A new ultrahigh performance liquid chromatography with diode array detection coupled to electrospray ionization and quadrupole time-of-flight mass spectrometry analytical strategy for fast analysis and improved characterization of phenolic compounds in apple products.

A new, rapid, selective and sensitive ultrahigh performance liquid chromatography with diode array detection coupled to electrospray ionization and quadrupole time-of-flight mass spectrometry (UHPLC-DAD-ESI-Q-ToF-MS) strategy using automatic and simultaneous acquisition of exact mass at high and low collision energy, MS(E), has been developed to obtain polyphenolic profile of apples, apple pomace and apple juice from Asturian cider apples in a single run injection of 22 min. MS(E) spectral data acquisition overcomes chromatographic co-elution problems, performing simultaneous collection of precursor ions as well as other ions produced as a result of their fragmentation, which allows resolving complex spectra from mixtures of precursor ions in an unsupervised way and eases their interpretation. Using this technique, 52 phenolic compounds of five different classes were readily characterized in these apple extracts in both positive and negative ionization modes. The spectral data for phenolic compounds obtained using this acquisition mode are comparable to those obtained by conventional LC-MS/MS as exemplified in this work. Among the 52 phenolic compounds identified in this work, 2 dihydrochalcones and 3 flavonols have been tentatively identified for the first time in apple products. Moreover, 2 flavanols, 4 dihydrochalcones, 9 hydroxycinnamic acids and 4 flavonols had not been previously reported in apple by ToF analysis to our knowledge.

Improvement using chemometrics in ion mobility coupled to mass spectrometry as a tool for mass spectrometry fragmentation studies: flavonoid aglycone cases.

A chemometric method for the treatment of ion mobility coupled to mass spectrometry (IMS/MS) data is proposed as a complementary tool for obtaining experimental evidence for the study of MS fragmentations, which can provide a direct and automatable methodology for characterising ionic series and the hierarchy of all product ions of an MS spectrum. Two MS/MS with ion mobility experiments have been designed: in the first, the intrinsic mobility of each ion is estimated, and in the second experiment, distributions of the ionic intensity of product ions fragmented after IMS separations are recorded. These mobilograms are aligned using the coshift algorithm and mathematically fitted using Classical Least Squares (CLS) to determine the mobility contributions from their precursor ions. Despite some limitations when studying low intensity ions and ions with similar ion mobility, CLS fitting improves the usage of IMS coupled with accurate mass spectrometry as a complementary tool in the study of MS fragmentation mechanisms and more notably, it offers an automatable and efficient alternative to MS3 experiments.

Feasibility study of FT-MIR spectroscopy and PLS-R for the fast determination of anthocyanins in wine.

The feasibility of using Fourier Transform Mid-Infrared Spectroscopy (FT-MIR) combined with Partial Least Squares Regression (PLS-R) for the determination of 12 anthocyanins (3-O-glucosides of delphinidin, cyanidin, petunidin, peonidin and malvidin, as well as acetic acid esters and p-coumaric acid esters of petunidin, peonidin and malvidin and caffeic acid ester of malvidin) and three sums (sum of non-acylated anthocyanins, sum of acetylated anthocyanins and sum of coumaroylated anthocyanins), in red wines has been tested. Reference values of anthocyanin concentrations by reverse-phase High Performance Liquid Chromatography with Diode Array Detection (HPLC-DAD) were used to calibrate the models. A Principal Component Analysis (PCA) was applied to these reference values and a differentiation of wine samples by wine type (young wines of 2005, young wines of 2004 and crianza and reserva wines) has been possible. A calibration model using PLS-R was built with 153 samples of Rioja wines and the prediction of the anthocyanin concentrations using this model was evaluated by internal and external validation sample sets. Most of the anthocyanins and their sums have been predicted with a Standard Error of Prediction (SEP) of 15-30% for young wines recently bottled. However, for young wines after one year of being bottled, and for crianza and reserva wines, these errors were unacceptable. The obtained results suggest that the model built for FT-IR instrument calibration is a useful tool for a quick determination of the anthocyanin content of young wines of the current vintage, but a careful robust external validated calibration of the technique is necessary in order to maintain the prediction errors within controlled limits.

Practical guidelines for characterization of O-diglycosyl flavonoid isomers by triple quadrupole MS and their applications for identification of some fruit juices flavonoids.

Fifteen flavonoid O-diglycosides with different interglycosidic linkage isomery and glycosylation position have been studied in order to analyze their fragmentation patterns. Initial separation was carried out using high performance liquid chromatography with diode array detection (HPLC/DAD) coupled to an electrospray ionization (ESI) interface and a triple quadrupole mass spectrometer. Some useful differences in their MS spectra have been found and discussed. As it has already been reported, [Y*]+/[Y0]+ ratio for flavanones and [Y1]+/[Y0]+ ratio for other flavonoids is specific for each isomeric interglycosidic linkage. In this work it has also been observed that the abundance of these ions is dependent on the position of glycosylation. On the basis of these differences, systematic guidelines for our experimental conditions have been proposed for the differentiation of not only isomeric interglycosidic linkage but also glycosylation position using collision-induced dissociation MS/MS (CID-MS/MS) spectra in positive mode. These results have been successfully applied for the characterization of three diglycosyl flavonoids found in Citrus fruit juices and these conclusions have also been extrapolated for characterizing two triglycosides in the same fruits.