the direct detection of lactate has to be conducted in the low wavelength UV region. This makes serious background interferences likely. There is also a reversephase separation using a mixture of phosphate and acetonitrile as mobile phase. For this separation, there are two detection methods: UV-VIS spectrophotometry (3) and fluorimetry (4). For both detection methods, the lactate has to be derivatized and therefore the methods are not convenient. Ion chromatography has also been used for the separation of lactate. The two published methods use conductimetric (5) and acoustic wave (6) detection. The conductimetric method requires several ion-exchange steps to reduce the background, resulting in a very complicated system. Besides, the chromatogram usually involves many anionic peaks that make data analysis difficult. The acoustic wave method, on the other hand, is limited by its lack of commercial availability and difficult data interpretation. In this paper we describe the development of a simple and practical LCEC method for the determination of lactate.