Caffeoylquinic Acid Isomers Research Articles

Fully automatic quantitation of eight different metabolites in coffee using 1H‐NMR spectroscopy and the PULCON methodology

AbstractBackgroundCoffee contains a plethora of constituents with some of them being especially important either due to their physiological effects or as quality markers. As quantitative proton nuclear magnetic resonance spectroscopy (1H‐NMR) has been established as a fast and reliable analytical tool its application was evaluated for the simultaneous quantitation of lactic acid, acetic acid, formic acid, caffeine, caffeoylquinic acid (CQA) isomers, N‐methylpyridinium, trigonelline, and 5‐hydroxymethylfurfural (HMF) in aqueous extracts of roasted Coffea arabica samples.ResultsSimultaneous quantitative determination was achieved by an automated analysis based on the PULCON methodology (pulse length‐based concentration determination). The method was validated regarding linearity, accuracy, precision, limit of detection (LOD), and limit of quantitation (LOQ). Recovery rates were between 76% (CQA) and 116% (HMF), and precision was between 1.7% (caffeine) and 10.3% (HMF). The LOD varied between 0.06 g/kg (HMF) and 1.35 g/kg (caffeine and CQA), with the LOQ being between 0.22 g/kg (HMF) and 4.87 g/kg (CQA). To verify the results of the 1H‐NMR method, caffeine, trigonelline, HMF, 3‐CQA, 4‐CQA, and 5‐CQA were additionally quantitated by HPLC‐DAD and the results were compared. The described 1H‐NMR method was additionally applied to coffee samples that contained different coffee defects. Results showed only slight changes in the concentrations of the analytes by adding defective beans to defect‐free coffee.DiscussionThe developed 1H‐NMR approach was proven to be fast (30 min), reliable, and precise. Thus, it is well suited to analyze several coffee constituents of interest in a large number of samples in, for example, quality control.

Protection against myocardial ischemia/reperfusion injury in mice by 3-caffeoylquinic acid isomers isolated from Saxifraga tangutica.

The development of ischemic heart disease (IHD) involves a variety of pathophysiological responses, such as mitochondrial dysfunction. Many compounds with antioxidant activity isolated from natural products have been shown to have significant effects on the prevention and treatment of cardiovascular diseases. However, little is known about the palliative effects of 3-caffeoylquinic acid isomers isolated from Saxifraga tangutica (S. tangutica) on myocardial ischemia/reperfusion injury (MIRI). Three isomers of 3-caffeoylquinic acid were isolated from S. tangutica and identified as neochlorogenic acid (Fr2-4-1-1, 18.5 mg), chlorogenic acid (Fr2-5-1-1, 81.7 mg) and cryptochlorogenic acid (Fr2-5-2-1, 15.0 mg) using medium-pressure liquid chromatography-high-pressure two-dimensional liquid chromatography. An in vitro DPPH assay showed that cryptochlorogenic acid (CCGA), neochlorogenic acid (NCGA) and chlorogenic acid (CGA) (in order of activity from strongest to weakest) possessed superior antioxidant activity. Langendorff's in vitro model was utilized to explore the protective effects of 3 caffeoylquinic acid isomers against MIRI. The ex vivo MIRI assay demonstrated that CCGA significantly improved hemodynamic function (P < 0.05), hemodynamic function-related indices (LVDP, RPP, +dP/dt and -dP/dt), and cell morphology in I/R myocardium tissues. In addition, the results of western blot analysis showed that mitochondrial biogenesis was significantly increased in I/R myocardial tissues after treatment with CCGA. In contrast, the activities of CGA and NCGA were lower. This is the first demonstration of efficient preparative isolation of 3-caffeoylquinic acid isomers (CGA, NCGA and CCGA) from S. tangutica. CCGA may be a promising approach for the treatment of cardiac I/R injury, especially for the regulation of mitochondrial biogenesis after MIRI.

Antioxidant Response, Phenolic Compounds and Yield of Solanum tuberosum Tubers Inoculated with Arbuscular Mycorrhizal Fungi and Growing under Water Stress.

