UHPLC-HRMS (Orbitrap) fingerprinting in the classification and authentication of cranberry-based natural products and pharmaceuticals using multivariate calibration methods

UHPLC-HRMS (Orbitrap) fingerprinting in negative and positive H-ESI mode was applied to the characterization, classification and authentication of cranberry-based natural and pharmaceutical products. HRMS data in full scan mode (m/z 100-1500) at a resolution of 70,000 full-width at half maximum was recorded and processed with MSConvert software to obtain a profile of peak intensities in function of m/z values and retention times. A threshold peak filter of absolute intensity (105 counts) was applied to reduce data complexity. Principal component analysis (PCA) and partial least squares-discriminant analysis (PLS-DA) revealed patterns able to discriminate the analyzed samples according to the fruit of origin (cranberry, grape, blueberry and raspberry). Discrimination among cranberry-based natural and cranberry-based pharmaceutical preparations was also achieved. Both, UHPLC-HRMS fingerprints in negative and positive H-ESI modes, as well as the data fusion of both acquisition modes, showed to be good chemical descriptors to address cranberry extracts authentication. Validation of the proposed methodology showed a prediction rate of 100% of the samples. Obtained data was further treated by partial least squares (PLS) regression to identify frauds and quantify the percentage of adulterant fruits in cranberry-fruit extracts, achieving prediction errors in the range 0.17-3.86%

Detection and quantitation of frauds in the authentication of cranberry-based extracts by UHPLC-HRMS (Orbitrap) polyphenolic profiling and multivariate calibration methods

UHPLC-HRMS (Orbitrap) polyphenolic profiling was applied to the characterization, classification and authentication of cranberry-based natural and pharmaceutical products. 53 polyphenolic standards were characterized to build a user accurate mass database which was then proposed to obtain UHPLC-HRMS polyphenolic profiles by means of ExactFinderTM software. Principal component analysis results showed a good sample discrimination according to the fruit employed. Regarding cranberry-based pharmaceuticals, discrimination according to the presentation format (syrup, sachets, capsules, etc.) was also observed due to the enhancement of some polyphenols by purification and preconcentration procedures. Procyanidin A2 and homogenistic, sinapic, veratric, cryptochlorogenic and caffeic acids showed to be important polyphenols to achieve cranberry-based products discrimination against the other studied fruits. Partial least square regression allowed the determination of adulterant percentages in cranberry-fruit samples. Very satisfactory results, with adulteration quantification errors lower than 6.0% were obtained even at low adulteration levels

Characterization, classification and authentication of fruit-based extracts by means of HPLC-UV chromatographic fingerprints, polyphenolic profiles and chemometric methods

HPLC-UV was applied to the analysis and characterization of fruit-based and fruit-processed products. A Kinetex C18 reversed-phase column was proposed under gradient elution for the determination of 17 polyphenols. Acceptable sensitivity (LODs below 0.16 mg/L), and good linearity (r2 higher then 0.995), precision (RSD below 6.8%), and method trueness (relative errors below 11%) were obtained. Data corresponding to polyphenolic peak areas and HPLC-UV chromatographic fingerprints were then analyzed by exploratory principal component analysis (PCA) to extract information of the most significant variables contributing to characterization and classification of analyzed samples regarding the fruit of origin. HPLC-UV chromatographic data was further treated by partial least square (PLS) regression to determine the percentages of adulteration in cranberry-fruit extracts. It was found that even mixture samples containing low percentages of adulterants could be distinguished from genuine cranberry extracts. Highly satisfactory results were obtained, with overall errors in the quantification of adulterations below 4.3%

Determination of phthalic acid esters in drinking water and olive oil by ultra-high performance liquid chromatography-electrospray-tandem mass spectrometry: Study of phthalate migration from plastic bottles to drinking water at different domestic exposure conditions

