Changes of phenolic acids and antioxidant activities during potherb mustard (Brassica juncea, Coss.) pickling

Phenolic acids in potherb mustard (Brassica juncea, Coss.) were determined and the effects of pickling methods on the contents of total free phenolic acids, total phenolic acids, total phenolics, and antioxidant activities were investigated. Gallic acid, protocatechuic acid, p-hydroxybenzoic acid, vanillic acid, caffeic acid, p-coumaric acid, ferulic acid, and sinapic acid were identified in the present study. The contents of total free phenolic acids, total phenolic acids and total phenolics in fresh potherb mustard were 84.8 ± 0.58 μg/g dry weight (DW), 539 ± 1.36 μg/g DW, and 7.95 ± 0.28 mg/g DW, respectively. The total free phenolic acids increased during the pickling processes, but the total phenolic acids, total phenolics, and antioxidant activities decreased. However, after 5 weeks of fermentation, all the pickling methods retained over 70% of total phenolic contents and above 65% of antioxidant capacities. The results indicated that pickling processes were relatively good methods for the preservation of phenolic acids and antioxidants for potherb mustard.     

Development and validation of a duplex real-time PCR method for the simultaneous detection of celery and white mustard in food

The developed duplex real-time PCR method allows the simultaneous detection of traces of potentially allergenic white mustard (Sinapis alba) and celery roots (Apium graveolens var. rapaceum), celery stalks (A. g. var. dulce) and leaf celery (A. g. var. secalinum). The duplex assay does not show any cross-reactivity with 64 different biological species, including various members of the Brassicaceae and Apiaceae family. In raw model sausages spiked with white mustard and celery roots, the LOD was found to be 0.001% white mustard and 0.005% celery. In model sausages brewed at 75–78 °C for 15 min the LOD was found to be 0.005% white mustard and 0.005% celery. The duplex real-time PCR assay was applied to check if commercial food products are labelled in compliance with the legal regulations.     

Chemical composition of white (Morus alba), red (Morus rubra) and black (Morus nigra) mulberry fruits

In this study, the chemical composition of white (Morus alba L.), red (Morus rubra L.) and black (Morus nigra L.) mulberry fruits grown in the East Anatolia Region of Turkey was investigated. The highest total phenolic and flavonoid contents were observed in black mulberry (1422 mg gallic acid equivalents/100 g fresh matter and 276 mg quercetin equivalents/100 g fresh matter). M. alba had the highest total fat content (1.10%), followed by M. nigra (0.95%) and M. rubra (0.85%), respectively. The major fatty acids in mulberry fruits were linoleic acid (54.2%), palmitic acid (19.8%) and oleic acid (8.41%), respectively. The total soluble solids content of mulberry species varied between 15.9% (M. rubra L.) and 20.4% (M. alba L.), acidity between 0.25% (M. alba L.) and 1.40% (M. nigra L.), pH between 3.52 (M. nigra L.) and 5.60 (M. alba L.), ascorbic acid 19.4 mg/100 g (M. rubra L.) and 22.4 mg/100 g (M. alba L.), respectively. Mineral compositions of the mulberry species were 0.83% N, 235 mg/100 g P, 1141 mg/100 g K, 139 mg/100 g Ca, 109 mg/100 g Mg, 60 mg/100 g Na, 4.3 mg/100 g Fe, 0.4 mg/100 g Cu, 4.0 mg/100 g Mn and 3.1 mg/100 g Zn, respectively.     

Natural occurrence of bisphenol F in mustard

Bisphenol F (BPF) was found in mustard up to a concentration of around 8 mg kg^?1. Contamination of the raw products or caused by the packaging could be ruled out. Also, the fact that only the 4,4?-isomer of BPF was detected spoke against contamination from epoxy resin or other sources where technical BPF is used. Only mild mustard made of the seeds of Sinapis alba contained BPF. In all probability BPF is a reaction product from the breakdown of the glucosinolate glucosinalbin with 4-hydroxybenzyl alcohol as an important intermediate. Hot mustard made only from brown mustard seeds (Brassica juncea) or black mustard seeds (Brassica nigra) contained no BPF. BPF is structurally very similar to bisphenol A and has a similar weak estrogenic activity. The consumption of a portion of 20 g of mustard can lead to an intake of 100–200 µg of BPF. According to a preliminary risk assessment, the risk of BPF in mustard for the health of consumers is considered to be low, but available toxicological data are insufficient for a conclusive evaluation. It is a new and surprising finding that BPF is a natural food ingredient and that this is the main uptake route. This insight sheds new light on the risk linked to the family of bisphenols.     