Solanum tuberosum (potato) is one of the most common crops worldwide; however, it is sensitive to water stress, which necessitates the identification of alternative tools to improve their production. Here, we evaluated the inoculation of two arbuscular mycorrhizal fungi (AMF) strains, Claroideoglomus claroideum (CC), Claroideoglomus lamellosum (HMC26), and the MIX (CC + HMC26) in yield and phenolic and antioxidant response using chromatographic and spectroscopic methods in potato crops, at increasing levels of water stress, namely, with 100% (0), 70% (S1), and 40% (S2) soil humidity. Two caffeoylquinic acid isomers were detected and their levels showed a tendency to increase under stress together with the AMF inoculation, reaching up to 19.2 mg kg-1 of 5-caffeoylquinic acid and 7.4 mg kg-1 of caffeoylquinic acid isomer when CC was inoculated, and potato plants grew at the highest water starvation condition (S2). Regarding antioxidant activities, a differentiated response was detected depending on the AMF strain, highlighting the effect of HMC26 on Trolox equivalent antioxidant capacity (TEAC) method and CC in cupric reducing antioxidant capacity (CUPRAC) method, reaching up to 1.5 μmol g-1 of TEAC in plants inoculated with HMC26 and 0.9 μmol g-1 of CUPRAC in plants inoculated with CC, both in potato tubers of plants growing under the S2 stress condition. Meanwhile, the use of AMF did not influence the number and biomass of the tubers, but significant changes in the biochemical properties of tubers were observed. The results suggest that specific AMF adaptations to water stress must be considered when inoculation procedures are planned to improve the yield and quality of tubers in potato crops.

Manufacture of active multilayer films made of functionalized pectin coated by polyhydroxyalkanoates: A fully renewable approach to active food packaging

Biodegradable active packaging, i.e. materials able to promote food preservation while avoiding plastic waste accumulation are expecting to play a key role fsor the manufacture of new generation materials.Bioactive films composed of citrus pectin (CP) added with extracts from spent coffee grounds (SCGs) were herein developed. To address the limitations of pectin-based materials, i.e. hydrophilicity and poor water barrier properties, an active multilayer film was prepared by coating the functionalized pectin by a two-steps dipping into polyhydroxyalkanoates (PHA) solutions characterized by different monomeric compositions. The PHA coating affected the stiffness of the film and its opacity, while improving its hydrophobicity and water vapor permeability. The dipping procedure did not compromise the stability of the bioactive compounds, rather it allowed to preserve their antioxidant and antimicrobial activities for longer times in comparison with uncoated functionalized film. Further, the multilayer material functionalized with extracts was able to delay the carotenoid degradation in mashed carrots likely because of the increased film opacity conferred by both phenols (caffeoylquinic acid isomers including chlorogenic acid) found in SCGs and PHAs layer suggesting that the new material may be used to extend the food shelf-life and potentially to enhance product health credentials.

A novel basis for monitoring the coffee roasting process: Isomerization reactions of 3-caffeoylquinic and 4-caffeoylquinic acids

The aim of this study was to investigate the changes of individual chlorogenic acids during the coffee roasting process under authentic commercial conditions. Unique chemical reactions of caffeoylquinic acid (CQA) and dicaffeoylquinic acid (diCQA) isomers were shown in different Coffea species (C. arabica vs. C. canephora), batch sizes (1 kg vs. 20 kg) and roaster designs. Temperature determination setups of different roaster machines appeared gave varying results. Isomerization reactions of CQA in C. arabica beans were observed between the end of the drying phase and the crack reaction. Regarding C. canephora beans, the slower degradation rate of 3-CQA and 4-CQA isomers after the crack reaction in contrast to C. arabica species was described for the first time.

Colon Bioaccessibility under In Vitro Gastrointestinal Digestion of Different Coffee Brews Chemically Profiled through UHPLC-Q-Orbitrap HRMS

Coffee represents one of the most traditionally consumed beverages worldwide, containing a broad range of human health–related compounds. According to previous studies, regular coffee consumption may display protective effects against colorectal cancer and other chronic diseases. The main goal of this research was to evaluate the bioaccessibility of phenolic content and variation in antioxidant capacity of three different types of coffee brews after simulated gastrointestinal digestion. This would allow to elucidate how antioxidant compounds present in coffee may exert their effect on the human body, especially in the colonic stage. Moreover, the content of bioactive compounds namely chlorogenic acids (CGAs, n = 11) and caffeine was also assessed throughout ultra-high-performance liquid chromatography followed by high-resolution Orbitrap mass spectrometry (UHPLC-Q-Orbitrap HRMS). The three main isomers of caffeoylquinic acid constituted the highest fraction of CGAs present in the samples, accounting for 66.0% to 70.9% of total CGAs. The bioaccessibility of coffee polyphenols significantly increased in digested samples from 45.9% to 62.9% at the end of the colonic passage, compared to the non-digested samples. These results point to the colonic stage as the major biological site of action of the active antioxidant coffee compounds.

