Phenolic profiles of Andean purple corn (Zea mays L.)

Qualitative and quantitative high performance liquid chromatography with diode array detection (HPLC-DAD) was performed to characterize the presence of phenolic compounds in Andean purple corn. Phenolic compounds were analyzed by separating them in two main fractions: a water fraction (WF) and an ethyl acetate fraction (EAF). The WF rich in anthocyanins revealed the presence of cyanidin-3-glucoside, pelargonidin-3-glucoside and peonidin-3-glucoside. The respective acylated anthocyanin glucoside forms of these compounds were also detected following alkaline hydrolysis. The EAF was composed of phenolic acids such as p-coumaric, vanillic acid, protocatechuic acid, flavonoids such quercetin derivatives and a hesperitin derivative. Alkaline and acid hydrolysis of the EAF revealed the presence of p-coumaric and ferulic acid as main components in four bound hydroxycinnamic acid forms present in the ethyl acetate fraction.     

Optimisation,characterisation and quantification of phenolic compounds in olive cake

The phenolic compounds in extracts from pressed olive cake were investigated. Free phenolic compounds were extracted from olive cake using methanol. To liberate bound phenolic compounds, the olive cake was subjected to basic and acidic hydrolysis followed by methanol extraction. The individual phenolic compounds and antioxidant activity of the extracts were determined. The highest total phenolic content and antioxidant activity were obtained using methanol extraction for 12 h at 70 °C. The RP-HPLC profiles for full-fat and defatted olive cake showed that protocatechuic acid, hydroxybenzoic acid, sinapic acid, p-coumaric acid, rutin and hesperidin were the predominant free phenolic compounds. Meanwhile, syringic acid, sinapic acid, caffeic acid and protocatechuic acid were the predominant bound phenolic acids. A positive correlation was observed between total phenolic content and antioxidant activity. The results indicated that most of the phenolic compounds in olive products were present in their free forms (75–90% of total phenolic content), while bound phenolic compounds were only a small proportion (10–25%) of total phenolic content.     

Metabolism of food phenolic acids by Lactobacillus plantarum CECT 748

Phenolic acids account for almost one third of the dietary phenols and are associated with organoleptic, nutritional and antioxidant properties of foods. This study was undertaken to assess the ability of Lactobacillus plantarum CECT 748^T to metabolize 19 food phenolic acids. Among the hydroxycinnamic acids studied, only p-coumaric, caffeic, ferulic and m-coumaric acids were metabolized by L. plantarum. Cultures of L. plantarum produced ethyl and vinyl derivatives from p-coumaric and caffeic acids, 4-vinyl guaiacol from ferulic acid, and 3-(3-hydroxyphenyl) propionic acid from m-coumaric acid. Among the hydroxybenzoic acids analysed, gallic acid and protocatechuic acid were decarboxylated to pyrogallol and catechol, respectively. Inducible enzymes seem to be involved, at least in m-coumaric and ferulic acid metabolism, since cell-free extracts from cultures grown in the absence of these phenolic acids were unable to metabolize them. Further work is needed for the identification of the enzymes involved, since the knowledge of the metabolism of phenolic compounds is an important issue for the food industry.     

High performance liquid and thin-layer chromatographic determination of phenolic acids in palm (Phoenix dactilifera) products

Phenolic acids have been determined in the Tunisian currant palm tree products, dates and legmi (a fermented sap extract). Reversed phase high performance liquid chromatography has been used and results confirmed for the more concentrated compounds by thin-layer chromatography on silica gel.Gallic acid, protocatechuic, p-hydroxybenzoic, syringic, vanillic, caffeic, p-coumaric and ferulic acids have been identified and the origin of these compounds is discussed.The analysed palm tree products are rich in phenolic acids and are characterized by a high concentration in one particular phenolic acid: ferulic acid for dates, gallic acid for legmi.     