Development and validation of a novel real-time PCR method for the detection of celery (Apium graveolens) in food

The paper presents a novel real-time PCR method allowing the detection of traces of celery (Apium graveolens) in complex food matrices. The method is based on the amplification of a sequence of the gene coding for the Apium graveolens NADPH-dependent mannose-6-phosphate reductase. It allows the detection of three commonly used celery varieties, celery roots (Apium graveolens var. rapaceum), celery stalks (Apium graveolens var. dulce) and leaf celery (Apium graveolens var. secalinum) and does not show any cross-reactivity with 64 biological species, including ten members of the Apiaceae family. The limit of detection, determined by analysing serially diluted celery extracts, is 10 pg celery DNA for all three celery varieties. In spiked model sausages, the LOD is 0.005% celery. The real-time PCR method was applied to 26 commercial food products. Celery DNA was found in one out of ten samples without any information about the presence of celery.     

Studies on the low-salt Chinese potherb mustard (Brassica juncea, Coss.) pickle. I—The effect of a homofermentative l(+)-lactic acid producer Bacillus coagulans on starter culture in the low-salt Chinese potherb mustard pickle fermentation

A homofermentative l(+)-lactic acid producer Bacillus coagulans B179 has been evaluated as starter cultures for low-salt potherb mustard pickle process. This study examined the impact of inoculation by B. coagulans B179 as starter culture on the spontaneous lab-scale fermentation process of potherb mustard pickle. Three different salt concentrations trials, i.e., 5 g/100 g, 8 g/100 g and 10 g/100 g, respectively, were carried out. As a result, the addition of starter culture of B. coagulans B179 to 5 g/100 g salinity fermentation showed an inhibiting effect upon the population of non-lactic acid bacteria (LAB) and marked increase in population of LAB and production of titratable acidity but did not significantly influence the pickle end products quantity of acid produced by fermentation as compared with a treatment without inoculation. However, in the higher salt concentrations, i.e., 8 g/100 g and 10 g/100 g NaCl trials, inoculation had no impact on the fermentation of the pickle. From the standpoint of fermentation, as a functional starter culture, inoculated B. coagulans B179 can effectively improve desired native LAB growth and inhibit the growth of undesired fungi, which will thereby allow it to shorten fermentation period and enhance the pickle quality in the low-salt potherb mustard pickle production.     

The antioxidative properties of mustard leaf (Brassica juncea) kimchi extracts on refrigerated raw ground pork meat against lipid oxidation

The efficacy of varying concentration of mustard leaf kimchi ethanolic extracts (MK) in retarding oxidative rancidity was tested with raw ground pork. Freshly ground pork meat was assigned to one of the following five treatments: control (no antioxidants); AC-0.02 (0.02% ascorbic acid); MK-0.05, 0.1, and 0.2 (0.05%, 0.1% and 0.2% MK, respectively). The pH of the samples decreased and the TBARS values and free fatty acids (%) increased considerably (P < 0.05) during storage. The total bacterial count was lower in MK-0.1 and MK-0.2 than the control during storage. The internal L∗ value and a∗ value decreased (P < 0.05) with the addition of MK. The internal b∗ value of MK treatments were higher (P < 0.05) than that for the control and increased incrementally with MK concentration. The TBARS values and free fatty acids (%) of MK-0.02 was lowest among the treatments. The peroxide value of the control increased until 7 days and reached the maximum value at a certain storage time and decreased thereafter. In the other treatments it increased. All treatments had lower concentration of conjugated dienes (P < 0.05) compared to the control sample, after the first day. Mustard leaf kimchi ethanolic extracts exhibited a protective effect against lipid oxidation in raw ground pork.     

Enantioselective degradation of fipronil in Chinese cabbage (Brassica pekinensis)

Chiral pesticide enantiomers often show different bioactivity and residual toxicity, but this property is usually ignored when evaluating the environmental risk and public safety. In this study, a convenient and precise chiral method was developed and validated for measuring fipronil enantiomers in Chinese cabbage (Brassica pekinensis) based on a high-performance liquid chromatography (HPLC) using (R,R) Whelk-O 1 column. Then the proposed method was successfully applied to the study of enantioselective degradation of fipronil in Chinese cabbage under field conditions. The results showed that the degradation of the two enantiomers in Chinese cabbage was proved to be enantioselective and followed pseudo first-order kinetics (R² ? 0.98). The (R)-enantiomer degraded faster than the (S)-enantiomer, resulting in the relative enrichment of (S)-enantiomer in residue. The detected metabolites MB46513 (desthio), MB45950 (sulfide) and MB46136 (sulfone) by GC–MS suggested that degradation was mainly contributed by oxidization, reduction and photodegradation. Due to the more insecticide activity and lower mammalian toxicity of S-form, the higher concentration of S-fipronil may result in higher activity in crop protection and lower risk to environment and human beings compared to the recemate. This result should be considered in future environmental risk and food safety evaluation.     

