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Miscellaneous: In-Vitro

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Development of a Cranberry Standard for Quantification of Insoluble Cranberry (Vaccinium macrocarpon Ait.) Proanthocyanidins.

Posted: 
August 29, 2019
Authors: 
Gullickson ER; Krueger CG; Birmingham A; Maranan M; Reed JD.
Journal: 
Journal of Agricultural & Food Chemistry 10.1021/acs.jafc.9b03696 [doi]
Abstract: 

Cranberry proanthocyanidins (PACs) can be partitioned into soluble PACs, which are extracted with solvents, and insoluble PACs, which remain associated with fibers and proteins after extraction. Most research on cranberry products only quantifies soluble PACs because proper standards for quantifying insoluble PACs are lacking. In this study, we evaluated the ability of a cranberry PAC (c-PAC) standard, reflective of the structural heterogeneity of PACs found in cranberry fruit, to quantify insoluble PACs by the butanol-hydrochloric acid (BuOH-HCl) method. For the first time, a c-PAC standard enabled conversion of BuOH-HCl absorbance values (550 nm) to a weight (milligram) basis, allowing for quantification of insoluble PACs in cranberries. The use of the c-PAC reference standard for sequential analysis of soluble PACs by the method of 4-(dimethylamino)cinnamaldehyde and insoluble PACs by the method of BuOH-HCl provides analytical tools for the standardization of cranberry-based ingredients.

Evaluation of the Degree of Polymerization of the Proanthocyanidins in Cranberry by Molecular Sieving and Characterization of the Low Molecular Weight Fractions by UHPLC-Orbitrap Mass Spectrometry.

Posted: 
August 29, 2019
Authors: 
Gardana, C. Simonetti, P.
Journal: 
Molecules. 24(8):1504
Abstract: 

4-dimethylammino-cinnamaldehyde (DMAC) assays quantify total proanthocyanidins (PACs) but do not provide qualitative PAC molecular weight distribution information and cannot discriminate between A- and B-type PACs. We developed an efficient method for assessing PAC molecular weight distributions. The PACs from three commercial cranberry extracts (A1-A3) were fractionated by molecular sieves with cut-offs of 3, 10, 30, 50, and 100 kDa, and each fraction was analyzed by DMAC assays. A1, A2, and A3 contained 27%, 33%, and 15% PACs, respectively. Approximately 28 PACs, 20 flavonols, and 15 phenolic acids were identified by UHPLC-DAD-Orbitrap MS in A1 and A3, while A2 contained only flavan-3-ols. Epicatechin was the main monomer in A1 and A3, and catechin was the main in A2. Procyanidin A2 was the main dimer in A1 and A3, representing more than 85% of the total dimers, while it constituted approximately only 24% of A2. A1 and A3 contained quercetin, isorhamnetin, myricetin, and their glycosides, which were totally absent in A2. In A1 and A3 the PACs were mainly distributed in the fractions 30-3 and <3 kDa, while in A2 more than 70% were present in the fraction less than 3 kDa. Overall, obtained data strongly suggests that A2 is not cranberry-derived, or is adulterated with another source of PACs.

Antioxidant Properties and Phenolic Compounds of Vitamin C-Rich Juices.

Posted: 
February 19, 2019
Authors: 
Nowak, D. Goslinski, M. Wojtowicz, E. Przygonski, K.
Journal: 
Journal of Food Science. 83(8):2237-2246.
Abstract: 

Many studies have shown that bioactive compounds, for example, polyphenols, and so on can play an important role in reducing oxidative stress and protect against various diseases. The sources of these compounds in the human diet include mainly fruit and good quality fruit juices, which may contain polyphenols but also other phytochemicals such as vitamin C. The purpose of the study was to analyze the antioxidant properties of vitamin C-rich juices, which underwent mild processing. The content of total polyphenols (TP, FBBB), total flavonoids (TF), total anthocyanins (TA), and vitamin C as well as the antioxidant capacity (DPPH, ABTS) were evaluated in commercial fruit juices rich in vitamin C (acerola, gojiberry, sea buckthorn, wild rose, cranberry, Japanese quince). Moreover, phenolic acids and selected flavonoids were determined by HPLC methods. Among the examined fruit juices, acerola and wild rose juices contained the highest amounts of vitamin C and total polyphenols, and had the highest antioxidant capacity. Acerola owes its high antioxidant properties mainly to vitamin C, whereas the antioxidant capacity of wild rose is also attributed to its rich content of flavonoids and phenolic acids. Sea buckthorn juice and Japanese quince juice had a lower antioxidant capacity, yet higher than determined for gojiberry and cranberry juices. Total anthocyanins were the highest in cranberry juice. The results showed that the analyzed juices were a valuable source of natural antioxidants. Generally, vitamin C-rich juices are also good source of polyphenols. Vitamin C and polyphenols act synergistically and define the antioxidant properties of juices.