Phenolic profiles of andean mashua (Tropaeolum tuberosum Ruíz & Pavón) tubers: Identification by HPLC-DAD and evaluation of their antioxidant activity

Qualitative high performance liquid chromatography with diode array detection (HPLC-DAD) was performed to characterize non-anthocyanin phenolic compounds in three different coloured mashua genotypes. The ORAC antioxidant activity contribution in the tubers related to the type of phenolic compounds present was also evaluated. Phenolic compounds were analysed by separating them into four main fractions: fraction I obtained by means of a liquid–liquid partition with ethyl acetate and fractions II, III and IV obtained by elution on a Sephadex LH-20 column. Fraction I revealed the presence of gallic acid, gallocatechin, procyanidin B₂ and epigallocatechin. Other phenolic compounds such as hydroxycinnamic and hydroxybenzoic acid derivatives, rutin and/or myricetin derivatives were also present in fraction I. Fraction II was mainly composed of epicatechin, hydroxycinnamic and hydroxybenzoic acid derivatives. Fraction III presented mainly anthocyanins for the purple coloured mashua tubers and rutin, hydroxycinnamic acid and hydroxybenzoic acid derivatives for the yellow coloured genotype. Fraction IV was composed of proanthocyanidins. Alkaline and acid hydrolysis of the different fractions revealed the presence of gallocatechin, epicatechin, p-coumaric acid, o-coumaric acid, cinnamic acid, protocatechuic acid, rutin and quercetin as the main phenolic moieties present. The proanthocyanidin fractions were the major contributors to the ORAC antioxidant activity of the mashua tubers for two of the three genotypes (34.7–39.2%). The results obtained in the present study confirm that mashua tubers constitute a promising source of antioxidant phenolics and could potentially be considered as a functional food with beneficial health effects.     

Phenolic profile of seventeen Portuguese wild mushrooms

Analysis of phenolic compounds in seventeen Portuguese wild mushroom species was carried out by high-performance liquid chromatography coupled to photodiode array detection (HPLC-DAD). Protocatechuic, p-hydroxybenzoic, p-coumaric and cinnamic acid were found and quantified. Fistulina hepatica showed the highest phenolic acids concentration (111.72 mg/Kg, dw) due to the significant contribution of protocatechuic (67.62 mg/Kg) and p-hydroxybenzoic (41.92 mg/kg) acids. The edible mushrooms analysed could be directly used in the human diet to combat oxidative stress, while inedible species could represent a source of extractable phenolic compounds to be used as additives in the food industry or as components in pharmaceutical and cosmetic formulations, due to their well-known antioxidant properties.     

Antioxidant capacity and phenolic composition of different woods used in cooperage

This study has investigated the antioxidant capacity of different woods used in cooperage and the effect of the botanical species of wood on this capacity. Quercus robur and Castanea sativa were the species with the highest antioxidant capacity, due to their relatively high content of phenolic compounds. However, the phenolic content of Prunus avium samples was very low, also reflected in its antioxidant capacity. All measurements of antioxidant capacity were consistent with the content and composition of the phenolic compounds detected in the wood samples. The major contributors to the antioxidant capacity were identified as phenolic acids, including gallic, protocatechuic, p-coumaric and ellagic acid and all the ellagitannins, due to their characteristic structure.     

Comparison of phenolic compound contents and antioxidant capacities of loquat (Eriobotrya japonica Lindl.) fruits

The characteristics, phenolic compound contants, and antioxidant capacities of 6 cultivars of loquat fruit grown in China were evaluated. HPLC was used to identify and quantify phenolics. Chlorogenic acid, neochlorogenic acid, 4-O-caffeoylquinic acid, protocatechuic acid, 4-hydroxybenzoic acid, caffeic acid, ferulic acid, ellagic acid, and o-coumaric acid were the main phenolic compounds of mature loquat fruits. Contents of chlorogenic acid, neochlorogenic acid, 4-hydroxybenzoic acid, protocatechuic acid, and o-coumaric acid were all significantly correlated with antioxidant capacities determined using DPPH, ABTS radical scavenging activity, and ferric reducing/antioxidant power assays. The ‘Taxiahong’ cultivar contained the highest amounts of total phenolics and flavonoids and the highest antioxidant capacity, while ‘Taipingbai’ showed the lowest. The high level of phenolic compounds and antioxidant capacities of some cultivars indicates that these cultivars can be sources of bioactive compounds that are relevant to human health.     