A UHPLC-ESI-MS/MS method for the determination of ten phthalates in mineral water and olive oil samples was developed. A hold-back column placed between pump and injection valve allowed preventing phthalate instrumental background contamination. Good instrumental limits of detection (1-9 µg/L), precisions (RSD <23.9%), and trueness (relative error <20%) were achieved. A migration study from PET bottles into mineral water at different domestic exposure conditions revealed than only diethyl phthalate migrated when exposed at 40 oC although at not-quantified levels up to six months of exposure. No matrix effect was present for mineral waters and only dicyclohexyl phthalate was detected. Matrix-matched calibration was proposed for the analysis of olive oils due to the huge matrix-effect encountered with the employed liquid-liquid extraction method. Among the multiple positives detected in olive oil samples, only dibutyl phthalate was found in one sample at the established EU specific migration level (0.3 mg/kg)

Determination of phthalic acid esters in drinking water and olive oil by ultra-high performance liquid chromatography-electrospray-tandem mass spectrometry: Study of phthalate migration from plastic bottles to drinking water at different domestic exposure conditions

A UHPLC-ESI-MS/MS method for the determination of ten phthalates in mineral water and olive oil samples was developed. A hold-back column placed between pump and injection valve allowed preventing phthalate instrumental background contamination. Good instrumental limits of detection (1-9 µg/L), precisions (RSD <23.9%), and trueness (relative error <20%) were achieved. A migration study from PET bottles into mineral water at different domestic exposure conditions revealed than only diethyl phthalate migrated when exposed at 40 oC although at not-quantified levels up to six months of exposure. No matrix effect was present for mineral waters and only dicyclohexyl phthalate was detected. Matrix-matched calibration was proposed for the analysis of olive oils due to the huge matrix-effect encountered with the employed liquid-liquid extraction method. Among the multiple positives detected in olive oil samples, only dibutyl phthalate was found in one sample at the established EU specific migration level (0.3 mg/kg)

Detection and quantitation of frauds in the authentication of cranberry-based extracts by UHPLC-HRMS (Orbitrap) polyphenolic profiling and multivariate calibration methods

UHPLC-HRMS (Orbitrap) polyphenolic profiling was applied to the characterization, classification and authentication of cranberry-based natural and pharmaceutical products. 53 polyphenolic standards were characterized to build a user accurate mass database which was then proposed to obtain UHPLC-HRMS polyphenolic profiles by means of ExactFinderTM software. Principal component analysis results showed a good sample discrimination according to the fruit employed. Regarding cranberry-based pharmaceuticals, discrimination according to the presentation format (syrup, sachets, capsules, etc.) was also observed due to the enhancement of some polyphenols by purification and preconcentration procedures. Procyanidin A2 and homogenistic, sinapic, veratric, cryptochlorogenic and caffeic acids showed to be important polyphenols to achieve cranberry-based products discrimination against the other studied fruits. Partial least square regression allowed the determination of adulterant percentages in cranberry-fruit samples. Very satisfactory results, with adulteration quantification errors lower than 6.0% were obtained even at low adulteration levels

UHPLC-HRMS (Orbitrap) fingerprinting in the classification and authentication of cranberry-based natural products and pharmaceuticals using multivariate calibration methods

UHPLC-HRMS (Orbitrap) fingerprinting in negative and positive H-ESI mode was applied to the characterization, classification and authentication of cranberry-based natural and pharmaceutical products. HRMS data in full scan mode (m/z 100-1500) at a resolution of 70,000 full-width at half maximum was recorded and processed with MSConvert software to obtain a profile of peak intensities in function of m/z values and retention times. A threshold peak filter of absolute intensity (105 counts) was applied to reduce data complexity. Principal component analysis (PCA) and partial least squares-discriminant analysis (PLS-DA) revealed patterns able to discriminate the analyzed samples according to the fruit of origin (cranberry, grape, blueberry and raspberry). Discrimination among cranberry-based natural and cranberry-based pharmaceutical preparations was also achieved. Both, UHPLC-HRMS fingerprints in negative and positive H-ESI modes, as well as the data fusion of both acquisition modes, showed to be good chemical descriptors to address cranberry extracts authentication. Validation of the proposed methodology showed a prediction rate of 100% of the samples. Obtained data was further treated by partial least squares (PLS) regression to identify frauds and quantify the percentage of adulterant fruits in cranberry-fruit extracts, achieving prediction errors in the range 0.17-3.86%