A filtration-based real-time PCR method for the quantitative detection of viable Salmonella enterica and Listeria monocytogenes in food samples

We developed a novel filtration-based method that can eliminate dead or severally damaged Salmonella enterica and Listeria monocytogenes in food samples. This new method can recover all viable bacteria in less than 30 min, and can be coupled with a subsequent bacterial DNA extraction and real-time PCR. No statically significant differences (p < 0.01) were found between real-time PCR results obtained separately from S. enterica and L. monocytogenes when different ratios of living and dead cells were used. The analytical sensitivity in both cases was 1 genome equivalent (GE), and the quantification was linear (R² > 0.9969) over a 5-log dynamic range with PCR efficiencies >0.9754. When compared with the standard microbiological methods for the detection of these foodborne pathogens, the relative accuracy was excellent ranging from 95.72% to 104.48%. Finally, we applied the pre-treatment method to the direct detection of viable forms of these foodborne pathogens in food samples using yogurt as a model, the results being similar to those obtained using pure cultures.     

A novel real-time polymerase chain reaction-based method for the detection and quantification of lactose-fermenting Enterobacteriaceae in the dairy and other food industries

The presence of lactose-fermenting Enterobacteriaceae and coliforms is routinely assessed to determine the hygienic quality of water and foods, particularly dairy products. This paper reports the use of lacZ-specific primers in an SYBR green I-based real-time PCR method for the easy and rapid detection of coliforms in dairy products. A large number of bacterial species were assayed to establish the specificity of the method. The sensitivity of the method was assessed using artificially contaminated cheeses. The limit of detection was 1 coliform cell in cheese samples enriched for 8 h in a culture medium. The entire procedure, including sample processing, enrichment, DNA extraction, and real-time PCR amplification, can be completed within 10 to 12 h, making it a single-day assay.     

Effect of water content and temperature on glucosinolate degradation kinetics in broccoli (Brassica oleracea var. italica)

The effect of water content and temperature on glucosinolate thermal degradation in broccoli (Brassica oleracea var. italica) was investigated. Broccoli was freeze dried obtaining batches with water content between 13% and 82% (a_w from 0.32 to 0.99). These samples were heated at different temperatures (from 60 to 120 °C) and glucosinolate levels were monitored. To rule out enzymatic breakdown, myrosinase was inactivated prior to heating. Degradation could be described by first-order kinetics for all glucosinolates and all water contents. In the temperature range 60–100 °C the sample with 13% showed the lowest degradation rate, whereas at 120 °C the degradation rate increased with the water content. This particular behavior was reflected by the high activation energy value of the driest sample. Several hypotheses to explain the observed behavior are discussed.     

Rapid detection of Listeria monocytogenes in food using culture enrichment combined with real-time PCR

A rapid method for the detection of Listeria monocytogenes in foods combining culture enrichment and real-time PCR was compared to the ISO 11290-1 standard method. The culture enrichment component of the rapid method is based on the ISO standard and includes 24 h incubation in half-Fraser broth, 4 h incubation in Fraser broth followed by DNA extraction and real-time PCR detection of the ssrA gene of L. monocytogenes. An internal amplification control, which is co-amplified with the same primers as the L. monocytogenes DNA, was also included in the assay. The method has a limit of detection of 1–5 CFU/25 g food sample and can be performed in 2 working days compared to up to 7 days for the ISO standard. A variety of food samples from retail outlets and food processing plants (n = 175) and controls (n = 31) were tested using rapid and conventional methods. The rapid method was 99.44% specific, 96.15% sensitive and 99.03% accurate when compared to the standard method. This method has the potential to be used as an alternative to the standard method for food quality assurance providing rapid detection of L. monocytogenes in food.     

Enantioselective degradation of diclofop-methyl in cole (Brassica chinensis L.)