Effect of hydrolysis of casein by plasmin on the heat stability of milk

This study examined the effect of hydrolysis of casein by added plasmin (6 mg L⁻¹) on the heat stability of raw, pre-heated, serum protein-free or concentrated skim milk. Plasmin activity markedly affected the heat stability–pH profile of skim milk and serum protein-free milk, apparently by altering the properties of the casein micelles. It is probable that changes in the surface charge of the micelles, as a result of the hydrolysis of caseins, contributed to this effect. Hydrolysis by plasmin reduced the zeta-potential of the casein micelles from ∼−19 to ∼−16 mV. The effect of hydrolysis of casein by plasmin on the heat stability of pre-heated milk was less pronounced, shifting the heat stability–pH profile to more alkaline values; the heat stability of concentrated milk was unaffected by plasmin. A very high (50 mg L⁻¹) level of added plasmin resulted in clearing of the skim milk; the L^* value decreased from ∼75 to ∼35 after 24 h incubation at 37 °C. Clearing was correlated with a change in casein micelle diameter from an initial value of ∼175 to ∼250 nm. It is suggested that plasmin-induced changes in zeta-potential may promote micellar aggregation or changes in micelle stucture.     

Effect of phenolic acids on glucose and organic acid metabolism by lactic acid bacteria from wine

The influence of phenolic (p-coumaric, caffeic, ferulic, gallic and protocatechuic) acids on glucose and organic acid metabolism by two strains of wine lactic acid bacteria (Oenococcus oeni VF and Lactobacillus hilgardii 5) was investigated. Cultures were grown in modified MRS medium supplemented with different phenolic acids. Cellular growth was monitored and metabolite concentrations were determined by HPLC-RI. Despite the strong inhibitory effect of most tested phenolic acids on the growth of O. oeni VF, the malolactic activity of this strain was not considerably affected by these compounds. While less affected in its growth, the capacity of L. hilgardii 5 to degrade malic acid was clearly diminished. Except for gallic acid, the addition of phenolic acids delayed the metabolism of glucose and citric acid in both strains tested. It was also found that the presence of hydroxycinnamic acids (p-coumaric, caffeic and ferulic) increased the yield of lactic and acetic acid production from glucose by O. oeni VF and not by L. hilgardii 5. The results show that important oenological characteristics of wine lactic acid bacteria, such as the malolactic activity and the production of volatile organic acids, may be differently affected by the presence of phenolic acids, depending on the bacterial species or strain.     

Changes in colour,antioxidant activities and carotenoids (lycopene, β-carotene,lutein) of marigold flower (Tagetes erecta L.) resulting from different drying processes

Marigold (Tagetes erecta L.) flower has long been used as a food colourant and ingredient in human food and animal feed. Drying is one of the most important processes for producing marigold powder. Therefore the effects of different drying processes, namely freeze drying (FD), hot air drying (HA) and combined far-infrared radiation with hot air convection (FIR-HA), on the colour, carotenoids (lycopene, β-carotene and lutein) and phenolic compounds of marigold flowers were evaluated. The results indicate that colour changes were less for FIR-HA dried marigold than after FD and HA drying. Different drying methods resulted in changes in the content of individual bioactive compounds. HA gave the highest content of β-carotene (15.5 mg/100 g dry weight (DW)), while FIR-HA and FD provided the highest levels of lutein and lycopene. The predominant phenolic acids in all samples of marigold are p-coumaric acid, ferulic acid and sinapic acid. Gallic, protocatechuic, caffeic, syringic, p-coumaric and ferulic acid showed the highest content in marigold after FIR-HA drying. These results demonstrate that FIR-HA should be considered as a suitable drying method for marigold with respect to preserving its colour, antioxidant properties and bioactive compounds and provided useful information for industrial production of marigold powder.     

