HMW Fraction Research Articles

Investigation of the Distribution and Binding Affinity of Copper to Size-Fractioned Dissolved Organic Matter (DOM) in a Constructed Wetland

This study investigated the distribution and binding affinity of dissolved copper (Cu) and organic carbon (OC) in size-fractioned dissolved organic matter (DOM) in a constructed wetland (CW). Two sites were studied: one at the inflow (P-1) and one within the wetland (P-2). The DOMs (<0.45 μm) were separated into six size fractions using a cross-flow ultrafiltration system. In the wetland (P-2), the concentrations of dissolved organic carbon (DOC) increased while the concentrations of Cu decreased. The high molecular weight fraction (1 kDa–0.45 μm, HMW) contained most of the OC mass (57.4–71.2% averages). On the other hand, Cu was almost equally distributed in HMW and low molecular weight fractions (<1 kDa, LMW) with mean HMW percentages of 50.3–51.3%. The mean Cu binding affinity to DOM ratios (CuBADOM) was 74.9 ± 24.0 μmol/g-C at site P-1 and 17.3 ± 2.6 μmol/g-C at site P-2. The CuBADOM ratios were decreased in wetlands of bulk and size-fractioned DOM (p < 0.001 to p = 0.073). The SUVA254 values for bulk DOM solution were 2.54 ± 0.15 and 1.68 ± 0.18 L/mg-C/m, and humidification index (HIX) values were 1.74 ± 0.16 and 2.09 ± 0.19 for sites P-1 and P-2, respectively. Optical indicators suggested that the wetland process decreased aromaticity but increased the humification degree of DOM. Furthermore, the CuBADOM ratios positively correlated with SUVA254 and HIX within the constructed wetland DOM but not in the influent DOM. Understanding the Cu distribution and binding affinity to size-fractioned DOM makes it possible to develop strategies to mitigate the potential effects of copper pollution in wetlands.

Investigating the molecular weight distribution of atmospheric water-soluble brown carbon using high-performance size exclusion chromatography coupled with diode array and fluorescence detectors

Atmospheric brown carbon (BrC) contain amounts of organic species, but their molecular weight (MW) distributions is still poorly understood. This study applied high-performance size exclusion chromatography (HPSEC) coupled with a diode array detector (DAD) and fluorescence detector (FLD) to characterize the MW distributions of typical chromophores and fluorophores within water-soluble BrC. The investigation focused on the spring season, encompassing both typical urban and rural aerosols. Our results showed that chromophores (at 254 and 365 nm), and humic-like and protein-like fluorophores identified by excitation-emission matrix parallel factor analysis (EEM-PARAFAC) within BrC were broadly distributed along the MW continuum (∼50–20,000 Da). This suggests that BrC mainly comprises complex chromophores and fluorophores with heterogeneous molecular sizes. High-MW (HMW, >1 kDa) species (66%–74%) dominated the chromophores at 254 and 365 nm. However, the latter chromophores were enriched with more HMW species. This result suggested that the HMW chromophores might contribute more to BrC absorption at longer wavelengths. The PARAFAC-derived fluorescent components also exhibited different MW distributions. Three humic-like substances (HULIS) were all dominated by HMW fractions (51%–74%), but protein-like fluorescent component (PLOM) enriched low-MW (LMW, <1 kDa) species (60%–66%). Furthermore, the molecular size (i.e., weight-averaged and number-averaged MW) and the ratios between HMW and LMW species decreased in the order highly-oxygenated HULIS > less-oxygenated HULIS > PLOM, indicating that the fluorophores with longer Em were generally related to larger MW. To our knowledge, this is the first report on the molecular size of individual fluorescent components within aerosol BrC. The results obtained here enhanced our knowledge of heterogeneous composition, complex physicochemical properties, and potential atmospheric fates of aerosol BrC.