Reprogramming of Tomato Leaf Metabolome by the Activity of Heat Stress Transcription Factor HsfB1.

Plants respond to high temperatures with global changes of the transcriptome, proteome, and metabolome. Heat stress transcription factors (Hsfs) are the core regulators of transcriptome responses as they control the reprogramming of expression of hundreds of genes. The thermotolerance-related function of Hsfs is mainly based on the regulation of many heat shock proteins (HSPs). Instead, the Hsf-dependent reprogramming of metabolic pathways and their contribution to thermotolerance are not well described. In tomato (Solanum lycopersicum), manipulation of HsfB1, either by suppression or overexpression (OE) leads to enhanced thermotolerance and coincides with distinct profile of metabolic routes based on a metabolome profiling of wild-type (WT) and HsfB1 transgenic plants. Leaves of HsfB1 knock-down plants show an accumulation of metabolites with a positive effect on thermotolerance such as the sugars sucrose and glucose and the polyamine putrescine. OE of HsfB1 leads to the accumulation of products of the phenylpropanoid and flavonoid pathways, including several caffeoyl quinic acid isomers. The latter is due to the enhanced transcription of genes coding key enzymes in both pathways, in some cases in both non-stressed and stressed plants. Our results show that beyond the control of the expression of Hsfs and HSPs, HsfB1 has a wider activity range by regulating important metabolic pathways providing an important link between stress response and physiological tomato development.

Chlorogenic acid abates male reproductive dysfunction in arsenic-exposed mice via attenuation of testicular oxido-inflammatory stress and apoptotic responses

Arsenic, a major environmental pollutant of global concern, is well-known for its reproductive toxicity. In this study, the protective potential of chlorogenic acid (CGA), a caffeoylquinic acid isomer abundantly found in many plants, was investigated against sodium arsenite (NaAsO2)-induced testicular dysfunctions. Adult male Swiss mice were either administered NaAsO2 alone at 5 mg kg−1 or co-treated with CGA at 100 mg kg−1 or 200 mg kg−1 body weight for 4 weeks. Results showed that NaAsO2-treated mice exhibited marked declines in testes weight, sperm count, and viability accompanied by decreases in sexual hormonal levels. Moreover, NaAsO2 toxicity evoked exhaustion of antioxidant markers (SOD, CAT, GPx, GR, and GSH), down-regulation of Nrf2 (nuclear factor erythroid 2–related factor 2) gene expression level, and elevations in malondialdehyde. Further, elevations in inflammatory cytokines (IL-1, TNF-α, and IL-6) together with the up-regulation of pro-apoptotic biomarkers (Bax and caspase- 3) and down-regulation of anti-apoptotic Bcl-2 were observed in NaAsO2 intoxication. Immunohistochemical analysis of testis sections of NaAsO2-treated mice showed high caspase-3 expression. These findings were well supported with testicular histopathological examination. However, pretreatment of mice with CGA resulted in noteworthy improvements in testicular damage induced by arsenic in a dose-dependent manner possibly mediated by the Nrf2 signaling pathway. Conclusively, CGA counteracted arsenic-induced testicular injury through its antioxidant, anti-inflammatory, and anti-apoptotic properties. Therefore, CGA could serve as a favorable intervention in the alleviation of arsenic-induced reproductive toxicity.