Phenolic acids in the meal of developing and stored barley seeds

The following free phenolic acids were identified in the meal of developing barley seeds: salicylic, p-hydroxybenzoic, vanillic, protocatechuic, o-, m- and p-coumaric, syringic, ferulic and sinapic acids. The highest concentration of these compounds was observed at 19–31 days after anthesis. At this time the highest amount of total ethanol-soluble, bound phenolic acids was also found. In the ripe seeds or seeds stored for 2–6 weeks after harvesting, the content of free and soluble bound phenolic acids was distinctly lower. No free caffeic or chlorogenic acids were detected in the seeds by the procedure applied.     

Supramolecular structure of the casein micelle

The supramolecular structure of colloidal casein micelles in milk was investigated by using a sample preparation protocol based on adsorption of proteins onto a poly-l-lysine and parlodion-coated copper grid, staining of proteins and calcium phosphate by uranyl oxalate, instantaneous freezing, and drying under a high vacuum. High-resolution transmission electron microscopy stereo-images were obtained showing the interior structure of casein micelles. On the basis of our interpretation of these images, an interlocked lattice model was developed in which both casein-calcium phosphate aggregates and casein polymer chains act together to maintain casein micelle integrity. The caseins form linear and branched chains (2 to 5 proteins long) interlocked by the casein-stabilized calcium phosphate nanoclusters. This model suggests that stabilization of calcium phosphate nanoclusters by phosphoserine domains of α_s1-, α_s2-, or β-casein, or their combination, would orient their hydrophobic domains outward, allowing interaction and binding to other casein molecules. Other interactions between the caseins, such as calcium bridging, could also occur and further stabilize the supramolecule. The combination of having an interlocked lattice structure and multiple interactions results in an open, sponge-like colloidal supramolecule that is resistant to spatial changes and disintegration. Hydrophobic interactions between caseins surrounding a calcium phosphate nanocluster would prevent complete dissociation of casein micelles when the calcium phosphate nanoclusters are solubilized. Likewise, calcium bridging and other electrostatic interactions between caseins would prevent dissociation of the casein micelles into casein-calcium phosphate nanocluster aggregates when milk is cooled or urea is added to milk, and hydrophobic interactions are reduced. The appearance of both polymer chains and small aggregate particles during milk synthesis would also be expected based on this interlocked lattice model of casein micelles, and its supramolecule structure thus exhibits the principles of self-aggregation, interdependence, and diversity observed in nature.     

Stability of casein micelles cross-linked by transglutaminase

In this study, caseins micelles were internally cross-linked using the enzyme transglutaminase (TGase). The integrity of the micelles was examined on solubilization of micellar calcium phosphate (MCP) or on disruption of hydrophobic interactions and breakage of hydrogen bonds. The level of monomeric caseins, determined electrophoretically, decreased with increasing time of incubation with TGase at 30°C; after incubation for 24 h, no monomeric β- or κ-caseins were detected, whereas only a small level of monomeric α_S1-casein remained, suggesting near complete intramicellar cross-linking. The ability of casein micelles to maintain structural integrity on disruption of hydrophobic interactions (using urea, sodium dodecyl sulfate, or heating in the presence of ethanol), solubilization of MCP (using the calcium-chelating agent trisodium citrate) or high-pressure treatment was estimated by measurement of the L*-value of milk; i.e., the amount of back-scattered light. The amount of light scattered by casein micelles in noncross-linked milk was reduced by >95% on complete disruption of hydrophobic interactions or complete solubilization of MCP; treatment of milk with TGase increased the stability of casein micelles against disruption by all methods studied and stability increased progressively with incubation time. After 24 h of cross-linking, reductions in the extent of light scattering were still apparent in the presence of high levels of dissociating agents, possibly through citrate-induced removal of MCP nanoclusters from the micelles, or urea- or sodium dodecyl sulfate-induced increases in solvent refractive index, which reduce the extent of light-scattering.     