Evaluating the phenolic composition and antioxidant properties of Georgia pecans after in vitro digestion

Acetonic crude extracts from raw and roasted Georgia pecans were prepared and subjected to in vitro gastrointestinal digestion. Digested and undigested extracts of raw pecan phenolics were then separated into low- and high-molecular-weight (LMW and HMW) fractions via Sephadex LH-20 column chromatography; afterwards, they were characterized by RP-HPLC-ESI-MS. The LMW fraction consisted primarily of flavan-3-ols and ellagic acid derivatives and following digestion there was an overall loss in phenolics from 16% to 100%. In the HMW fraction, procyanidins with degrees of polymerization ranging from dimers to hexamers were present. Following digestion, a loss in higher oligomeric procyanidins was observed, along with a significant increase in quantity of dimers. This increase was attributed to the dimerization of (+)-catechin/(−)-epicatechin and the scission of larger procyanidins, mainly tetramers to hexamers. The loss of phenolics following digestion, as seen by HPLC characterization, was reflected in reduced total phenolics content and antioxidant capacity, as determined by selected in vitro antioxidant assays.

In vivo anti-tumor activity of Lignosus rhinocerus TM02® using a MCF7-xenograft NCr nude mice model

Ethnopharmacological relevanceLignosus rhinocerus (Cooke) Ryvarden (also known as Tiger Milk mushroom, TMM), is a basidiomycete belonging to the Polyporaceae family. It has been documented to be used by traditional Chinese physicians and indigenous people in Southeast Asia to treat a variety of illnesses, such as gastritis, arthritis, and respiratory conditions, as well as to restore patients' physical well-being. TMM has also been used in folk medicine to treat cancer. For example, people from the indigenous Kensiu tribe of northeast Kedah (Malaysia) apply shredded TMM sclerotium mixed with water directly onto breast skin to treat breast cancer, while Chinese practitioners from Hong Kong, China prescribe TMM sclerotium as a treatment for liver cancer. L. rhinocerus has previously been demonstrated to possess selective anti-proliferative properties in vitro, however pre-clinical in vivo research has not yet been conducted. Aim of studyThis study aimed to examine the anti-tumor activities of L. rhinocerus TM02®, using two different sample preparations [cold water extract (CWE) and fraction] via various routes of administration (oral and intraperitoneal) on an MCF7-xenograft nude mouse model. This study also investigated the inhibitory effect of TM02® CWE and its fractions against COX-2 in vitro using LPS-induced RAW264.7 macrophages, on the basis of the relationship between COX-2 and metastasis, apoptosis resistance, as well as the proliferation of cancer cells. Materials and methodsThe first preparation, L. rhinocerus TM02® sclerotium powder (TSP) was dissolved in cold water to obtain the cold water extract (CWE). It was further fractionated based on its molecular weight to obtain the high (HMW), medium (MMW) and low (LMW) molecular weight fractions. The second preparation, known as the TM02® rhinoprolycan fraction (TRF), was obtained by combining the HMW and MMW fractions. TSP was given orally to mimic the daily consumption of a supplement; TRF was administered intraperitoneally to mimic typical tumorous cancer treatment with a rapid and more thorough absorption through the peritoneal cavity. Another experiment was conducted to examine changes in COX-2 activity in LPS-induced RAW264.7 macrophages after a 1-h pre-treatment with CWE, HMW, and MMW. ResultsOur results revealed that intraperitoneal TRF-injection (90 μg/g BW) for 20 days reduced initial tumor volume by ∼64.3% (n = 5). The percentage of apoptotic cells was marginally higher in TRF-treated mice vs. control, suggesting that induction of apoptosis as one of the factors that led to tumor shrinkage. TSP (500 μg/g BW) oral treatment (n = 5) for 63 days (inclusive of pre-treatment prior to tumor inoculation) effectively inhibited tumor growth. Four of the five tumors totally regressed, demonstrating the effectiveness of TSP ingestion in suppressing tumor growth. Although no significant changes were found in mouse serum cytokines (TNF-α, IL-5, IL-6 and CCL2), some increasing and decreasing trends were observed. This may suggest the immunomodulatory potential of these treatments that can directly or indirectly affect tumor growth. Pre-treatment with CWE, HMW and MMW significantly reduced COX-2 activity in RAW264.7 macrophages upon 24 h LPS-stimulation, suggesting the potential of L. rhinocerus TM02® extract and fractions in regulating M1/M2 polarization. ConclusionBased on the findings of our investigation, both the rhinoprolycan fraction and crude sclerotial powder from L. rhinocerus TM02® demonstrated tumor suppressive effects, indicating that they contain substances with strong anticancer potential. The antitumor effects of L. rhinocerus TM02® in our study highlights the potential for further explorations into its mechanism of action and future development as a prophylactic or adjunct therapeutic against tumorous cancer.

