Phenolic Composition, Antiradical, Antimicrobial, and Anti-Inflammatory Activities of Propolis Extracts from North East Spain.

Antioxidant-related parameters and anti-inflammatory and antimicrobial activities against Listeria monocytogenes were assessed in eight North East Spain poplar propolis samples. Propolis extracts (PEs) were obtained using 70% ethanol (PEE) and methanol (PME). Yield and total phenol compounds were higher in PEE. Phenolic acids were analyzed by a high-performance liquid chromatograph-diode array detector. Caffeic and ferulic acids were quantified in all PEE and PME. All samples contained p-coumaric acid (quantified in 6 PEE and in 3 PME). Ascorbic acid was detected in all propolis, but mainly quantified in PME (≤0.37 mg/g PE). Biological properties were tested on PEE. As for antiradical activities, trolox equivalent antioxidant capacity (TEAC) [against 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)•+], ranged between 578 and 4620 µmol trolox/g, 2,2-diphenyl-1-picrylhydrazyl (DPPH) (against DPPH free radical), between 0.049 and 0.094 mg/mL, antioxidant activity against hydroxyl (•OH) radical (AOA), between 0.04 and 11.01 mmol uric acid/g, and oxygen radical absorbance capacity (ORAC) against peroxyl (ROO•) radical between 122 and 3282 µmol trolox/g. Results of TEAC, AOA, and ORAC were significantly correlated. IC50 anti-inflammatory activity ranged from 1.08 to 6.19 mg/mL. Propolis showed higher inhibitory activity against L. monocytogenes CECT934 and L. monocytogenes CP101 by agar well diffusion (P < .05) (10.5 and 10.2 mm, respectively) than against L. monocytogenes CP102 (7.0 mm). Data of this research show that North East Spain propolis may be of interest for pharmaceutical and food industry use.

Relationships among Hydrogen Peroxide Concentration, Catalase, Glucose Oxidase, and Antimicrobial Activities of Honeys.

Honey is a natural sweetener made by bees that exhibits antimicrobial activity, mainly related to its H2O2 content. The aim of this work was to research the H2O2 concentration of 24 Spanish honeys from different botanical origins, studying their possible correlation with glucose oxidase (GOx), catalase (CAT), and anti-Staphylococcus aureus activities (minimal inhibition concentration (MIC), minimal bactericidal concentration (MBC), and percentage of inhibition at 5% (w/v) honey against Staphylococcus aureus), as well as possible correlations among all the analyzed parameters. The results showed that the H2O2 concentration did not depend on the botanical origin of the honeys. There were neither correlations between the H2O2 concentration and the activities of GOx and CAT, nor between GOx and antimicrobial activity. However, CAT and antimicrobial activities were positively correlated. Therefore, CAT could be successfully used as a possible marker of the antimicrobial activity of honeys against Staphylococcus aureus. Furthermore, a linear regression model has been fitted to explain the antimicrobial activity from CAT and GOx activity and H2O2 concentration. Although H2O2 is one of the compounds involved in honey's antibacterial activity, this capacity also strongly depends on other honey components (such as low water activity, acidity, osmolarity, and phenolic compounds). The very high anti-Staphylococcus aureus activity exhibited by all samples could be interesting for commercial honey-based formulations also helping to promote local beekeeping.

Impact of the Drying Procedure and Botanical Origin on the Physico-Chemical and Potentially Bioactive Properties of Honey Powders.

This study was aimed at researching the impact of the drying procedure (using the most appropriate honey-maltodextrin concentration for each drying technique) and botanical origin of honey on the physicochemical and potentially bioactive properties of honey powders that were made using maltodextrin as a carrier. The research was carried out with thyme, lavender, vetch and multifloral honey dehydrated using vacuum drying and freeze drying. The analysed parameters were moisture, water activity, colour, glass transition temperature, powder recovery, hygroscopic index and rate, tapped density, solubility, and phenolics as well as antiradical (ABTS•+, ROO•, •OH and O2•-), anti-inflammatory and antimicrobial (against Staphylococcus aureus, Escherichia coli and Listeria monocytogenes) activities. Freeze drying provided the highest recoveries. Powders obtained using freeze drying showed higher moisture and solubility as well as lower glass transition temperature, density and hygroscopicity than those obtained using vacuum drying. Hygroscopicity, glass transition temperature and antimicrobial activity against St. aureus depended on the drying procedure-honey concentration. Colour, anti-O2•- activity and antimicrobial activity against L. monocytogenes depended on the botanical origin of the raw honey. Moisture, solubility, density, total phenolic content, anti-ABTS•+ and anti-ROO• activities as well as anti-inflammatory activity and antimicrobial activity against E. coli depended on the drying procedure-honey concentration and botanical origin.