BIOACTIVE COMPONENTS OF VACCINIUM MACROCARPON AND ITS ANTIOXIDANT ACTIVITY: AN IN-VITRO STUDY

Posted: 
February 19, 2019
Authors: 
Krishnaeswari V, Manikandan S, Vijayakumar J
Journal: 
International Journal of Pharmaceutical Sciences and Research, DOI: 10.13040/IJPSR.0975-8232.10(1).438-44
Abstract: 

Cranberries (Vaccinium macrocarpon) contain many bioactive compounds and have some biological activities and beneficial health properties. This study aimed to screen phytochemicals of cranberry fruits from the different solvent, to estimate the total phenolic and flavonoid content of cranberry fruits and their antioxidant effect in-vitro by DPPH, superoxide and nitric oxide radical scavenging assay. Phytochemical screening of various extracts such as aqueous, ethanol, chloroform, acetone and petroleum ether of cranberry fruit extracts, revealed the presence of flavonoids, cardiac glycosides, phenols, coumarins, terpenoids, and betacyanin. The cranberry extracts were evaluated for phenol and flavonoid content with Gallic acid (GA) and Quercetin (Q) as standard. The optimum yield of phenol and flavonoid content were found in ethanol fruit extract 13.07 mg Gallic acid Equivalents (GAE)/g and 9.02 mg Quercetin Equivalents (QE)/g of cranberry. The cranberry extracts were evaluated for antioxidant activities by DPPH (1,1– diphenyl -2- picrylhydrazyl) radical scavenging assay. Among five different solvents used, maximum antioxidant activity was found in ethanolic fruit extract (81.4%) followed by others. The IC50 values of ethanolic cranberry extract in superoxide radical scavenging activity and Nitric oxide radical scavenging assay are 61.1 µg/ml and 54.7 µg/ml. The IC50 values showed a strong antioxidant activity of the extracts. The powerful antioxidant effect attributed to the greater amount of phenol and flavonoid compound in the ethanolic cranberry extract.

Comparative Evaluation of Grape Seed and Cranberry Extracts in Preventing Enamel Erosion: An Optical Emission Spectrometric Analysis.

Posted: 
February 19, 2019
Authors: 
Nandakumar M, Nasim I.
Journal: 
J Conserv Dent. 2018 Sep-Oct;21(5):516-520. doi: 10.4103/JCD.JCD_110_18.
Abstract: 

INTRODUCTION:Dental erosion is defined as the loss of tooth structure due to chemical process that does not involve bacteria. The management of such a condition calls for a comprehensive approach to identifying the cause and treating it.AIM:The aim of this study is to comparatively evaluate the role of grape seed extract (GSE) and cranberry extract (CE) in preventing dental erosion using optical emission spectrometry.MATERIALS AND METHODS:Prepared enamel specimens were subjected to the erosive challenge using HCl for 10 s, followed by immersion in experimental natural groups and control fluoride group for 30 s and artificial saliva for 60 min. This cycle was repeated three times. The amounts of calcium and phosphorous present in the acid solution after 1st, 2nd, and 3rd erosive challenges were determined for each group using induced coupled plasma-optical emission spectrometry.RESULTS:The cumulative calcium and phosphorous release after the 1st, 2nd, and 3rd erosive challenges were found to be the least in SnF2 group, followed by GSE group and then in CE group.CONCLUSION:The protective of GSE and CE was inferior to the gold standard control group of stannous fluoride role, against enamel erosion. GSE showed better remineralizing effect; however, there was no statistically significant difference between the two groups.

Some New Findings Regarding the Antiadhesive Activity of Cranberry Phenolic Compounds and Their Microbial-Derived Metabolites against Uropathogenic Bacteria.