Understanding the influence of pre-ozonation on the formation of disinfection byproducts and cytotoxicity during post-chlorination of natural organic matter: UV absorbance and electron-donating-moiety of molecular weight fractions.

Pre-ozonation can reduce the formation of disinfection byproducts (DBPs) and related adverse effects during subsequent chlorination, but the change of each molecular weight (MW) fraction during each step of combined pre-ozonation and post-chlorination has not been well illustrated. In this study, it was investigated in terms of electron-donating-moieties (EDMs) and UVA254 for a representative natural organic matter from Suwanee river (SRNOM). Pre-ozonation suppressed the post-chlorination of SRNOM through oxidation of almost all EDMs (>85%) and UVA254 (>90%) in high MW fractions (HMW, >3.2kDa) and moderate EDMs (43%) and UVA254 (72%) in medium MW fractions (MMW, 1.0-3.2kDa). Furthermore, pre-ozonation led to comparable abatements of EDMs and UVA254 for HMW fractions, but lower abatement of EDMs than UVA254 for MMW fractions. However, when t-BuOH was used as an •OH-quencher, pre-ozonation led to a few instances in which there were higher abatements of EDMs than UVA254 for the MMW fraction. These findings suggested that the HMW fraction dominantly underwent ring-cleavage of phenols via O3- or •OH-oxidation. Differently, the MMW fraction underwent ring-cleavage of phenols and quinones-formation via O3-oxidation, but occasionally underwent hydroxylation and hydro-phenol formation via •OH-oxidation. Because of forehand elimination of reactive moieties (e.g. EDMs), pre-ozonation obviously inhibited the formation of representative DBPs (67%-84% inhibition), total organic chloride (51% inhibition) and cytotoxicity (31% inhibition), but may have promoted the formation of carbonyl-DBPs (trichloroacetone and chloral hydrate). When compared with UVA254, EDMs would better for surrogate of DBPs formation. EDM abatement surrogated the formation of total organic chlorine (TOCl) and cytotoxicity following a two-stage phase, possibly because the speciation of DBPs and transformation products varied with the abatement of EDMs.

Baseline distributions and sources of polycyclic aromatic hydrocarbons (PAHs) in reef-associated sediments of Vembar group of Islands, Gulf of Mannar, India

The study aims to investigate the source and concentration of PAH fractions in the reef sediments of the Vembar group of Islands, Gulf of Mannar, India. The concentration of PAHs ranged from 0.36 to 15.98 ng/g. The reef environment was less contaminated with low-molecular-weight PAH fractions. The accumulation of the LMW-PAH fraction was very less, whereas the HMW fraction was derived from pyrolytic sources. The level of low and high molecular weight PAHs was lower than the level of Effective Range Median (ERM) and Effective Range Low (ERL).The calculated total TEQ value ranged from ND to 10.24 ng/g in the reef sediments.

The release inhibition of organic substances from microplastics in the presence of algal derived organic matters: Influence of the molecular weight-dependent inhibition heterogeneities