Easy Nitrite Analysis of Processed Meat with Colorimetric Polymer Sensors and a Smartphone App.

We have developed an in situ methodology for determining nitrite concentration in processed meats that can also be used by unskilled personnel. It is based on a colorimetric film-shaped sensory polymer that changes its color upon contacting the meat and a mobile app that automatically calculates the manufacturing and residual nitrite concentration by only taking digital photographs of sensory films and analyzing digital color parameters. The film-shaped polymer sensor detects nitrite anions by an azo-coupling reaction, since they activate this reaction between two of the four monomers that the copolymer is based on. The sensory polymer is complemented with an app, which analyzes the color in two different digital color spaces (RGB and HSV) and performs a set of 32 data fittings representing the concentration of nitrite versus eight different variables, finally providing the nitrite concentration of the test samples using the best fitting curve. The calculated concentration of nitrite correlates with a validated method (ISO 2918: 1975) usually used to determine nitrite, and no statistically significant difference between these methods and our proposed one has been found in our study (26 meat samples, 8 prepared, and 18 commercial). Our method represents a great advance in terms of analysis time, simplicity, and orientation to use by average citizens.

Chemical Extraction and Gastrointestinal Digestion of Honey: Influence on Its Antioxidant, Antimicrobial and Anti-Inflammatory Activities.

The effect of chemical extraction and in vitro digestion of different kinds of honey on bioactive compounds (total phenolic compounds and flavonoids) and biological activities (antioxidant, antimicrobial and anti-inflammatory) was investigated. The antioxidant activity was evaluated against three radicals (ABTS•+, ROO•, •OH), and the antimicrobial activity was studied against five bacteria (Staphylococcus aureus, Listeria monocytogenes, Escherichia coli, Streptococcus mutans and Pseudomona aeruginosa) and one yeast (Candida albicans). The results show that in comparison with raw honeys, the methanolic extracts exhibited lower values for phenols, flavonoids and antioxidant activity and higher anti-inflammatory and antimicrobial activities against L. monocytogenes. The higher anti-inflammatory activity indicates a possible use of dried honey extracts in the pharmaceutical and cosmetic industries. The digested honeys showed higher total phenolics and higher antioxidant activity than the pre-digested honeys, as well as higher antimicrobial activity against S. aureus and L. monocytogenes, which underlines the possible antioxidant and antimicrobial effects of honey in the human body after the digestion process.

Polymer film as starch azure container for the easy diastase activity determination in honey.

A new original application for a polyacrylic film based on the monomers 2-(dimethylamino)ethyl methacrylate (NNDA), 2-hydroxyethyl acrylate (2HEA) and methylmethacrylate (MMA) as a starch azure container has been set up for a simple determination of honey diastase activity. The proposed method is based on the correlation of reducing sugars generated during the enzymatic process with the Schade reference assay. The polyacrylic film is charged with starch azure acting as a container for this substance; thus, the starch does not interfere in the measurement of reducing sugars, so that the diastase activity is easily calculated. The method has been contrasted with Schade method, showing good correlation and differences under 0.4% between methods in some honey samples. The polyacrylic film has great potential for the routine honey diastase activity assessment in small laboratories, dramatically reducing analysis time and cost.

A simple one-pot determination of both total phenolic content and antioxidant activity of honey by polymer chemosensors.

We have developed a new method for the rapid (2 h) and inexpensive (materials cost < 0.02 €/sample) "2-in-1" determination of the total phenolic content (TPC) and the antioxidant activity (AOX) in honey samples. The method is based on hydrophilic colorimetric films with diazonium groups, which react with phenols rendering highly colored azo groups. The TPC of the sample is correlated to its trolox equivalent antioxidant capacity (TEAC). The intensity of the color allows us to determine both TPC and TEAC of the sample by the analysis of a picture taken with a smartphone that is analysed by the use of the color-definition-parameters (RGB). The controlled light conditions and the systematic use of the same camera avoid the periodical calibration of the system improving the efficiency of the method. Thus, it is a simple method carried out by non-specialized personnel and it involves much lower money and time investment compared to traditional methods.

Phenolic Profile, Antioxidant Capacities and Enzymatic Inhibitory Activities of Propolis from Different Geographical Areas: Needs for Analytical Harmonization.