The stereoselective degradation of the racemic diclofop-methyl and its chiral degradation product, diclofop (the hydrolysate of diclofop-methyl), in cole has been investigated. Both enantiomers of diclofop-methyl and diclofop were extracted by organic solvent and detected by chiral high-performance liquid chromatography–diode array detector (DAD). Cellulose-tris-(3,5-dimethylphenylcarbamate)-based chiral column was used for the chiral separation of the four enantiomers applying a mixture of n-hexane and 2-propanol (98:2) concluding 0.1% TFA as mobile phase at a flow rate of 1.0 mL/min. The assay method was linear over a range of concentrations (0.5–250 mg L⁻¹), and the mean recovery was more than 60% for all the enantiomers. The limit of detection for all the enantiomers was 0.2 μg g⁻¹ in plant. Racemic diclofop-methyl was foliar sprayed to cole. Stereoselective behaviour was observed in the diclofop-methyl degradation process. The (S)-diclofop-methyl dissipated faster than (R)-diclofop-methyl. In the case of diclofop, the generation and degradation rates of (S)-enantiomer were higher than (R)-enantiomer in the plant.     

Contact and fumigant activities of aromatic plant extracts and essential oils against Lasioderma serricorne (Coleoptera: Anobiidae)

Methanol extracts from 30 aromatic medicinal plant species and five plant essential oils were tested for their insecticidal activities against Lasioderma serricorne (F.) adults using direct contact application and fumigation methods. Responses varied with plant material and exposure time. Good insecticidal activity against L. serricorne adults was achieved with extracts of Agastache rugosa whole plant, Cinnamomum cassia bark, Illicium verum fruit and Foeniculum vulgare fruit as well as cinnamon (C. cassia), horseradish (Cocholeria aroracia) and mustard (Brassica juncea) oils applied at 3.5 mg/cm² in a filter paper diffusion method. Over 90% mortality at 3 days after treatment was achieved with an extract of Acorus calamus var. angustatus rhizome. Cinnamon, horseradish and mustard oils, at 0.7 mg/cm², were highly toxic to the adult beetles 1 day after treatment. In a fumigation test with the beetle adults, insecticidal activity of horseradish oil, mustard oil and Foeniculum fruit extract was much more effective in closed cups than in open ones, indicating that the insecticidal activity of these materials was largely attributable to fumigant action. These naturally occurring plant extracts and essential oils could be useful for managing populations of L. serricorne.     

Nutrient contents of kale (Brassica oleraceae L. var. acephala DC.)

Fructose, glucose and sucrose, as the major soluble sugars and citric and malic acids, as the major organic acids, were identified and determined in kale (Brassica oleraceae L. var. acephala DC., black cabbage) leaves. Fructose was the predominant sugar (2011 mg 100 g⁻¹ dry wt) identified, followed by glucose (1056 mg 100 g⁻¹ dry wt) and sucrose (894 mg 100 g⁻¹ dry wt). The contents of citric and malic acids were at 2213 and 151 mg 100 g⁻¹ dry wt in the leaves. The 16:0, 18:2n − 6 and 18:3n − 3 fatty acids were the most abundant fatty acids in the leaves. Considering the level of these fatty acids, 18:3n − 3 was found to be the highest (85.3 μg g⁻¹ dry wt), contributing 54.0% of the total fatty acid content. Linoleic acid (18:2n − 6), being the second most abundant fatty acid was present at 18.6 μg g⁻¹ dry wt, contributing 11.8% of the total fatty acid content. In the seed oil of kale, 22:1n − 9 was the most abundant fatty acid (4198 μg g⁻¹ dry wt, 45.7%), with 18:2n − 6 (1199 μg g⁻¹ dry wt, 12.3%) and 18:1n − 9 (1408 μg g⁻¹ dry wt, 14.8%) being the second next most abundant fatty acids. The most abundant amino acid was glutamic acid (Glu) which was present at 33.2 mg g⁻¹ dry wt. Aspartic acid, which was the second most abundant amino acid, was present at 27.6 mg g⁻¹ dry wt and accounted for 10.2% of the total amino acid content of kale leaf. The amino acid content was assessed by comparing the percentages of the essential amino acids in kale leaf versus those of a World Health Organization (WHO) standard protein. The protein of kale leaf compares well with that of the WHO standard. Only one amino acid, lysine, had a score that fell below 100%; the lysine score of kale leaf was 95%. This study attempts to contribute to knowledge of the nutritional properties of the plant. These results may be useful for the evaluation of dietary information.     

Determination of ergosterol in canola (Brassica napus L.) by liquid chromatography

A sensitive and precise method is described to assay the oilseed canola (Brassica napus L.) for ergosterol, a fungal metabolite indicating spoilage. The ground seed is refluxed in methanol, and the methanol extract is saponified with potassium hydroxide. After addition of water, the mixture is partitioned into n-hexane. The n-hexane extract is dried, reconstituted, and applied to a silica solid-phase extraction cartridge, which is then washed with carbon tetrachloride, and eluted with acetone. The acetone eluate is acetylated, and the ergosterol determined as the acetate by liquid chromatography using a reverse-phase column and absorbency detection at 282 nm. Acetylation is necessary since an unidentified constituent of the matrix co-chromatographs with free ergosterol, and the ergosterol peak cannot otherwise be resolved. Using a 2-h acetylation at 60°C, recoveries of 10 and 25 ppm added ergosterol were 96.3% and 94.0%, respectively (n=4, relative SD=3.8% in each case). Results using a convenient overnight acetylation at 37°C were similar and not significantly different. The limit of detection was approximately 0.3 ppm.     