As the emerging contaminants, the behavior and fate of microplastics (MPs) were highly related to the interactions with surrounding organic matters. However, information on the effects of molecular sizes of organic matters on the interaction is still lacking. In this study, the bulk algal-derived organic matter (AOM) samples were obtained and further fractionated into high molecular weight (HMW-, 1kDa-0.45 μm) and low molecular weight (LMW-, < 1 kDa) fractions. The interaction between MPs [polyethylene (PE) and polystyrene (PS)] and these MW-fractionated AOMs were characterized by dissolved organic carbon, fluorescence and absorbance spectroscopy, and fourier transform infrared (FTIR) analysis. Results showed that presence of AOM could effectively inhibit the release of additives from MPs. Further analysis found that the inhibition extents decreased in the order of HMW- > bulk > LMW-AOM. The absorbance and fluorescence spectroscopy showed that aromatic protein-like substances in HMW fraction exhibited higher adsorption affinity to MPs than the bulk and LMW counterparts. The strong sorption of aromatic substances may offer more binding sites for additives to inhibit the release of organic substances. Moreover, two dimensional FTIR correlation spectroscopy revealed that the HMW non-aromatic substances were preferentially adsorbed onto PS, which led to an enhanced adsorption capacity to additives by forming H-bonding. Therefore, the MW- and component-dependent heterogeneities of AOM samples must be fully considered in evaluating the environmental behavior of MPs.

Molecular weight-dependent heterogeneities in photochemical formation of hydroxyl radical from dissolved organic matters with different sources

Dissolved organic matter (DOM) is ubiquitous in aquatic ecosystem and characterized by a wide range of molecular weight (MW) distribution. In this study, a total of nine bulk DOM samples, including five International Humic Substances Society (IHSS) standards and four naturally collected samples, were fractionated into low MW (LMW-, <1 kDa) and high MW (HMW-, 1 kDa~0.45 μm) fractions, with MW-dependent heterogeneities in photochemical formation of hydroxyl radical (HO) was investigated. The formation rate of HO (RHO) for the bulk samples were 4.60–7.27 × 10−12 M/s/mg-C/L for IHSS standards and 4.63–7.66 × 10−12 M/s/mg-C/L for naturally collected samples. Regardless of sample types, the LMW fraction was found to exhibit generally higher RHO values than the HMW counterparts. For IHSS standards, the RHO decreased from 4.68–8.46 × 10−12 M/s/mg-C/L for LMW fraction to 3.67–6.66 × 10−12 M/s/mg-C/L for HMW fraction, and for naturally collected samples, the value of RHO decreased from 5.21–12.04 × 10−12 M/s/mg-C/L for LMW fraction to 3.25–6.49 × 10−12 M/s/mg-C/L for HMW counterpart. A positive correlation between the net RHO and the normalized intensities of fluorescent peak [Em/Ex: (400–500)/(230–250) nm] was found, showing that the HO formation was strongly coupled to the abundance of humic-like substances. The results indicate that aquatic DOM is an important pool for HO formation, and characterization of MW distribution rather than average MW is thus required to explain the DOM-induced formation potential of HO.

Adsorption of cyanobacterial extracellular polymeric substance on colloidal particle: Influence of molecular weight.

Cyanobacterial extracellular polymeric substances (EPSs) in aquatic environments are easily adsorbed onto colloidal particles, whereas the adsorption behavior as affected by molecular weight (MW) properties remained unknown till now. Herein, the bulk cyanobacterial EPS matrix (<0.45μm) was fractionated into high MW (HMW-, 1kDa~0.45μm) and low MW (LMW-, <1kDa) fractions, with MW-dependent adsorption heterogeneities onto TiO2 colloids exploring through batch experiment, UV-Vis and fluorescence spectroscopy, and two dimensional Fourier transform infrared correlation spectroscopy (2D-FTIR-COS). About two-thirds of total organic matters within bulk EPS matrix were distributed in the HMW fraction, leaving one-third in the LMW fraction. Compared to LMW-EPS, the HMW counterpart exhibited higher aromaticity and richness of autochthonous protein-like substances, showing evident MW-dependent differences in abundance and composition. The adsorption capacity based on the measurement of total abundance, UV-Vis and fluorescent spectra all decreased in sequence of HMW->Bulk>LMW-EPS, demonstrating obvious MW-dependent adsorption heterogeneities. During adsorption, the values of SUVA254 in residual supernatants exhibited an initial decrease followed by gradual increase for all samples, suggesting that the preferentially adsorbed aromatic substances can be subsequently replaced by the non-aromatic moieties. 2D-FTIR-COS further revealed that the carboxylic groups of proteins were preferentially adsorbed onto colloidal surface, followed by the CC functional groups and then the CH groups of polysaccharides, which accounted for the variations of SUVA254 values in the supernatants. This study demonstrated that the adsorption behavior of EPS matrix was highly MW-dependent, and detailed characterization on size fractionation is thus needed in future studies.

