Albumin quantification by alternative chemometric-assisted dye-binding methods robust towards pH and ionic strength variations

Albumin is one of the most common proteins in human fluids, making it one of the most studied and quantified; it is also used as a biological marker for a wide range of pathologies, such as diabetes mellitus, renal diseases, and liver malfunctions. Clinically, it is currently determined using UV-Vis spectroscopy with sulfonephthalein indicators such as Bromophenol Blue (BPB), Bromocresol Green (BCG), and Bromocresol Purple (BCP) at acidic pH levels between 3.5 and 5. These procedures, generally referred to as dye-binding methods, are favored for their rapidity, simplicity, low cost of reagents and instrumentation, and potential for automation. However, they have some disadvantages, including the risk of contamination, the need for skilled operators in manual analysis, low selectivity, matrix effects, time sensitivity, inaccuracy, and low robustness towards pH and ionic strength (I) variations. In this study, new alternative dye-binding methods are proposed using Bovine Serum Albumin (BSA) as model albumin and five different sulfonephthalein dyes, the three aforementioned dyes, Chlorophenol Red (CPR), and Bromothymol Blue (BTB). The successful application of Partial Least Squares regression allows quantifying the protein using each dye and in a range of pH between 3.5 and 9 and of I between around 0.01 and 0.5 M. In addition, most of the multivariate quantification models show an interesting robustness towards both pH and I since the same model can be applied to quantify BSA in samples with much different pH (over 3 log units) and I values.