1 H-NMR metabolomics profiling of recombinant tobacco plants holding a promoter of a sesquiterpene cyclase.

Nicotiana tabacum is a plant model intensively used in the bio-engineering pharmaceutical industry as a platform to produce drugs and therapeutic agents. Currently, no information regarding the non-targeted metabolome of transgenic tobacco containing recombinant regulatory sequences is available. To compare the proton nuclear magnetic resonance (1 H-NMR) metabolomics profiling of a recombinant Nicotiana tabacum strain containing a promoter of a sesquiterpene cyclase from Capsicum annuum driving GUS expression, versus wild-type samples. Methodology The non-targeted 1 H-NMR metabolome was obtained and processed by principal component analysis (PCA) and orthogonal projection to latent structures discriminant analysis (OPLS-DA). The differential metabolites were quantified by quantitative NMR. PCA and OPLS-DA revealed 37 metabolites including 16 discriminant compounds for transgenic samples. Ethanol (0.4 mg g-1 ), the main differential compound, was exclusively detected in transgenic tobacco; however, high levels of formate (0.28 mg g-1 ) and acetate (0.3 mg g-1 ) were simultaneously observed in the same group of samples. Cembratriene-4,6-diol, an antitumour and neuroprotective compound, and capsidiol, a known phytoalexin, increased by about 30% in transgenic samples. In addition, the endogenous levels of the antioxidant caffeoylquinic acid isomers increased by 50% in comparison to those of wild-type tobaccos. Our results support the occurrence of metabolic differences between wild type and transgenic tobacco containing a promoter of a Capsicum sesquiterpene cyclase gene. Interestingly, the recombinant transgenic strain studied accumulated high amounts of added value compounds with biological activity.

One-Step Isolation of Monoterpene Indole Alkaloids from Psychotria leiocarpa Leaves and Their Antiviral Activity on Dengue Virus Type-2

The leaf MeOH extract of Psychotria leiocarpa (Rubiaceae) showed in vitro non-cytotoxic and anti-dengue virus serotype 2 (DENV2) activity in human hepatocarcinoma cell lineage (HepG2). A one-step and cost-effective reversed-phase solid-phase extraction method based on high-performance liquid chromatography (HPLC) parameters allowed the isolation, directly from this bioactive extract, of the monoterpene indole alkaloids: N-glucopyranosyl vincosamide (1), vincosamide (2) and strictosidinic acid (3). The chemical structures were characterized based on 1D and 2D nuclear magnetic resonance (NMR), UV and high-resolution mass spectra (HRMS). The methodology has also allowed yielding a polyphenolic-rich fraction that was analyzed by high-performance liquid chromatography coupled to diode array detection and electrospray ionization tandem mass spectrometry (HPLC-DAD-ESI-MS/MS) revealing two flavonol triglycosides (4, 5) and three caffeoylquinic acid isomers (6-8). Compound 3 is reported for the first time in P. leiocarpa and all the phenolic compounds (4-8) are described for the first time in the genus Psychotria. Compounds 1-3 showed to be non-cytotoxic and anti-dengue active towards DENV2, highlighting vincosamide (2).

Chlorogenic acid prevents hepatotoxicity in arsenic-treated mice: role of oxidative stress and apoptosis

Arsenic is a potent and toxic heavy metal found in the environment that causes health problems, including liver disease, in humans and animals. Chlorogenic acid (CA) is the most abundant caffeoylquinic acid isomer present in plants. This study aims to assess how CA protects the liver tissue following sodium arsenite (NaAsO2)-induced toxicity in mice. Male Swiss mice were allocated into 5 groups: Control, intragastrically administered CA (200 mg/kg), intragastrically administered NaAsO2 (5 mg/kg), and two groups administered with CA (100 and 200 mg/kg) and NaAsO2. CA was administered 30 min before NaAsO2 and all the mice were treated daily for 28 days. To investigate the biochemical, histopathological, immunohistochemical, and molecular changes, blood and liver samples were collected. NaAsO2 treatment increased the liver function biomarkers such as alanine transaminase, aspartate transaminase, alkaline phosphatase, and total bilirubin. Lipid and nitric oxide production was elevated. Glutathione content and the activities of superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase decreased, indicating a disturbance in redox homeostasis. Histopathological examination revealed a granular degeneration of hepatocytes, infiltration of inflammatory cells, and centrilobular hepatocyte necrosis. Furthermore, tumor necrosis factor-α and interleukin-1β were upregulated upon NaAsO2 treatment, suggesting the induction of inflammation. Moreover, NaAsO2 triggered apoptosis in the liver by upregulating Bax and caspase-3 and downregulating Bcl-2. However, CA abrogated the biochemical, molecular, and histological changes, reflecting its hepatoprotective role in response to NaAsO2 treatment. Our findings demonstrate that CA could be a potential therapeutic to minimize NaAsO2-induced hepatic injury.

Evaluation of Antifungal Phenolics from Helianthus tuberosus L. Leaves against Phytophthora capsici Leonian by Chemometric Analysis.