Degradation of Peptide-Bound Maillard Reaction Products in Gastrointestinal Digests of Glyoxal-Glycated Casein by Human Colonic Microbiota.

A large portion of Maillard reaction products (MRPs) cannot be absorbed in the upper gut and therefore may be further decomposed and utilized by colonic microbiota (CM). This work reported the stability of UV-absorbent MRPs, fluorescent MRPs and peptide-bound N(ε)-(carboxymethyl)-lysine (CML) in high molecular weight (HMW, >10 kDa), medium molecular weight (MMW, 1-10 kDa), and low molecular weight (LMW, <1 kDa) gastrointestinal digests of glyoxal-glycated casein in the presence of CM. Fluorescent MRPs showed high stability, whereas UV-absorbent MRPs may be partially decomposed. A higher depletion rate of CML was found in the LMW fraction (38.7%) than in the MMW (21.7%) and HMW (9.6%) fractions. The 16S rRNA sequencing results revealed both beneficial and detrimental changes in CM composition induced by the glycated fractions. Generation of short-chain and branched-chain fatty acids in fermentation solutions with glycated fractions was significantly suppressed compared with that in fermentation solution with unglycated digests. This work revealed the possible interplay between peptide-bound MRPs and CM.

Amino acid enantiomers in old and young dissolved organic matter: Implications for a microbial nitrogen pump

Abstract Dissolved organic nitrogen (DON) represents the largest reservoir of fixed N in the surface ocean and a significant portion accumulates in the deep sea, where it can persist for millennial time scales. However, like the dissolved organic carbon (DOC) pool, the origin and composition of long-lived, refractory DON remains largely unknown. In recent years, the “microbial carbon pump” hypothesis has emerged from abundant evidence showing that microbial processes are primarily responsible for refractory DOC accumulation. However, a similar mechanism for DON has rarely been investigated. In the study of DON, spectroscopic evidence has indicated a primarily amide composition, implying a dominant contribution from peptides. Therefore, if an analogous “microbial nitrogen pump” controls refractory DON accumulation, the amino acid component should bear increasing signatures of microbial origin with increasing age. Here we investigate the microbial sequestration of N via the production of refractory DON, for the first time considering together DOM Δ14C with amino acid (AA) molar abundance (Mol%) and D/L ratio (as a tracer for prokaryotic input). Measurements were made on a unique set of high and low molecular weight (HMW, LMW) DOM isolates with 14C ages and chemical compositions generally consistent with semi-labile and refractory DOM respectively. The samples were collected in the North Pacific Subtropical Gyre where deep waters contain some of the oldest DOC in the world ocean. We observe higher D/L ratios in older, LMW DOM isolates for almost all analyzed AAs. Using mass spectral data, we also quantify three D-AAs in all samples (D-valine, D-phenylalanine, and D-leucine), which have not previously been confirmed in ocean DOM. These newly identified D-AAs are concentrated in the LMW refractory DOM fraction and have oceanographically consistent depth profiles. Our results suggest that several novel D-AA subgroupings may be unique tracers for different prokaryotic source processes. D-alanine appears to have largely independent cycling from the other D-AAs with a connection to the production of HMW DON, which we hypothesize is linked to water column peptidoglycan. In contrast, D-leucine, D-valine, and D-phenylalanine appear to be most strongly related to the production of LMW DON. Trends in both the HMW and LMW fractions suggest a linkage to sinking particles and local microbial transformations, implying that LMW DON has a direct biological source rather than originating from successive microbial reprocessing of HMW DON. Taken together, our observations are consistent with the dominant production of refractory LMW DON by prokaryotic organisms and suggests that different AA sub-groupings that can be used to track different processes within the DON pool.