Posted: 
February 19, 2019
Authors: 
González de Llano D, Liu H, Khoo C, Moreno-Arribas MV, Bartolomé B.
Journal: 
J Agric Food Chem. 2019 Feb 12. doi: 10.1021/acs.jafc.8b05625
Abstract: 

Findings concerning the antiadhesive activity of cranberry phenolic compounds and their microbial-derived metabolites against Gram-negative ( Escherichia coli ATCC 53503 and DSM 10791) and Gram-positive ( Enterococcus faecalis 04-1) bacteria in T24 cells are reported. A-Type procyanidins (A2 and cinnamtannin B-1) exhibited antiadhesive activity (at concentrations ≥250 μM), a feature that was not observed for B-type procyanidins (B2). The metabolites hippuric acid and α-hydroxyhippuric acid also showed effective results at concentrations ≥250 μM. With regard to conjugated metabolites, sulfation seemed to increase the antiadhesive activity of cranberry-derived metabolites as 3-(3,4-dihydroxyphenyl)propionic acid 3- O-sulfate presented active results, unlike its corresponding nonsulfated form. In contrast, methylation decreased antiadhesive activity as 3,4-dihydroxyphenylacetic acid was found to be active but not its corresponding methylated form (4-hydroxy-3-methoxyphenylacetic acid). As a whole, this work sustains the antiadhesive activity of cranberry-derived metabolites as one of the mechanisms involved in the beneficial effects of cranberries against urinary tract infections.

Advantages of a Validated UPLC-MS/MS Standard Addition Method for the Quantification of A-Type Dimeric and Trimeric Proanthocyanidins in Cranberry Extracts in Comparison with Well-Known Quantification Methods.

Posted: 
September 4, 2018
Authors: 
Dooren, I. van Foubert, K. Theunis, M. Naessens, T. Pieters, L. Apers, S.
Journal: 
Journal of Pharmaceutical and Biomedical Analysis;148:32-41.
Abstract: 

The berries of Vaccinium macrocarpon, cranberry, are widely used for the prevention of urinary tract infections. This species contains A-type proanthocyanidins (PACs), which intervene in the initial phase of the development of urinary tract infections by preventing the adherence of Escherichia coli by their P-type fimbriae to uroepithelial cells. Unfortunately, the existing clinical studies used different cranberry preparations, which were poorly standardized. Because of this, the results were hard to compare, which led sometimes to conflicting results. Currently, PACs are quantified using the rather non-specific spectrophotometric 4-dimethylaminocinnamaldehyde (DMAC) method. In addition, a normal phase HPTLC-densitometric method, a HPLC-UV method and three LC-MS/MS methods for quantification of procyanidin A2 were recently published. All these methods contain some shortcomings and errors. Hence, the development and validation of a fast and sensitive standard addition LC-MS/MS method for the simultaneous quantification of A-type dimers and trimers in a cranberry dry extract was carried out. A linear calibration model could be adopted for dimers and, after logarithmic transformation, for trimers. The maximal interday and interconcentration precision was found to be 4.86% and 4.28% for procyanidin A2, and 5.61% and 7.65% for trimeric PACs, which are all acceptable values for an analytical method using LC-MS/MS. In addition, twelve different cranberry extracts were analyzed by means of the newly validated method and other widely used methods. There appeared to be an enormous variation in dimeric and trimeric PAC content. Comparison of these results with LC-MS/MS analysis without standard addition showed the presence of matrix effects for some of the extracts and proved the necessity of standard addition. A comparison of the well-known and widely used DMAC method, the butanol-HCl assay and this newly developed LC-MS/MS method clearly indicated the need for a reliable method able to quantify A-type PACs, which are considered to be the pharmacologically active constituents of cranberry, since neither the DMAC or butanol-HCl assays are capable of distinguishing between A and B-type PACs and therefore cannot detect adulterations with, for example, extracts with a high B-type PAC content. Hence, the combination of the DMAC method or butanol-HCl assay with this more specific LC-MS/MS assay could overcome these shortcomings.

Determination of Flavanols by Liquid Chromatography with Fluorescence Detection. Application to the Characterization of Cranberry-Based Pharmaceuticals Through Profiling and Fingerprinting Approaches.