Phytophthora capsici Leonian causes destructive economical losses in pepper production, and a promising source of natural fungicides- Helianthus tuberosus leaves was reported. The antifungal activities of different extracts and compounds from H. tuberosus leaves against the phytopathogen, P. capsici Leonian, were examined by chemometric analysis, including HPLC-MS/MS and multivariate data analyses. Principal component analysis and orthogonal partial least squares-discriminate analysis were applied to examine the four groups of H. tuberosus leaves samples, including crude extracts obtained by different methods, including refluxing, macerating, and refluxing under vacuum; four fractions, namely, petroleum ether (PE), chloroform (Chl), ethyl acetate (EA), and n-butanol (NB) fractions; the samples of three H. tuberosus cultivars; and the samples at three growth stages of cultivar Nan Yu. The phenolics contents were categorized based on 3,5-Dicaffeoylquinic acid (3,5-DiCQA), 1,5-Dicaffeoylquinic acid (1,5-DiCQA), 3-O-Caffeoylquinic acid (3-CQA), and 4,5-Dicaffeoylquinic acid (4,5-DiCQA), which were predominant in all the samples. Antifungal activity assay revealed that Chl and NB fractions were more active against P. capsici Leonian with lower IC50(half of maximal inhibitory concentration) values, whereas partial least squares-discriminate analysis suggested caffeoylquinic acid isomer(4-CQA), methyl-quercetin glycoside(MQG), and caffeic acid(CA) might be the main active components in H. tuberosus leaves against P. capsici Leonian. Furthermore, microscopic evaluation demonstrated structural deformities in P. capsici Leonian treated with Chl and NB fractions, indicating the antifungal effects of H. tuberosus leaves. These results imply that H. tuberosus leaves with a high concentration of phenolics might be a promising source of natural fungicides.

Assessment of the potential of Artemisia annua L. as a source of biologically active substances in Belarus

Aim. The aim of the work was to assess the composition of biologically active compounds in plants of Artemisia annua L. Methods. The plants were grown on the experimental plot of the Central Botanical Garden of the National Academy of Sciences of Belarus. Essential oil from the above-ground mass was isolated by the method of water-steam distillation. To analyze the content of flavonoids and hydroxycinnamic acids, we used the method of quantitative extraction and spectrophotometric determination of the total content of hydroxycinnamic acids in the presence of flavonoids in plant extractive substances, and the method of Folin-Chocalteu was used to determine the content of phenolic compounds. Analysis of the essential oils was performed by GC and NMR. The analysis of the content of phenolic acids in extracts was performed by HPLC. Results. The genetic potential of A. annua was evaluated under conditions of the central agroclimatic zone of Belarus. The yield and composition of essential oils, the content of phenolic compounds and flavonoids, the composition of phenolic acids were studied. Conclusions. A. annua in the conditions of the central agroclimatic zone of Belarus contains essential oil, which is dominated by isoartemisia ketone, β-selinene, β-myrcene and camphor. In the above-ground mass phenolic acids including chlorogenic acid and isomers of caffeoylquinic acid were predominant in the composition of phenolic compounds.&#x0D; Keywords: annual wormwood, essential oil, phenolic compounds, phenolic acids.

Identification, Quantification, and Antioxidant Activity of Hydroalcoholic Extract of Artemisia campestris from Algeria.

Our study aimed to investigate the chemical profile of hydroalcoholic extract of Algerian Artemisia campestris and its antioxidant activity. The hydroalcoholic extract of Algerian A. campestris was investigated for its phenolic constituents using high performance liquid chromatography (HPLC)-diode array detection (DAD)-electrospray ionization (ESI)-mass spectrometer (MS)/MS. The in vitro antioxidant activity and total phenolic content were also evaluated via oxygen radical absorbance capacity and Folin-Ciocalteu assays, respectively. HPLC-DAD-ESI-MS/MS analysis revealed that the main tentatively identified compounds were caffeoylquinic acid isomers, flavonoids, and benzoic acid derivatives. Additionally, the hydroalcoholic extract exhibited a promising antioxidant activity value of 120.5±10.4 μmol Trolox equivalent antioxidant capacity/g dry weight (DW), and a strong correlation exists between this activity and the total phenolic content value of 102.09±1.65 mg/g gallic acid equivalents DW. The hydroalcoholic extract of A. campestris is a promising candidate for the production of naturally occurring antioxidant agents.