Propolis is a resinous vegetal exudate modified by bees, and is interesting as a preservative and potentially functional product. This work dealt with studying the common phenolic profiles and antioxidant capacities of 13 bee propolis from different geographical areas. Both hyaluronidase and angiotensin converting enzyme (ACE) inhibitory activities were also assessed and related when possible with particular phenolic compounds. High performance liquid chromatography-ultraviolet detection (HPLC-UV) analysis showed that every propolis contained p-coumaric acid (1.2-12.2 mg/g) and ferulic acid (0.3-11.0 mg/g). Pinocembrin, catechin, and caffeic acid phenethyl ester (CAPE) plus galangin were the main flavonoids. Antioxidant activities were higher than 280 µmol trolox/g for trolox equivalent antioxidant capacity (TEAC), 0.099 mmol uric acid/g for radical-scavenging effect on hydroxyl radicals, and 0.19 mg/mL for half maximal inhibitory concentration (IC50) of antioxidant activity against superoxide anion radical. Working with solutions of 10 mg/mL propolis, hyaluronidase inhibitory activity ranged between 0% and 68.20%, being correlated to ferulic acid content. ACE inhibitory effect determined by HPLC was higher than 78%, being correlated with catechin and p-coumaric acid. Therefore, propolis could be useful for food, pharmaceutical, and cosmetic companies, also helping to reduce risk factors for diseases related to oxidative damage, inflammatory processes, and hypertension. This research also highlights the necessity for harmonized analysis methods and the expression of results for propolis.

Evolution of hydroxymethylfurfural content of honeys from different climates: influence of induced granulation.

Knowing the 'best-before period once opened' is of paramount importance for consumers, because it gives information about how long a given food keeps its quality. The hydroxymethylfurfural (HMF) content is related to honey's aging and heating. We researched the evolution of HMF of 60 unheated honeys originating from continental and oceanic climate areas, recording the moment at which the HMF increase begins to speed up. We also researched the influence of induced granulation on HMF evolution. All samples were stored at room temperature and analysed every 5 months throughout 30 months. Storage had an effect on HMF contents. On the contrary, induced granulation did not show any influence on this parameter. HMF constantly increased in all samples but much more markedly after 20 months. At 20 months, the percentages of samples with HMF higher than 40 mg/kg ranged between 2.9% and 8.0%. Nevertheless, at 25 months these percentages clearly increased, ranging from 22.9% to 72.0%. For honeys from both the continental and oceanic climate areas, taking HMF contents into account, 20 months could be proposed as the 'best-before period once opened'.

Capillary zone electrophoresis method for the determination of inorganic anions and formic acid in honey.

A capillary zone electrophoresis method for the determination of inorganic anions and formic acid in honey samples was developed for the first time. The complete separation of chloride, nitrate, sulfate, phosphate, and formic acid was achieved with a simple electrolyte composed by 2 mM potassium dichromate as the carrier solution and background absorbance provider and 0.05 mM tetraethylenepentamine (TEPA) as electro-osmotic flow suppressor (pH 4.00). Injection was performed hydrostatically by elevating the sample at 10 cm for 10 s. The running voltage was -27 kV at 25 degrees C. Indirect UV absorption detection was achieved at 254 nm. The detection limit was in the range between 0.03 and 20 mg/kg, and the quantification limits ranged from 1.52 to 20.6 mg/kg. The calibration graphs were linear in the concentration range from the quantification limit to at least 2.5 g/kg for chloride, 0.25 g/kg for nitrate, 0.75 g/kg for sulfate, 1.50 g/kg for phosphate, and 0.75 g/kg for formic acid. Precision data in the honey samples analyzed showed repeatability and reproducibility relative standard deviations lower than 1.4 and 2.4% for migration time and lower than 1.8 and 4.3% for anion content, respectively. Recoveries of anions in honey samples analyzed ranged from 94.4 to 99.8%. Ten honey samples were analyzed to test the proposed method. Mean contents of 260.5, 3.93, 60.5, 139.4, and 209.3 mg/kg were found, respectively, for chloride, nitrate, sulfate, phosphate, and formic acid in analyzed honeys. These results agreed with literature data.

Rapid determination of nonaromatic organic acids in honey by capillary zone electrophoresis with direct ultraviolet detection.

A rapid capillary zone electrophoresis (CZE) method with direct ultraviolet (UV) detection has been set up and developed to determine the most important nonaromatic organic acids in honey with a really simple treatment of the sample. The determination of oxalic, formic, malic, succinic, pyruvic, acetic, lactic, citric, and gluconic acids has been carried out in 4 min. The electrolyte composition was phosphate as the carrier buffer (7.5 mM NaH(2)PO(4) and 2.5 mM Na(2)HPO(4)), 2.5 mM tetradecyltrimethylammonium hydroxide (TTAOH) as electroosmotic flow modifier, and 0.24 mM CaCl(2) as selectivity modifier, with the pH adjusted at 6.40 constant value. The running voltage was -25 kV at a thermostated temperature of 25 degrees C. The injections were performed in hydrodynamic mode (30 s), and the detection mode was UV direct at 185 nm. Validation parameters of the method as detection and quantification limits, linearity, precision (repeatability and reproducibility), and recovery were also studied. The advantages related to the technique such as simplicity, short analysis times, and low consumption of chemicals as well as the good validation parameters obtained for this method permit it to be considered as adequate for routine analysis in honey.

