Daniel Esquivel-Alvarado

Liquid chromatography-mass spectrometry (LC-MS)-based metabolomics has become an important tool to increase our understanding of how diet affects human health. However, public and commercial mass spectral libraries of dietary metabolites are limited, resulting in the greatest challenge in converting mass spectrometry data into biological insights. In this study, we constructed an LC-MS/MS ginger library as an example to demonstrate the importance of dietary libraries for discovering food biomarkers. The functional and exposure biomarkers of ginger were investigated using plasma samples from mice treated with control and ginger extract diets. Our results showed clear discrimination between the metabolome of mice on normal and ginger extract diets. Using the in-house ginger library, we identified 20 ginger metabolites that can be used as exposure biomarkers of ginger. However, without the LC-MS/MS ginger library, none of the ginger metabolites could be accurately identified based on online mass databases. In addition, ginger treatment significantly impacts the endogenous metabolome, especially the purine metabolism and phenylalanine, tyrosine, and tryptophan biosynthesis. Overall, we demonstrated that the construction of LC-MS/MS spectra dietary libraries would enhance the ability to identify potential dietary biomarkers and correlate potential health benefits associated with food consumption.

The interaction between A‐type interflavan bonds from cranberry proanthocyanidins (PAC) and surface virulence factors of extra‐intestinal pathogenic Escherichia coli (ExPEC) was studied. Electrospun nanofibers (ESNF) were fabricated using PAC and polycaprolactone (PCL) solutions and their physical and chemical properties were characterized. The ability of PAC:PCL composite ESNF to interact with and entrap ExPEC strain 5011 (ExPEC‐5011) was evaluated in vitro by plate culturing and when formulated as a biofilter and nanocoating. As a biofilter, the PAC:PCL ESNF exhibited a dose‐dependent ability to entrap ExPEC‐5011. Images from scanning electron and fluorescent microscopies revealed that ESNF sections with higher amounts of PAC led to higher bacterial entrapment. The effectiveness PAC:PCL ESNF to bind ExPEC when applied as a nanocoating was studied using ESNF‐coated polyvinyl chloride intermittent catheter. Results indicate that ExPEC‐5011 was entrapped well into the PAC:PCL ESNF coating on the catheter. Overall, our results suggest that incorporating the biomolecule PAC in ESNF is a potential means for applications requiring bacterial entrapment, such as biofunctionalization, biofiltration, and surface coating, among others.

Synthesis of proanthocyanidin-cinnamaldehydes pyrylium products (PCPP) was achieved by the condensation reaction of proanthocyanidins (PAC) with cinnamaldehyde and four cinnamaldehyde derivatives. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) spectra of PCPP show masses that correspond to (epi)catechin oligomers attached to single, double, or triple moieties of cinnamaldehydes. Synthesized PCPP exhibited fluorescence at higher excitation and emission wavelengths than PAC. Results indicate that PCPP were more bioactive for agglutinating extra-intestinal pathogenic Escherichia coli (ExPEC) compared to PAC. Scanning electron microscopy indicates that PCPP interact with ExPEC surface structures and suggests that PCPP have a higher affinity with the fimbriae-like structures of ExPEC than PAC. Fluorescent microscopy performed on in vitro and in vivo agglutination assays show that PCPP were entrapping ExPEC in a web-like network, thus demonstrating agglutination of ExPEC. This study demonstrated the potential of PCPP to improve our understanding of the temporal and dynamic interactions of PAC in in vitro and in vivo studies.

Background: Cranberry proanthocyanidins (c-PAC) are oligomeric structures of flavan-3-ol units, which possess A-type interflavan bonds. c-PAC differs from other botanical sources because other PAC mostly have B-type interflavan bonds. Cranberry products used to alleviate and prevent urinary tract infections may suffer from adulteration, where c-PAC are replaced with less expensive botanical sources of PAC that contain B-type interflavan bonds. Objective: Identifying the presence of A-type interflavan bonds in cranberry fruit and dietary supplements. Methods: Thirty-five samples reported to contain A-type PAC (cranberry fruit and cranberry products) and 36 samples reported to contain B-type PAC (other botanical sources) were identified and differentiated using MALDI-TOF MS, deconvolution of overlapping isotope patterns, and principal component analysis (PCA). Results: Our results show that both MALDI-TOF MS and deconvolution of overlapping isotope patterns were able to identify the presence of A-type interflavan bonds with a probability greater than 90% and a confidence of 95%. Deconvolution of MALDI-TOF MS spectra also determined the ratio of A-type to B-type interflavan bonds at each degree of polymerization in cranberry fruit and cranberry products, which is a distinguishing feature of c-PAC in comparison to other botanical sources of PAC. PCA shows clear differences based on the nature of the interflavan bonds. Conclusions: MALDI-TOF MS, deconvolution of overlapping isotope patterns of MALDI-TOF MS spectra, and PCA allow the identification, estimation, and differentiation of A-type interflavan bonds in cranberry-based foods and dietary supplements among other botanical sources containing mostly B-type interflavan bonds.

Proanthocyanidins (PAC) are oligomers and polymers of flavan-3-ols with putative health benefits. PAC are prevalent in a wide variety of natural products and dietary supplements. An inter-laboratory study was conducted to validate the 4-(dimethylamino)cinnamaldehyde (DMAC) colorimetric assay using a 96-well plate spectrophotometer for the accurate quantification of proanthocyanidins (PAC) in cranberry products and to evaluate the comparison of the procyanidin A2 (ProA2) dimer and cranberry PAC (c-PAC) reference standards. Four test materials analyzed in this study included cranberry fiber powder, cranberry extract powder, concentrated cranberry juice, and a solution of cranberry PAC (30%, w/v). The samples were homogenized, extracted, sonicated, centrifuged, and analyzed using a 96 well plate spectrophotometer. the linearity for both ProA2 and c-PAC standards were determined from 4.053 to 50.666 µg/mL and from 13.520 to 135.95 µg/mL, respectively. The relative standard deviation of repeatability (RSD_r) values for the four materials analyzed using both ProA2 and c-PAC standards met the Standard Method Performance Requirements (SMPR®). The inter-laboratory precision using the Horwitz ratio (HorRat) values for the four materials analyzed using both ProA2 and c-PAC standards satisfies the acceptance range in Appendix K of the Official Methods of Analysis (2003): Guidelines for Dietary Supplements and Botanicals. The limit of quantification (LOQ) was estimated to be 3.16 µg/mL. The results produced from this study demonstrate the utility of the c-PAC standard over the ProA2 standard and the advantages of using a 96-well plate spectrophotometer for the accurate quantification of PAC.