Posted: 
September 4, 2018
Authors: 
Bakhytkyzy, I. Nunez, O. Saurina, J.
Journal: 
Journal of Pharmaceutical and Biomedical Analysis; 2018. 156:206-213.
Abstract: 

In this work, a new method based on reversed-phase high-performance liquid chromatography (HPLC) with fluorescence detection (FLD) was established for the determination of catechins and related oligomeric proanthocyanidins (PACs) in cranberry-based pharmaceuticals. Compounds were recovered by liquid extraction using methanol/water/hydrochloric acid (60:39:1, v:v:v) as the extraction solvent. The chromatographic separation was carried out using a core-shell C18 column under an elution program based on 0.1% formic acid in water and methanol as the components of the mobile phase. The flow rate was 0.4 mL min-1 and the injection volume was 5 micro L. Chromatograms were acquired at 280 nm by UV-vis absorption and at lambda ex 280 nm and lambda em 347 nm by fluorescence spectroscopy. Compared to UV detection, FLD demonstrated both increased sensitivity and selectivity to avoid interfering signals from other phenolic compounds present in the samples. Data resulting from the analysis of cranberry-based products was exploited to tackle an exploratory characterization and classification using principal component analysis. Samples were clustered according to their compositions and those enriched with PACs with antibacterial activity were clearly distinguished from the others.

Development of a Thiolysis HPLC Method for the Analysis of Procyanidins in Cranberry Products.

Posted: 
September 4, 2018
Authors: 
Gao C; Cunningham DG; Liu H; Khoo C; Gu L.
Journal: 
Journal of Agricultural & Food Chemistry. 66(9):2159-2167
Abstract: 

The objective of this study was to develop a thiolysis HPLC method to quantify total procyanidins, the ratio of A-type linkages, and A-type procyanidin equivalents in cranberry products. Cysteamine was utilized as a low-odor substitute of toluene-alpha-thiol for thiolysis depolymerization. A reaction temperature of 70 degreeC and reaction time of 20 min, in 0.3 M of HCl, were determined to be optimum depolymerization conditions. Thiolytic products of cranberry procyanidins were separated by RP-HPLC and identified using high-resolution mass spectrometry. Standards curves of good linearity were obtained on thiolyzed procyanidin dimer A2 and B2 external standards. The detection and quantification limits, recovery, and precision of this method were validated. The new method was applied to quantitate total procyanidins, average degree of polymerization, ratio of A-type linkages, and A-type procyanidin equivalents in cranberry products. Results showed that the method was suitable for quantitative and qualitative analysis of procyanidins in cranberry products.

Evaluation of Polyphenol Anthocyanin-Enriched Extracts of Blackberry, Black Raspberry, Blueberry, Cranberry, Red Raspberry, and Strawberry for Free Radical Scavenging, Reactive Carbonyl Species Trapping, Anti-Glycation, Anti- Beta -Amyloid Aggregation.

Posted: 
September 4, 2018
Authors: 
Ma H, Johnson SL, Liu W, DaSilva NA, Meschwitz S, Dain JA, Seeram NP
Journal: 
International Journal of Molecular Sciences;19(2):461.
Abstract: 

Glycation is associated with several neurodegenerative disorders, including Alzheimer's disease (AD), where it potentiates the aggregation and toxicity of proteins such as beta -amyloid (A beta ). Published studies support the anti-glycation and neuroprotective effects of several polyphenol-rich fruits, including berries, which are rich in anthocyanins. Herein, blackberry, black raspberry, blueberry, cranberry, red raspberry, and strawberry extracts were evaluated for: (1) total phenolic and anthocyanins contents, (2) free radical (DPPH) scavenging and reactive carbonyl species (methylglyoxal; MGO) trapping, (3) anti-glycation (using BSA-fructose and BSA-MGO models), (4) anti-A beta aggregation (using thermal- and MGO-induced fibrillation models), and, (5) murine microglia (BV-2) neuroprotective properties. Berry crude extracts (CE) were fractionated to yield anthocyanins-free (ACF) and anthocyanins-enriched (ACE) extracts. The berry ACEs (at 100 micro g/mL) showed superior free radical scavenging, reactive carbonyl species trapping, and anti-glycation effects compared to their respective ACFs. The berry ACEs (at 100 micro g/mL) inhibited both thermal- and MGO-induced A beta fibrillation. In addition, the berry ACEs (at 20 micro g/mL) reduced H2O2-induced reactive oxygen species production, and lipopolysaccharide-induced nitric oxide species in BV-2 microglia as well as decreased H2O2-induced cytotoxicity and caspase-3/7 activity in BV-2 microglia. The free radical scavenging, reactive carbonyl trapping, anti-glycation, anti-A beta fibrillation, and microglial neuroprotective effects of these berry extracts warrant further in vivo studies to evaluate their potential neuroprotective effects against AD.

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