A Highly Sensitive and Rapid Assay for Protein Determination in Human Urine and Penicillin Using a Rayleigh Light-Scattering Technique with Benzeneazo-8-acetylamino-1-naphthol-3,6- disulfonic Acid Sodium Salt

Abstract

The quantitative analysis of protein is very vital in many fields, such as clinical analysis, medical research, protein industry and food chemistry. Many methods for the determination of proteins are continuously being put forward, including the Turbidity method, the Kjeldahl method, spectrophotometry, fluorophotometry and so on. [1][2][3][4][5][6][7] Recently, the Rayleigh lightscattering (RLS) technique has become a new and interesting method with many advantages. For instance, it can be performed using an ordinary fluorescence spectrometer. Also, the detection limit of proteins is quite low. This method has been widely used in biomacromolecules, such as proteins and nucleic acids, inorganic ion determinations and so on. [8][9][10][11][12] In the present work, based on the interaction of proteins with benzeneazo-8-acetylamino-1-naphthol-3,6-disulfonic acid sodium salt (also called azophloxine, AP, see Fig. 1) as a probe, a novel determination method was developed using the RLS technique. Azophloxine cannot be employed for protein determination using the spectrophotometric method for lower sensitivity, but can be served as a novel probe for the RLS technique with higher sensitivity. This expands the area of biomacromolecular probes for the RLS method.A microemulsion, as the medium, is introduced into protein determination, and can enhance the sensitivity of this method. Therefore, this assay is much more sensitive than most of the reported light-scattering methods, as displayed in Table 1. [13][14][15][16][17][18][19][20][21][22][23][24][25][26] The content of protein in human urine can provide much information in clinical analysis, and is often used as a reference for disease. At the same time, penicillin is also used in this work, which is a novel sample for protein determination using the RLS technique. If residual protein in penicillin is in excess, it can cause allergic reactions in many individuals, and also affect the yield of penicillin. It is thus necessary to develop a novel, rapid and accurate method for the determination of protein. The proposed method has been successfully applied to the determination of protein in human urine and penicillin samples. ExperimentalApparatus A Shimadzu Model UV-3101 spectrophotometer was employed for absorption spectral records. A Perkin-Elmer Model LS-55 luminescence spectrometer with 1.0 cm quartz cells was used to measure the intensity and spectra of RLS. A PHS-3B acidimeter was used in determining pH values. A simple and sensitive method was conducted for the determination of trace amounts of proteins with benzeneazo-8-acetylamino-1-naphthol-3,6-disulfonic acid sodium salt (azophloxine, AP) using a Rayleigh light-scattering (RLS) technique. At pH 2.60 and in the presence of an emulsifier OP microemulsion, the RLS of AP can be greatly enhanced by proteins, owing to the interaction between AP and protein. The enhanced intensity is proportional to the concentration of proteins. Four proteins, including bovine serum albumin (BSA), human serum albumin (HSA), lysozyme (...

On-line Vapor-Phase Generation Followed by Fourier-Transform Infrared Spectrometry for the Quantitative Analysis of Water-Soluble Penicillin G in Pharmaceutical Formulations

“…Previous methods for measuring PENG in different real samples, such as pharmaceutical formulations, ovine milk, human plasma and wastewater, include methods based on spectrometry, [3][4][5] electrophoresis, 6,7 high-performance liquid chromatography (HPLC), 8,9 gas chromatography, 10 colorimetry, 11 and electrochemistry. 12 However, some of these methods are insensitive, tedious, time consuming or expensive.…”