Preparative isolation of caffeoylquinic acid isomers from Kuding tea by salt‐containing aqueous two‐phase extraction and purification by high‐speed countercurrent chromatography

AbstractCaffeoylquinic acid and its derivatives are important groups of natural plant phenols with antioxidant and neuroprotective properties. In this study, a salt‐containing aqueous system consisting of ethanol/ammonium sulfate/water 32:21:47 wt% was used to enrich these polar phenols from crude Kuding tea extract. In a one‐step extraction, the yields of 1‐O‐caffeoylquinic acid and 3‐O‐caffeoylquinic acid can be up to three times that obtained by using a 1‐butanol/water system. To purify polar caffeoylquinic acid, a two‐step high‐speed countercurrent chromatography method with a conventional organic/water system and a salt‐containing aqueous system was introduced. In the first step, dicaffeoylquinic acids and their derivatives were separated from the crude extract using n‐hexane/ethyl acetate/methanol/water (0.05% formic acid) 1:5:2.5:5 v/v. And caffeoylquinic acids and polar 1,3‐O‐dicaffeoylquinic acid were collected as a mixed fraction. In the second step, the mixture was futher purified using a 1‐butanol/ethanol/saturated ammonium sulfate/water (9:1:6:4, v/v) solvent system. 1‐O‐Caffeoylquinic acid, 3‐O‐caffeoylquinic acid, chlorogenic acid, and 1,3‐O‐dicaffeoylquinic acid were well separated with high purities of 90%. The chemical structures of the isolated compounds were identified by liquid chromatography with diode array detection and electrospray ionization mass spectrometry using a standard reference.

Fatty acids-based microemulsion liquid chromatographic determination of multiple caffeoylquinic acid isomers and caffeic acid in honeysuckle sample

A green and efficient microemulsion liquid chromatographic (MELC) method using fatty acid as co-surfactant and electrochemical detection was established and validated for the determination of four caffeoylquinic acid isomers and caffeic acid in honeysuckle samples. The influences of each individual component within the isocratic oil-in-water (O/W) microemulsion mobile phase were systematically investigated, such as the type and concentration of co-surfactant, concentration of sodium dodecyl sulphate (SDS), organic modifier addition, type and concentration of oil phase, pH and detection voltage. Results indicated that excellent resolution was achieved using 3.0% w/v of propionic acid, 0.5% w/v of ethyl acetate, 1.0% w/v of SDS, 5% w/v acetonitrile, 90.5% v/v of water and 25 mM sodium dihydrogen phosphate at pH = 3 as microemulsion mobile phase and 0.8 V as the optimal voltage value. Under the optimal condition, analytical performance of developed method was evaluated. The detection limits were below 17.3 ng/mL and intra-day and inter-day precisions by relative standard deviations (RSD%) were between 0.5% and 3.6%. Satisfactory recovery (in the range of 83.8–109.1%) with good repeatability lower than 4.7% (n = 3) was obtained. Therefore, the developed O/W MELC method was rapid, precise and accurate for simultaneous determination of neochlorogenic acid, chlorogenic acid, isochlorogenic acid A and isochlorogenic acid C in honeysuckle samples, with contents of 2.6, 28.7, 18.1 and 5.2 mg/g, respectively.

Chlorogenic acid isomers directly interact with Keap 1-Nrf2 signaling in Caco-2 cells

Chlorogenic acid (CGA) exists as multiple isomers (e.g., 3-CQA, 4-CQA, 5-CQA, 3,4-diCQA, 3,5-diCQA, and 4,5-diCQA) in foods such as coffee beverages, fruits and vegetables. This study aimed to investigate relative activities of these six different CGA isomers to modify redox biology in inflamed Caco-2 cells that involved Nrf2 signaling. Caco-2 cells were pre-treated with individual CGA isomers to assess the relative effectiveness to mitigate oxidative stress. Isomer-specific capacity of different CGA isomers for direct free radical scavenging activity and potential endogenous control of oxidative stress were determined using chemical assays and cell-based experiments, respectively. Molecular dynamics simulations of the CGA and Keap1-Nrf2 complex were performed to predict CGA structure-specific interactions. Results demonstrated that dicaffeoylquinic acid (diCQA including 3,4-diCQA, 3,5-diCQA, and 4,5-diCQA) isomers had greater (p < 0.05) affinity to ameliorate oxidative stress through direct free radical scavenging activity. This observation corresponded to greater (p < 0.05) capacity to activate Nrf2 signaling compared to caffeoylquinic acid (CQA including 3-CQA, 4-CQA, and 5-CQA) isomers in inflamed differentiated Caco-2 cells. Simulations revealed that differences between the ability of CQA and diCQA to interact with the Keap1-Nrf2 complex may be due to differences in relative orientation within this complex. The observed CGA isomer-specific affinity for CQA to activate Nrf2 signaling was confirmed by nuclear translocation of Nrf2 induced by CGA and greater (p < 0.05) upregulation of genes related to Nrf2 expression.

