A simplified visible light spectroscopic method for the detection and assay of monensin

Monensin is a polyether carboxylic ionophore antibiotic existing as a mixture of four isomers denoted simply as A, B, C and D.
calendar icon 11 October 2019
clock icon 6 minute read

As a part of an ongoing undergraduate research project at Palm Beach Atlantic University we wanted to develop a simple method to detect the presence of monensin, monitor the kinetics when reacted with acidified vanillin in dilute solution and quantitatively measure its concentration in the parts per million range. Based on our method we were able to detect monensin in solutions of aqueous methanol using a visible light spectrophotometer at 520 nm to a level of about 3 X 10-6 M, which is approximately 2 mg/L or 2 ppm. In addition to developing this method for our own in-house research, we share it in the hopes that it might be utilised in the animal health industry or at least in a teaching lab at the undergraduate level where an HPLC equipped with a post-column reactor is not available.

Running the Monensin – Vanillin reaction

50 mL of each solution were combined in a 250 mL Erlenmeyer flask that had been placed in a 70 C water bath. The vanillin solution was clear and colorless. The filtered monensin solution was clear but light brown in color. Prior to mixing, small samples of the individual solutions were withdrawn with a 1 mL micropipette and used to zero the absorbance reading of the spectrophotometer. The reaction mixture slowly developed a pink color which continued to intensify to a cherry red over the course of 90 minutes. 1 mL samples, withdrawn every 5 minutes for the first hour and then every 10 minutes thereafter were placed in cuvettes and their absorbance read after 5 seconds. A graph of absorbance vs. time demonstrated that at 90 minutes the slope of the curve approached zero signifying that the reaction was complete (Graph I).

Quantitative assay of Monensin using a Beer’s Law graph

A Beer’s Law plot of absorbance vs. molarity of successively diluted solutions of the reaction mixture after 90 minutes was prepared using Microsoft Excel. Below is the graph showing the results. Also included is the equation for the straight line and the correlation coefficient, (Graph II). A Thermo Scientific Genesys 30 visible spectrophotometer was used to determine all measurements.

Conclusions

Successive 2:1 MeOH dilutions of the reaction mixture
Successive 2:1 MeOH dilutions of the reaction mixture

As a part of an ongoing undergraduate research project at Palm Beach Atlantic University we wanted to develop a simple method to detect the presence of monensin, monitor the kinetics when reacted with acidified vanillin in dilute solution and quantitatively measure its concentration in the parts per million range. Based on our method we were able to detect monensin in solutions of aqueous methanol using a visible light spectrophotometer at 520 nm to a level of about 3 X 10-6 M, which is approximately 2 mg/L or 2 ppm. In addition to developing this method for our own in-house research, we share it in the hopes that it might be utilised in the animal health industry or at least in a teaching lab at the undergraduate level where an HPLC equipped with a post-column reactor is not available.

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