The distribution of phenolic acids in pumpkin’s hull-less seed,skin, oil cake meal,dehulled kernel and hull

The phenolic acids in whole hull-less seed, its skin and oil cake meal, dehulled kernel and hull of pumpkin (Cucurbita pepo) were separated into free, esterified and insoluble-bound forms, which were then identified and quantified by high-performance liquid chromatography with a photodiode array detector. In all samples, protocatechuic, p-hydroxybenzoic, vanillic, trans-p-coumaric, ferulic, trans-sinapic acids and p-hydroxybenzaldehyde were quantified. Caffeic acid was present in all samples except in hulls, while syringic acid was not detectable only in skin and oil cake meal. p-Hydroxybenzoic acid was the dominant phenolic compound in all investigated samples, with 34.7%, 52.0%, 51.4%, 67.4% and 51.8% found in hull-less seed, oil cake meal, skin, dehulled kernels and hulls, respectively, based on total phenolic acid content. Most phenolic acids were present in bound (esterified and insoluble) form, from 50.6% in skin to 84.1% in hull-less seed.     

Study of the inhibitory activity of phenolic compounds found in olive products and their degradation by Lactobacillus plantarum strains

Lactobacillus plantarum is the main species responsible for the spontaneous fermentation of Spanish-style green olives. Olives and virgin oil provide a rich source of phenolic compounds. This study was designed to evaluate inhibitory growth activities of nine olive phenolic compounds against four L. plantarum strains isolated from different sources, and to explore the L. plantarum metabolic activities against these phenolic compounds. None of the nine compounds assayed (oleuropein, hydroxytyrosol, tyrosol, as well as vanillic, p-hydroxybenzoic, sinapic, syringic, protocatechuic and cinnamic acids) inhibited L. plantarum growth at the concentration found in olive products. Oleuropein and tyrosol concentrations higher than 100 mM were needed to inhibit L. plantarum growth. On the other hand, sinapic and syringic acid showed the highest inhibitory activity since concentrations ranging from 12.5 to 50 mM inhibited L. plantarum growth in all the strains analyzed. Among the nine compounds assayed, only oleuropein and protocatechuic acid were metabolized by L. plantarum strains grown in the presence of these compounds. Oleuropein was metabolized mainly to hydroxytyrosol, while protocatechuic acid was decarboxylated to catechol. Metabolism of oleuropein was carried out by inducible enzymes since a cell-free extract from a culture grown in the absence of oleuropein was unable to metabolize it. Independent of their isolation source, the four L. plantarum strains analysed showed similar behaviour in relation to the inhibitory activity of phenolic compounds, as well as their ability to metabolize these compounds.     

Proteolysis in rennet casein-based cheese analogues

Rennet caseins manufactured from mid- or late-lactation milk were used to prepare pilot- and commercial-scale Mozzarella-type cheese analogues. Mid-lactation rennet casein and analogues made from it contained low levels of plasmin; late-lactation rennet caseins and analogues made from them had higher levels of plasmin while pilot-scale analogues had higher levels of plasmin than commercial-scale analogues. The cheese analogues were stored at 8°C for 32 weeks during which time total bacterial counts increased from ∼10⁴ to 10⁷–10⁸ cfu g⁻¹, with non-starter lactic acid bacteria dominating the microflora. Proteolysis, as reflected by pH 4.6-soluble nitrogen and free amino acids, increased on storage of all the analogues. The increase was greater in the analogues manufactured from late-lactation rennet caseins than in those manufactured from mid-lactation rennet casein and in pilot-scale analogues than in commercial-scale analogues, reflecting the differences in plasmin levels as well as slight differences in composition. Polyacrylamide gel electrophoretograms of the cheese analogues showed that β-casein was hydrolysed more extensively than α_s1-casein and that the concentration of γ-caseins increased on storage. The degree of hydrolysis of β-casein was greater in the cheese analogues manufactured from late-lactation rennet caseins than in those manufactured from mid-lactation rennet casein and in the pilot-scale analogues than in the commercial-scale analogues reflecting the corresponding plasmin levels. Seventeen peptides in the pH 4.6-soluble extract of the cheeses were isolated and partially sequenced; all originated from the N-terminal region of β-casein; 9 peptides had Lys₂₉ of β-casein as the N-terminal, 6 peptides had Arg₁ of β-caesin as the N-terminal and the remaining two peptides had Ile₃₀ and Phe₃₃ of β-casein as the N-terminal, respectively. The pattern of casein hydrolysis in the cheese analogues suggests that plasmin was the primary proteolytic agent contributing to initial hydrolysis of the caseins while microbial proteinases and peptidases may have contributed to the high levels of free amino acids.     