Precise method for the measurement of catalase activity in honey.

An improved method is reported for the determination of catalase activity in honey. We tested different dialysis membranes, dialysis fluid compositions and amounts, dialysis temperatures, sample amounts, and dialysis times. The best results were obtained by dialysis of 7.50 g sample in a cellulose dialysis sack, using two 3 L portions of 0.015 M sodium phosphate buffer (pH 7.0) as the dialysis fluid at 4 degrees C for 22 h. As in previous methods, catalase activity was determined on the basis of the rate of disappearance of the substrate, H202, with the H202 determined spectrophotometrically at 400 nm in an assay system containing o-dianisidine and peroxidase. Trials indicated that the best solvent for the o-dianisidine was 0.2 M sodium phosphate buffer, pH 6.1; the best starting H202 concentration was 3 mM; the best HCl concentration for stopping the reaction was 6 N; and the best sample volume for catalase measurement was 7.0 mL. Precision values (relative standard deviations for analyses of 10 subsamples of each of 3 samples) were high, ranging from 0.48% for samples with high catalase activity to 1.98% for samples with low catalase activity.

Significance of nonaromatic organic acids in honey.

Although organic acids represent < 0.5% of honey's constituents, they make important contributions to the organoleptic, physical, and chemical properties of honey. To date, approximately 30 nonaromatic organic acids have been identified in honey, but relatively little attention has been paid to these components. This article reviews the current literature related to the significance of nonaromatic organic acids in honey; it was written with a goal of attracting researchers to study these interesting honey components. Previous research contributions on nonaromatic organic acids in honey may be classified into five main areas: (i) the antibacterial activities of these acids, (ii) the antioxidant activities of these acids, (iii) the use of these acids as possible indicators of incipient fermentation, (iv) the use of these acids for treatment of Varroa infestation, and (v) the use of these acids as factors for the characterization of both botanical and geographical origins of honeys. We conclude that nonaromatic organic acids are of interest for diverse reasons and that there is a particular need for studies regarding their possible antibacterial and antioxidant activities.

High-pressure liquid chromatographic determination of ascorbic acid in cooked sausages.

The purpose of this paper was to study and optimize both extraction and high-pressure liquid chromatography (HPLC)-UV detection procedures to develop a proper method for the determination of ascorbic acid content in cooked sausages. A simple and sensitive reversed-phase HPLC method for the NH2-bonded phase has been described for the determination of ascorbic acid content in cooked sausages. Various extracting agents were tested to solubilize the vitamin, with 5% (wt/vol) metaphosphoric acid giving the best results. Samples were chromatographed with UV detection at 248 nm on a 25-cm Spherisorb NH2 cartridge with a 0.4-cm inside diameter with a mixture of 0.02 M potassium phosphate buffer solution (pH 3.6) and acetonitrile (40:60, vol/vol) for the mobile phase. The method's precision within a day was 1.2%, and its precision between days was 3.8%. The detection limit was 1.6 mg/100 g. Recovery ranged from 91.4 to 96.2% for ascorbic acid added to meat samples. Twenty samples of six different products were analyzed in duplicate. For the samples analyzed, the mean value for ascorbic acid ranged between 21.555 and 24.899 mg/100 g of fresh weight.

Rapid determination of minority organic acids in honey by high-performance liquid chromatography.

A rapid high-performance liquid chromatographic method for the determination of organic acids in honey is reported. Malic, maleic, citric, succinic and fumaric acids were identified and quantified in 15 min. First time repeatibility, reproducibility and recoveries were determined out for these acids in honey samples. Maleic acid was also quantified for first time by a chromatographic method. The organic acids were removed from honey by using a solid-phase extraction procedure with anion-exchange cartridges. Previously, the solution of honey was adjusted to pH 10.50 with 0.1 M NaOH and stirred for 15 min at room temperature. Then, this solution was adjusted to pH 5.00 with 0.1 M H2SO4. This procedure was carried out to avoid interferences in the baseline. The chromatographic separation was achieved with only one Spherisorb ODS-2 S5 column thermostated at 25 degrees C. Metaphosphoric acid (pH 2.20) was used as mobile phase at a flow-rate of 0.7 ml/min. Organic acids were detected with a UV-vis detector (215 nm). The precision results showed that the relative standard deviations of the repeatability and reproducibility were < or =3.20% and < or =4.86%, respectively. The recoveries of the organic acids ranged from 62.9 to 99.4%. Under optimum conditions the detection limits ranged from 0.0064 to 7.57 mg/kg and the quantification limits ranged from 0.025 to 10.93 mg/kg.