Extraction of canola seed (Brassica napus) oil using compressed propane and supercritical carbon dioxide

This study aimed to investigate the extraction of canola seed (Brassica napus) oil using supercritical carbon dioxide and compressed propane as solvents. The extractions were performed in a laboratory scale unit at temperatures and pressures of 40, 50 and 60 °C and 20, 22.5 and 25 MPa for carbon dioxide and 30, 45 and 60 °C and 8, 10 and 12 MPa for propane extractions, respectively. The results indicated that pressure and temperature were important variables for the CO₂ extraction, while temperature is the most important variable for the extraction yield with propane. The extraction with propane was much faster than that with carbon dioxide. The characteristics of the extracted oil, that is, the oxidative stability determined by DSC and the chemical profile of fatty acids determined by gas chromatography, were similar for the two solvents. The overall extraction curves were well described by the Sovová model.     

Multiplex real-time PCR for detection and quantification of mycotoxigenic Aspergillus, Penicillium and Fusarium

The most agriculturally and economically important classes of mycotoxins are produced by species of Aspergillus, Penicillium, and Fusarium. Rapid methods to detect mycotoxigenic fungi could help prevent mycotoxins from entering the food chain. The purpose of this research was to develop a multiplex real-time PCR assay to detect and quantify multiple species of mycotoxigenic fungi. A pair of broad-spectrum PCR primers was designed for amplification of the internal transcribed spacer (ITS) regions of rDNA from the mycotoxigenic species. An in silico analysis of the primers revealed the presence of amplification in more than 40 Aspergillus species, 23 Fusarium species, and 32 Penicillium species as well as 64 other fungal genera. Genus-specific Taqman probes were designed from the ITS sequences of the most important mycotoxigenic species of Fusarium, Penicillium, and Aspergillus. The specificity of the probes was established against a wide range of fungal species. As a multiplex assay, the linear range of detection was 1 pg to 10 ng of DNA. The assay was validated by analyzing fungal growth in distiller's grain (DG), an animal feedstock that is a by-product when ethanol is produced from corn. This assay could be used as an initial step to evaluate the mycotoxigenic potential of DG and various other agricultural commodities.     

Characterization of polyphenol oxidase from broccoli (Brassica oleracea var. botrytis italica) florets

Polyphenol oxidase (PPO) from broccoli florets was extracted and purified through (NH₄)₂SO₄ precipitation, ion-exchange and gel filtration chromatography. The molecular weight was estimated to lie between 51.3 and 57 kDa by sodium dodecyl sulphate-polyacrylamide gel electophoresis (SDS-PAGE) and gel filtration. The effects of substrate specificity, pH, and sensitivity to various inhibitors: citric acid, ascorbic acid, sodium sulphate and EDTA (sodium salt of ethylenediaminetetraacetic acid) of partially purified PPO were investigated. Polyphenol oxidase showed the best activity toward catechol (K_M = 12.34 ± 0.057 mM, V_max = 2000 ± 8736 U/ml/min) and 4-methyl catechol (K_M = 21 ± 0.087 mM, V_max = 28.20 ± 0.525 U/ml/min). The optimum pH for broccoli PPO was 5.7 with catechol and 4-methylcatechol as substrates. The most effective inhibitor was sodium sulphate.     

Interference of phenolic compounds in Brassica napus, Brassica rapa and Sinapis alba seed extracts with the Lowry protein assay

The protein content of aqueous extracts of Brassica napus, Brassica rapa and Sinapis alba meal was determined by the Lowry and Kjeldahl nitrogen assays. Phenolic compounds interfered with the Lowry method to different extents based on the lines studied as well as the extraction procedure used. Three ways to correct for this interference were studied; acid precipitation of the protein before analysis, analyzing in the presence and absence of copper and the binding of free phenolics using non-ionic, porous polystyrene (Amberlite XAD-4). Analysis in presence and absence of copper, and using the difference in absorption at 660 nm between these analyses, proved to be the best way to correct for phenolic interference in the Lowry assay. Extractability of Cruciferae seed phenolics may be pH dependant thus the contribution of phenolics to the Lowry protein assay varies with the pH used for extraction.