Ability of Lactobacillus brevis strains to degrade food phenolic acids

The potential to degrade 15 food phenolic acids was investigated for several Lactobacillus brevis strains isolated from different sources. All the strains analysed in this study showed a similar metabolism on phenolic acids. Among the cinnamic acids assayed, only p-coumaric, ferulic and caffeic acids were metabolized by the L. brevis strains. These acids were decarboxylated to produce their corresponding vinyl derivatives. Contrarily to the results previously reported on Lactobacillus plantarum, the L. brevis strains analysed in this study were unable to subsequently reduce or metabolize these vinyl derivatives. In L. brevis, vinyl phenol, vinyl catechol, and vinyl guaiacol were the final metabolic products from p-coumaric, caffeic or ferulic acids, respectively. From the benzoic acids analysed, and similarly to L. plantarum strains, only gallic and protocatechuic acids were modified by L. brevis strains. Both acids were decarboxylated to pyrogallol and catechol, respectively. Currently, the enzymes involved in the metabolism of phenolic acids in L. brevis remain uncharacterized.     

Sodium caseinate-stabilized fat globules inhibition of the rennet-induced gelation of casein micelles studied by Diffusing Wave Spectroscopy

Sodium caseinate (NaCas)-stabilized oil-in-water emulsions were added to skim milk and the rennet-induced aggregation was observed in situ using light scattering and dynamic oscillatory rheology. The gelation of the recombined milk was greatly inhibited by the addition of the oil droplets, at volume fractions >0.025. The development of the turbidity parameter, 1/l*, and the apparent hydrodynamic radius during renneting were determined using diffusing wave spectroscopy. Although the recombined milk samples contained two scattering particles, namely, casein micelles and fat globules, the latter overwhelmingly contributed to the overall light-scattering signal. This made possible to follow the behaviour of NaCas-stabilized fat globules during the gelation process. The enzymatic reaction associated with the hydrolysis of micellar κ-casein was not significantly affected by the presence of the NaCas-stabilized fat globules. However, the emulsion droplets impeded the aggregation of rennet-altered casein micelles preventing the formation of a gel network. The inability of renneted casein micelles to develop a gel network can be attributed in part to an altered equilibrium between soluble and micellar calcium phosphate, caused by the association of soluble Ca²⁺ with casein molecules, but mostly can be attributed to the effect of non-adsorbed caseins on the surface of the casein micelles.     

Phenolic Profile and Antioxidant Activity of Polish Meads

The purpose of this study was to evaluate phenolic profile and antioxidant activity of ten different meads of 1:1 and 1:2 types, produced with addition of fruit juices, root spices, and herbs. The total phenolic content in meads varied from 15.27 to 70.80 mg/dm³. The meads originated from dark honeys exhibited the highest antioxidant activity. The predominant phenolic compounds in meads were hydroxybenzoic acids, especially gallic and protocatechuic acid, originated mainly from honeys. Whereas, among the hydroxycinnamic acids, the major phenolic was chlorogenic acid, derived mainly from plant additives used in meads production. A principal component analysis was applied in order to differentiate the investigated meads. The 1:2 meads type could be best described by chlorogenic acid. Among the 1:1 meads type, those made from the dark honeys could be best described by gallic, p-coumaric, and protocatechuic acid while those made with addition of blackcurrant and raspberry juice could be described by caffeic and ferulic acid.