Performance Characteristics of Asphalt Binders containing Sodium-Alginate Hollow Fibers and Recycled Materials

Self-healing products such as hollow-fibers filled with an asphalt rejuvenator present an emerging technology that would enhance an asphalt mixture’s resistance to cracking damage. The objective of this study was to evaluate the rejuvenating efficiency of sodium-alginate fibers containing a rejuvenator product using asphalt binder blends containing extracted binder from recycled materials. The effects of adding extracted binder from recycled materials and sodium-alginate fibers on asphalt binder blends were evaluated by conducting a series of chemical and binder tests. HP-GPC and FTIR test results showed that the addition of fibers in blends containing recycled materials resulted in an increase in the HMW/LMW ratios. Some of the added polymeric fibers are thought to have increased the HMW fraction, thus leading to increase in the HMW/LMW ratio. The increase of the HMW fraction suggests that some of the fibers, which are polymers, caused the increase in the HMW/LMW ratios. MSCR test results showed that a binder blend with extracted binder from recycled materials and sodium-alginate fibers would have less rutting susceptibility than a conventional virgin binder would.

Partitioning of Cd, Zn and Fe in the Tissues and Cytosols of Blood Cockles (Anadara granosa) from the Gulf of Thailand

Anadara granosa is a bivalve blood cockle occurring abundantly in tropical estuarine intertidal sediments. The accumulation of Cd, Zn and Fe and their partitioning at the sub-cellular level in the cytosol has been investigated in the gills, digestive gland, body and foot of A. granosa from seven estuaries in the Gulf of Thailand. Cadmium and Zn were bioaccumulated predominantly in the gills with smaller concentrations distributed equally throughout the other tissues. Iron was present at relatively low concentrations in gills but was higher, and more evenly distributed, in the digestive gland, body and foot. The concentrations of metallothionein-like protein (MTLP) were found to be in the range 1–11 mg g−1 (dry weight) with the highest values being in the digestive gland at all sites. Partitioning of the metals in the cytosols between low molecular weight (LMW) compounds, MTLP and high molecular weight compounds (HMW) indicated that 16–39% of total Cd, 6.5–15% of total Fe and < 6% of total Zn were bound with MTLP. However, 23–27% of total Zn was bound with the HMW fraction, ascribed potentially to its association with the blood pigment haemoglobin (Hb). The metal-handling strategy, involving Zn, is likely important to blood cockles in supporting their uptake of dissolved oxygen in under-saturated tropical waters.

Characterizing the phenolic constituents and antioxidant capacity of Georgia peaches

Acetonic crude phenolic extracts of six Georgia peach cultivars were prepared and separated into low- and high-molecular-weight (LMW and HMW) fractions by Sephadex LH-20 column chromatography. Further characterization via RP-HPLC-ESI-MS identified the main phenolics as hydroxycinnamates, (+)-catechin, and proanthocyanidins with degrees of polymerization up to seven. The LMW phenolics of the commercial cultivar, ‘July Prince’, were further chromatographed and examined by RP-HPLC-ESI-MS. Derivatives of phenolic acids and flavan-3-ols, along with eriodictyol and quercetin diglycosides, were identified. Antioxidant capacities of the LMW and HMW fractions were determined using in vitro assays. H-ORACFL and FRAP assays gave values of 872 to 2428 μmol Trolox eq./100 g f.w. and 309 to 432 μmol Fe2+ eq./100 g f.w., respectively. The total phenolics content (TPC) was also measured; correlations between TPCs and antioxidant assays indicated that the HMW fractions of peach extracts were major contributors to the antioxidant capacity of the cultivars analyzed.

