Technical note: Nontargeted detection of adulterated plant proteins in raw milk by UPLC-quadrupole time-of-flight mass spectrometric proteomics combined with chemometrics

Abstract

We built and validated a chemometric model to detect possible milk adulteration with plant proteins. Specifically, we extracted proteins in raw milk, treated with tryptic digestion, and obtained peptide fingerprints by UPLC-quadrupole time-of-flight-mass spectrometry with proteomics to differentiate authentic milks from their counterparts adulterated with nonmilk proteins. This approach is able to detect soybean and pea powder-adulterated milks at as low as 1% (wt/wt). Additionally, we obtained the characteristic peptide sequences for milk authentication by principal component analysis. The prediction accuracies for milk authentication by partial least-squares-discriminant analysis were greater than 95%. These results indicated that peptide fingerprints with the chemometric analysis could be successfully applied for milk quality control.

Detection of plant protein in adulterated milk using nontargeted nano‐high‐performance liquid chromatography–tandem mass spectroscopy combined with principal component analysis

“…An approximate linear relationship of data points dependent on protein levels could be observed for samples adulterated with SP. Results of PCA in the current study are similar to those reported in other literature (Cordewener et al., ; Lu et al., ). Our results show that the adulterated samples could be separated from authentic milk for adulterants proteins.…”

“…In the loading plot (Figure 3b are similar to those reported in other literature (Cordewener et al, 2009;Lu et al, 2017). Our results show that the adulterated samples could be separated from authentic milk for adulterants proteins.…”

“…Among them, β-conglycinin, glycinin, and trypsin inhibitor from SP and vincilin, convicilin, legumin, and provicilin from PP were also identified in other studies, using UHPLC-QTOF MS proteomics, as reported by Lu et al (2017). Meanwhile, proteins identified from SDS-PAGE are also presented in the results of LC-MS/MS identification.…”

“…Combined with multivariable statistics, a variety of nontargeted detection methods have been proposed to identify plant protein adulterated in raw milk. Partial least squares‐discriminant analysis (PLS‐DA) and principal component analysis (PCA) using the fingerprints of intact protein flow injection mass spectra (MS) and ultra‐high‐performance liquid chromatography (UHPLC)–quadrupole time‐of‐flight (QTOF)‐MS were able to detect SP and PP in adulterated milks at the 1% level (Du et al., ; Lu et al., ). Based on the chromatographic files of authentic and adulterated milk powder obtained by UHPLC with UV detection at 215 nm, the t test approach and multivariate Q statistic from a SIMCA model would classify milk powder with SP at 1% and 3% levels correctly and failed to recognize adulterated samples with brown rice and hydrolyzed wheat protein below 10% (Jablonski, Moore, & Harnly, ).…”

“…Detection of soy, pea, wheat, and rice protein at 0.1%-0.2% of sample weight in milk powder was realized by a rapid turbidimetric measure based on the absorbance of the resuspended pellet solution (Scholl, Farris, & Mossoba, 2014), whereas these methods fail to present the origin of these adulterants. With amino acid sequences revealed by fragmented peptides, mass spectrometry (MS) is successful in the identification of plant protein added to milk products (Cordewener et al, 2009;Lu, Liu, Gao, Lv, & Yu, 2017;Luykx et al, 2007). Highperformance liquid chromatography (HPLC)-mass spectrometry (MS) can identify numerous peptides from major seed proteins of soy and pea in the adulterated milk powder, after borate buffer extraction and tryptic digestion (Luykx et al, 2007).…”