Liver Damage using Suicide Genes : A Model for Oval Cell Activation

Liver regeneration from the facultative hepatic stem cells, the oval cells, takes place in situations in which liver regeneration from pre-existing hepatocytes is prevented. Different models have been used to stimulate oval cell response. Many of them involve the use of carcinogenic agents with or without partial hepatectomy. In this study we show that adenovirus-mediated gene transfer of the suicide gene thymidine kinase followed by ganciclovir administration caused hepatotoxicity of variable intensity. Rats with moderate elevation in serum transaminases recovered normal liver architecture few weeks after adenovirus injection. In contrast, rats with severe liver damage exhibited a marked and persisting activation of oval cells accompanied by ductular hyperplasia. In some rats, such lesion eventually evolved to cholangiofibrosis and in one rat to cholangiocarcinoma. Deposition of fibronectin and increased number of hepatic stellate cells were found in association with oval cells and cholangiofibrotic lesions. Hepatocyte growth factor was hyperexpressed in the livers with intense oval cell response or ductular proliferation, suggesting a participation of this factor in those lesions. In summary, our data demonstrate activation of oval cell response after gene transfer of thymidine kinase followed by ganciclovir administration. These findings indicate that high doses of this therapy causes liver damage together with an impairment in hepatocellular regeneration

Characterization, classification and authentication of fruit-based extracts by means of HPLC-UV chromatographic fingerprints, polyphenolic profiles and chemometric methods

HPLC-UV was applied to the analysis and characterization of fruit-based and fruit-processed products. A Kinetex C18 reversed-phase column was proposed under gradient elution for the determination of 17 polyphenols. Acceptable sensitivity (LODs below 0.16 mg/L), and good linearity (r2 higher then 0.995), precision (RSD below 6.8%), and method trueness (relative errors below 11%) were obtained. Data corresponding to polyphenolic peak areas and HPLC-UV chromatographic fingerprints were then analyzed by exploratory principal component analysis (PCA) to extract information of the most significant variables contributing to characterization and classification of analyzed samples regarding the fruit of origin. HPLC-UV chromatographic data was further treated by partial least square (PLS) regression to determine the percentages of adulteration in cranberry-fruit extracts. It was found that even mixture samples containing low percentages of adulterants could be distinguished from genuine cranberry extracts. Highly satisfactory results were obtained, with overall errors in the quantification of adulterations below 4.3%

Supplementation of diets for weaned piglets withL-Valine and L-Glutamine+ L-Glutamic acid

The objective of this study was to evaluate the effects of supplementation of diets for weaned piglets with L-valine and L-glutamine + L-glutamic acid on performance, frequency of diarrhea, organ weight, digesta pH, intestinal morphology, and economic viability. Seventy-two piglets with a live weight of 7.53 ± 0.84 kg and 24 days of age were used. The animals were submitted to the following four treatments from 24 to 46 days of age: diet not supplemented with amino acids (control diet, CD); diet supplemented with glutamine + glutamic acid (GD); diet supplemented with glutamine + glutamic acid + valine (GVD), and diet supplemented with valine (VD). Two sequential phases (pre-initial I and pre-initial II) with a duration of 12 and 11 days, respectively, were established. A completely randomized design, consisting of six repetitions and three pigs per experimental unit, was used. Nine days after weaning, at 32 days of age, a piglet per pen was slaughtered for the evaluation of organ weight, digesta pH and intestinal morphology. All animals received a single diet from days 47 to 65. No effects on performance were observed during the pre-initial phases I and II; however, when the whole study period was considered (24 to 65 days of age), piglets fed GVD consumed less feed and exhibited better feed conversion than animals of the VD group. With respect to morphometric parameters, GD provided a greater ileal crypt depth than CD and VD. There was an economic advantage of diets supplemented with L-valine and L-glutamine + L-glutamic acid, validating their use in diets for weaned piglets until 46 days of age.</span