Use of FT-IR for the Quantification of Water in Biomolecules

Abstract

Water (10⁻³ mg H₂O in 1 mg of biopolymer) was extracted from biological compounds with acetonitrile and determined by FT-IR spectrometry with the use of a KBr transmission cell. A nitrogen-gas-purged glove box was employed during the extraction to avoid errors due to water absorption from ambient laboratory air. The subtractive compensation for concomitant water in acetonitrile was further accomplished by using a reference spectrum of acetonitrile kept under the same experimental conditions as those for preparing the acetonitrile extracts of water from the testing samples. This technique was applied to the quantification of water in peptides and enzymes. The advantages of this technique over the Karl Fisher titration are discussed, along with measurements for loss of weight on drying in vacuo.