Ceramic microfiltration – influence of pretreatment on operational performance

A sustainable membrane operation often requires pretreatment of the feed liquor to improve its technical and economic feasibility. This paper reports the impact of pretreatment on the performance of ceramic microfiltration for several pilot studies at different locations. Four different pretreatment processes were investigated: (1) in-line coagulation (to remove high molecular weight, HMW, dissolved organic carbon, DOC); (2) ion exchange (to remove low molecular weight, LMW, DOC); (3) ozone (for disinfection, taste and odor control, and modifying the character of DOC) (4) ion exchange followed by in-line coagulation (for almost complete removal of DOC). Pretreatment in all cases was needed to control membrane fouling, to establish a technically and economically feasible process. These studies seem to show that the DOC's HMW (which includes biopolymers) and LMW fractions (the latter includes humics/acids), are primarily responsible for the TMP increase after filtration followed by backwashing (irreversible fouling). Removing one of these organic fractions often results in more stable operation. Ozonation in all cases led to better operation, but is not always economically feasible. The feasibility of ozone as pre-treatment depends largely on the initial ozone demand, and whether or not there are secondary treatment targets (e.g., higher virus removal, taste, and/or odor).

Effect of Black Tea and Black Tea Pomace Polyphenols on α-Glucosidase and α-Amylase Inhibition, Relevant to Type 2 Diabetes Prevention.

This study evaluates the potential mechanism of action and bioactivity of black tea and black tea pomace for type 2 diabetes prevention via inhibition of carbohydrate hydrolyzing enzymes. Black tea leaves were extracted in hot water and black tea pomace was extracted in 70% acetone. The phenolic content of the water extract (WBT) and pomace acetone extracts (AOBT) were 5.77 and 8.9 mg/mL, respectively, both based on the same concentration of solid tea in the extract. The water extract was subjected to C18 extraction and the resulting hydrophobic fraction (HBBT) was further subjected to LH-20 extraction to recover a low molecular weight phenolic enriched fraction (LMW) and a high molecular weight enriched fraction (HMW). The phenolic content of the LMW and HMW fraction were 1.42 and 2.66 mg/mL, respectively. Among water extracts the HMW fraction was most bioactive against α-glucosidase (IC50 = 8.97 μg/mL) followed by HBBT fraction (IC50 = 14.83 μg/mL). However, the HBBT fraction was the most bioactive fraction against α-amylase (IC50 = 0.049 mg/mL). The black tea pomace (AOBT) had significant α-glucosidase inhibitory activity (IC50 = 14.72 μg/mL) but lower α-amylase inhibitory activity (IC50 = 0.21 mg/mL). The phenolic profiles for LMW and HMW fractions were evaluated using HPLC and the differences between the two profiles were identified. Further research is underway to identify and evaluate the phenolic compounds that are present in the HMW fraction. Our findings suggest that black tea and black tea pomace has potential for carbohydrate hydrolyzing enzyme inhibition and this activity depends on high molecular weight phenolic compounds.

Differentiation of degrees of ripeness of Catuai and Tipica green coffee by chromatographical and statistical techniques

The quality of green coffee is influenced by the degree of ripeness of the fruit at harvest. The aim of this study was to identify chemical markers differentiating between degrees of ripeness. Two coffee varieties, Catuai and Tipica, from the same farm were analysed using the following parameters and methods: caffeine and chlorogenic acid content using high-performance liquid chromatography (HPLC), sucrose content using hydrophilic interaction chromatography, high-molecular weight fraction (HMW) using high-performance size-exclusion chromatography (HPSEC) and volatile compounds using headspace solid phase micro extraction gas chromatography/mass spectrometry. The best method for differentiating between degrees of ripeness was found to be principal component analysis (PCA) based on HPLC data. HPSEC showed differences in the HMW fraction for different degrees of ripeness and both coffee varieties. Volatile profiles allowed separation of both varieties; yet, except for ripe Catuai, no separation was achieved for the degree of ripeness.

Fractionation of five technical lignins by selective extraction in green solvents and characterisation of isolated fractions

