Study of the effect of culture mediums on the amino acid metabolites for Corynebacterium glutamicum using high‐speed micellar electrokinetic chromatography

Abstract

Corynebacterium glutamicum (C. glutamicum) is a well‐known workhorse for the industrial production of amino acids. Different carbon, nitrogen, and sulfur source may force the bacterium to produce specific metabolites. In this work, a method of high‐speed MEKC with LIF detection was developed to rapidly analyze the amino acid metabolites released by C. glutamicum, which is fed with different culture mediums. Corynebacterium glutamicum was cultured in microbial fuel cells to monitor its metabolism process and collect its metabolites. In the CE system, a microliter‐scale sample reservoir was designed and applied to perform tiny volume sample injection. With the assistance of microwave, the derivatization time for amino acids with FITC was greatly shortened to 6 min. Under the optimized condition, the eight candidate amino acids of metabolites could be rapidly separated within 2 min. The whole analysis process for real samples, from sampling to determination, could be shortened to less than 10 min. The results showed that C. glutamicum could produce additional l‐lysine and l‐valine as the metabolites when fed with glucose and l‐methionine, respectively. The method proved that culture mediums used to feed C. glutamicum had great effect on the bacterium's metabolites.

Separation and determination of 3‐hydroxyaspartate by online concentration capillary electrophoresis/laser‐induced fluorescence with microwave‐assisted derivatization

“…Similarly, the fluorescence signal was enhanced until 30 min (Figure 1B). Compared with the short reaction time of several minutes or seconds reported in a literature [29–31], the longer time needed in this experiment was probably caused by the following two factors: First, the constant‐temperature mode of the microwave oven was used and the microwave power was only about 70 W to maintain the temperature at 70°C. Second, the volume of sample was just 200 μL, which induced lower microwave energy absorbed.…”

“…Similarly, the fluorescence signal was enhanced until 30 min (Figure 1B). Compared with the short reaction time of several minutes or seconds reported in a literature [29][30][31], the longer time needed in this experiment was probably caused by the following two factors: First, the constant-temperature mode of the microwave oven was used and the microwave power was only about 70 W to maintain the temperature at 70 • C. Second, the volume of sample was just 200 μL, which induced lower microwave energy absorbed. Actually, we tried to increase the volume of sample to 4 mL, the optimal F I G U R E 1 Impact of (A) reaction temperature and (B) reaction time on microwave-assisted derivatization F I G U R E 2 Electropherograms of microwave-assisted derivatization (red solid line) and water bath derivatization (black dotted line) under optimal conditions.…”

“…Similarly, the fluorescence signal was enhanced until 30 min (Figure 1B). Compared with the short reaction time of several minutes or seconds reported in a literature [29][30][31], the longer time needed in this experiment was probably caused by the following two factors: First, the constant-temperature mode of the microwave oven was used and the microwave power was only about 70 W to maintain the temperature at 70 As a result of the above optimization and comparative study on the derivatization method, microwave-assisted derivatization was chosen as a more efficient method with the following conditions: 10.0 mmol/L (pH = 8.5) borax, ratio of NBD-Cl to HA as 14:1, temperature of 70 • C, and derivatization time of 30 min.…”

Capillary and microchip electrophoresis method development for amino acid monitoring during biopharmaceutical cultivation

“…MEKC with laser induced fluorescence (LIF) detection was employed to analyze eight AA metabolites for Corynebacterium glutamicum after microwave‐assisted fluorescein isothiocyanate derivatization. [ 12 ] Celá et al . developed MEKC methods employing CE with fluorescence detection to analyze AAs in culture media for human embryos.…”

“…MEKC with laser induced fluorescence (LIF) detection was employed to analyze eight AA metabolites for Corynebacterium glutamicum after microwave-assisted fluorescein isothiocyanate derivatization. [12] Celá et al developed MEKC methods employing CE with fluorescence detection to analyze AAs in culture media for human embryos. [13][14][15] In samples of spent medium, 21 naphthalene-2,3-dicarboxyaldehyde (NDA) derivatized AAs were separated in a 45 cm effective length capillary using CE with light-emitting diode-induced fluorescence detection.…”