Detection of Chymotrypsin by Optical and Acoustic Methods

Abstract

Chymotrypsin is an important proteolytic enzyme in the human digestive system that cleaves milk proteins through the hydrolysis reaction, making it an interesting subject to study the activity of milk proteases. In this work, we compared detection of chymotrypsin by spectrophotometric dynamic light scattering (DLS) and quartz crystal microbalance (QCM) methods and determined the limit of chymotrypsin detection (LOD), 0.15 ± 0.01 nM for spectrophotometric, 0.67 ± 0.05 nM for DLS and 1.40 ± 0.30 nM for QCM methods, respectively. The sensors are relatively cheap and are able to detect chymotrypsin in 3035 min. While the optical detection methods are simple to implement, the QCM method is more robust for sample preparation, and allows detection of chymotrypsin in non-transparent samples. We give an overview on methods and instruments for detection of chymotrypsin and other milk proteases.

Detection of Sub-Nanomolar Concentration of Trypsin by Thickness-Shear Mode Acoustic Biosensor and Spectrophotometry

“…In addition, due to covalent binding of casein molecules at the self-assembled MUA, the casein layer is compactly packed, which creates additional restriction of access of trypsin to the cleavage sites. A similar conclusion was also obtained for chymotrypsin detection [57]. In contrast, at AuNPs, the casein is physically adsorbed at the gold surface.…”

“…Further modification with MCH resulted in a significant red shift around 60 nm accompanied by the broadening of the spectrum. This broadening is indicative of an aggregation of nanoparticles This is due to the replacement of the β-casein protective layer with MCH, which in turn makes the nanoparticles closer to each other [57,58]. The modification of AuNPs with MCH resulted in a significant red shift, indicating strong aggregation of the nanoparticles.…”

Detection of chymotrypsin using a peptide sensor based on graphene oxide modified with sulfhydryl groups and gold nanoparticles

“…The limit of detection (LOD) based on 3S/N was 0.25 pg mL −1 . Compared with other methods for detecting chymotrypsin, the detection limit reported by Mu et al 36 was 1.3 μg mL −1 , Gao et al 37 was 7.39 pg mL −1 , Piovarci et al was 1.40 nM, 8 and Fu et al was 0.4 nM. 9 This study has reported a label-free detection method using a important material, GO with sulfhydryl groups, compared with Gao's paper.…”

“…[1][2][3] Therefore, developing a reliable and sensitive detection method for chymotrypsin is very important for testing pancreatic function. 4 Traditional methods such as liquid crystals (0.1 ng mL À1 ), 5 mass spectroscopy (MS, 30 pM), 6,7 quartz crystal microbalance (QCM, 1.40 nM), 8 and surface-enhanced Raman scattering (SERS, 0.4 nM) 9 are sensitive and accurate and can be used to evaluate the function of the exocrine pancreas. Voltammetry and atomic force microscopy can be used to characterize the enzyme kinetic parameters in the proteasome catalyzed-hydrolysis of oligopeptide sequences (i.e., chymotrypsin activities) 10 but are not suitable for the detection of chymotrypsin itself.…”

Detection of Oenological Polyphenols via QCM-D Measurements

“…In a typical biosensor, a biologically active molecule (probe) decorates a surface exposed to the sample [ 6 , 7 ], and interacts with the analyte [ 8 ]. Due to their versatility and high performance, acoustic-wave-based biosensors find a variety of applications, such as for the detection of proteins [ 9 , 10 , 11 ] and enzymes [ 12 , 13 , 14 ] and for fast diagnostics [ 15 ]. To date, there has been a growing interest in the exploitation of acoustic sensors in food science [ 16 ], opening the way to new sciences such as precision oenology [ 17 ].…”