Lignins from softwood, hardwood, grass and wheat straw were fractionated by selective extraction at ambient temperature using green solvents like acetone/water solutions of 10, 30, 50, 70 and 90% (v/v) acetone and ethyl acetate. A comparison between the isolated fractions and unfractionated lignins was made in terms of extraction yield, lignin solubility factor, molecular weight distribution and functional group composition. Low molecular weight (LMW) lignin fractions with narrow dispersity are obtained by extraction with ethyl acetate and acetone–water solution containing 30% acetone, with yields depending on the type and the functional group content of lignins. A significant amount (56%) of the organosolv hardwood lignin with low molecular weight (Mw=1868g/mol) and low dispersity was isolated from ethyl acetate. Insoluble fractions with very high molecular weight (Mw between 10 and 17kg/mol) are obtained in low yield from acetone–water solutions with 50, 70 and 90% acetone. LMW lignins are in general less condensed and have lower aliphatic hydroxyl content than parent lignins while the HMW fractions have a higher content of condensed hydroxyls. Principal component analysis on the chemical composition of lignins and isolated fractions determined from 31P NMR data showed the high heterogeneity of the technical lignins. Partial least squares models based on FT-IR spectral data were developed to predict the functional group content determined by quantitative 31P NMR analysis of technical lignins and lignin fractions. This approach can be used to develop simple, rapid and accurate analytical tools to monitor and control the selective fractionation of lignin.

Cocoa brew inhibits in vitro α-glucosidase activity: The role of polyphenols and high molecular weight compounds

Cocoa brew showed a dose-dependent inhibitory activity against α-glucosidase (IC50 7.87mg/mL). The cocoa brew was fractionated by ultrafiltration in a low molecular weight fraction (LMW<10kDa) and two melanoidin-rich fractions called high molecular weight (HMW>30kDa) and intermediate molecular weight (IMW 10–30kDa) fractions. All fractions tested caused some inhibition with IMW that was the most active (IC50 2.37mg/mL). LMW fraction was separated with Sephadex LH-20 in an unbound (containing monomeric and dimeric catechins) and a bound fraction. All the inhibitory activity was recovered in the unbound fraction. All the phenolic compounds identified with HPLC showed α-glucosidase inhibitory activity. IMW and HMW fractions were fractionated by ethanol precipitation. The fractions from IMW precipitated with 75 and 25% ethanol were found to contain power inhibitors of α-glucosidase activity (IC50 0.87 and 1.01mg/mL, respectively). In the HMW sample, the fractions precipitated with 50 and 75% ethanol were found to be active against α-glucosidase activity. Most of the inhibitory activity against α-glucosidase of cocoa brew was due to the LMW fraction (56%) whereas IMW and HMW contributed for about 47% of the inhibitory activity. This study suggests that different components of cocoa may influence α-glucosidase activity.

Online Stable Isotope Analysis of Dissolved Organic Carbon Size Classes Using Size Exclusion Chromatography Coupled to an Isotope Ratio Mass Spectrometer

Stable isotopic content of dissolved organic carbon (δ(13)C-DOC) provides valuable information on its origin and fate. In an attempt to get additional insights into DOC cycling, we developed a method for δ(13)C measurement of DOC size classes by coupling high-performance liquid chromatography (HPLC)-size exclusion chromatography (SEC) to online isotope ratio mass spectrometry (IRMS). This represents a significant methodological contribution to DOC research. The interface was evaluated using various organic compounds, thoroughly tested with soil-water from a C3-C4 vegetation change experiment, and also applied to riverine and marine DOC. δ(13)C analysis of standard compounds resulted in excellent analytical precision (≤0.3‰). Chromatography resolved soil DOC into 3 fractions: high molecular weight (HMW; 0.4-10 kDa), low molecular weight (LMW; 50-400 Da), and retained (R) fraction. Sample reproducibility for measurement of δ(13)C-DOC size classes was ±0.25‰ for HMW fraction, ± 0.54‰ for LMW fraction, and ±1.3‰ for R fraction. The greater variance in δ(13)C values of the latter fractions was due to their lower concentrations. The limit of quantification (SD ≤0.6‰) for each size fraction measured as a peak is 200 ng C (2 mg C/L). δ(13)C-DOC values obtained in SEC mode correlated significantly with those obtained without column in the μEA mode (p < 0.001, intercept 0.17‰), which rules out SEC-associated isotopic effects or DOC loss. In the vegetation change experiment, fractions revealed a clear trend in plant contribution to DOC; those in deeper soils and smaller size fractions had less plant material. It was also demonstrated that the technique can be successfully applied to marine and riverine DOC without further sample pretreatment.