Preparative separation of seven phenolic acids from Xanthii Fructus using pH-zone-refining counter-current chromatography combined with semi-preparative high performance liquid chromatography.

Phenolic acids represented by caffeoylquinic acids in Xanthii Fructus have various pharmacological activities such as anti-inflammatory, anti-nociceptive, anti-oxidative and anti-allergic effects. In this study, pH-zone-refining counter-current chromatography was successfully applied in the segmentation of crude samples and further separation of phenolic acids from Xanthii Fructus. We initially segmented 1.6 g of the crude sample to yield three sample fractions using a two-phase solvent system composed of EtOAc–ACN–H2O (4 : 1 : 5, v/v/v) with 10 mM TFA added to the organic phase as the stationary phase and 10 mM NH3·H2O added to the aqueous phase as the mobile phase. The first fraction was separated using EtOAc–H2O (1 : 1, v/v) (10 mM TFA was added in the upper phase and 20 mM NH3·H2O was added in the lower phase) solvent system, the second fraction containing low-content compounds was separated using semi-preparative high performance liquid chromatography, and the third fraction contained one pure compound. As a result, seven phenolic acids including six caffeoylquinic acid isomers (3-caffeoylquinic acid, 4-caffeoylquinic acid, 5-caffeoylquinic acid, 1,5-O-dicaffeoylquinic acid, 3,5-O-dicaffeoylquinic acid, and 4,5-O-dicaffeoylquinic acid) and caffeic acid were successfully isolated from Xanthii Fructus with purities above 90%. This study demonstrated that pH-ZRCCC is an efficient preparative separation method for phenolic acids, especially isomeric caffeoylquinic acids, from natural products.

Acidity and Antioxidant Activity of Cold Brew Coffee

The acidity and antioxidant activity of cold brew coffee were investigated using light roast coffees from Brazil, two regions of Ethiopia, Columbia, Myanmar, and Mexico. The concentrations of three caffeoylquinic acid (CQA) isomers were also determined. Cold brew coffee chemistry was compared to that of hot brew coffee prepared with the same grind-to-coffee ratio. The pH values of the cold and hot brew samples were found to be comparable, ranging from 4.85 to 5.13. The hot brew coffees were found to have higher concentrations of total titratable acids, as well as higher antioxidant activity, than that of their cold brew counterparts. It was also noted that both the concentration of total titratable acids and antioxidant activity correlated poorly with total CQA concentration in hot brew coffee. This work suggests that the hot brew method tends to extract more non-deprotonated acids than the cold brew method. These acids may be responsible for the higher antioxidant activities observed in the hot brew coffee samples.

Preparative separation of caffeoylquinic acid isomers from Lonicerae japonicae Flos by pH-zone-refining counter-current chromatography and a strategy for selection of solvent systems with high sample loading capacities

Caffeoylquinic acid derivatives exhibit anti-inflammatory, antioxidant, and antibacterial activities. We successfully applied pH-zone-refining counter-current chromatography (pH-ZRCCC) to separation of isomeric caffeoylquinic acids from Lonicerae japonicae Flos using a two-phase solvent system composed of ethyl acetate–n-butanol–acetonitrile–water (3:1:1:5, v/v/v/v). Trifluoroacetic acid (10 mM) was added to the upper phase as a retainer and ammonium hydroxide (10 mM) was added to the lower phase as an eluter. As a result, 167.8 mg of chlorogenic acid, 15.9 mg of isochlorogenic acid B, 103.4 mg of isochlorogenic acid C, and 156.0 mg of isochlorogenic acid A were obtained from 1.2 g of crude extract, and all compound purities were over 96%. In addition, by comparing the numeric values of partition coefficient of compounds, it was found that larger differences between the K values of adjacent compounds in the solvent systems resulted in higher sample loading capacities. pH-ZRCCC method is an efficient preparative separation of isomeric caffeoylquinic acids from natural products.