Side‐Chain Liquid‐Crystalline Model for Starch

A chiral side‐chain liquid‐crystalline polymeric model for the structure and physical properties of starch was reviewed and extended [1—6]. The lamellae in starch are considered in terms of three components: backbone, side‐chains and double helices. It is the degree of mobility of these three components, coupled with the helix‐coil transition, which gives starch its distinctive properties. The model is used to help explain phenomena involved during hydration, gelatinisation, plasticisation, freezing, and morphogenesis. The new technique of scanning microfocus X‐ray diffraction is used to study double helix orientation and the tilt of the lamellae. Using integration of the double helix orientation along closed contours in single potato granules it is shown that they conain a “+1” hedgehog disclination. The effect of the chirality of the double helices on the phase behaviour is highlighted.     

Small epitope‐linker modules for PCR‐based C‐terminal tagging in Saccharomyces cerevisiae

PCR‐mediated gene modification is a powerful approach to the functional analysis of genes in Saccharomyces cerevisiae. One application of this method is epitope‐tagging of a gene to analyse the corresponding protein by immunological methods. However, the number of epitope tags available in a convenient format is still low, and interference with protein function by the epitope, particularly if it is large, is not uncommon. To address these limitations and broaden the utility of the method, we constructed a set of convenient template plasmids designed for PCR‐based C‐terminal tagging with 10 different, relatively short peptide sequences that are recognized by commercially available monoclonal antibodies. The encoded tags are FLAG, 3 × FLAG, T7, His‐tag, Strep‐tag II, S‐tag, Myc, HSV, VSV‐G and V5. The same pair of primers can be used to construct tagged alleles of a gene of interest with any of the 10 tags. In addition, a six‐glycine linker sequence is inserted upstream of these tags to minimize the influence of the tag on the target protein and maximize its accessibility for antibody binding. Three marker genes, HIS3MX6, kanMX6 and hphMX4, are available for each epitope. We demonstrate the utility of the new tags for both immunoblotting and one‐step affinity purification of the regulatory particle of the 26S proteasome. The set of plasmids has been deposited in the non‐profit plasmid repository Addgene ( http://www.addgene.org ). Copyright © 2009 John Wiley & Sons, Ltd.     

PCR‐RFLP and DAMD‐PCR genotyping for Salvia species

BACKGROUND: Identification of genotypes in Salvia is complicated owing to the morphological similarity and common occurrence of natural hybridisation within Salvia species. Species‐ and genotype‐specific DNA markers are very useful for plant identification, breeding and preservation programmes and can also provide a general overview on the prediction of plant essential oil yield. RESULTS: Polymerase chain reaction restriction fragment length polymorphism (PCR‐RFLP) was used for identification of species‐specific chloroplast and mitochondrial organelle DNA markers, and directed amplification of minisatellite DNA polymerase chain reaction (DAMD‐PCR) was used for genotyping of plant materials. Application of PCR‐RFLP resulted in species‐specific DNA markers, and use of DAMD‐PCR resulted in reproducible DNA patterns that are useful in Salvia genetic studies. Multivariate cluster analysis and principal coordinate analysis indicated that there were relationships between DNA marker patterns and essential oil yields at the species level. CONCLUSION: Results showed that genetic variations in Salvia are wide, and DNA patterns of relatedness among plant species appeared to correlate with essential oil yields. Further studies are required to confirm the application of PCR‐RFLP and DAMD‐PCR markers for selection of Salvia species with higher essential oil yield. Copyright © 2008 Society of Chemical Industry     

Development of formulations for reduced‐sugar and sugar‐free agar‐based fruit jellies

Because of the increasing consumer awareness of health‐related issues, there is a rising demand for noncariogenic, reduced‐energy confectionery products. This study was carried out to develop formulations for reduced‐sugar and sugar‐free, agar‐based jelly products. This was obtained by substituting sucrose and glucose syrup through a combination of sugar replacers, fibre, gelling agents and sweeteners. The application of a combination of polydextrose, oligofructose, sucralose and erythritol resulted in a sensory sweetness profile that was comparable to that of a sugar‐containing standard product. Concerning texture and sensory properties, reduced‐sugar and sugar‐free jellies processed using the respective formulations were comparable to the sugar‐containing standard products. Consumer evaluation using the Just‐about‐right technique exhibited satisfactory acceptance of the sugar‐free jellies.     

Preparation of Chitosan‐Alginate Nanoparticles for Trans‐cinnamaldehyde Entrapment

Trans‐cinnamaldehyde incorporated chitosan‐alginate nanoparticles were synthesized using the ionic gelation and polyelectrolyte complexation technique. Alginate, chitosan, calcium chloride, and trans‐cinnamaldehyde at predetermined concentrations were complexed electrostatically to optimize particle size and loading efficiency. A final methodology using optimized processing parameters (for example, stirring time, homogenization time, equilibration time, and droplet size) was developed. The best working alginate to chitosan mass ratio was determined to be 1.5:1 at a pH dispersion of 4.7. Particle size (166.26 nm) and encapsulation efficiency (73.24%) were further optimized at this mass ratio using an alginate:calcium chloride mass ratio of 4.8:1, alginate:trans‐cinnamaldehyde mass ratio of 37.5:1, a 18 gauge syringe needle, stirring times of 90 min, 15 min of homogenization at 21000 rpm, and equilibration time of 24 h. Optimized nanoparticles showed increased stability (6 wk) and translucency in solution. The final radical scavenging effect of loaded particles in apple juice was 62% and trans‐cinnamaldehyde was just as available to react in free form as it was in inclusion complexes. The final nanoparticle system with modified and optimized processing parameters reduced the size by 43.6% and increased entrapment efficiency by 17.2%. Nanoparticles resembled a spherical shell and core type arrangement (that is, spherical, distinct, and regular) and were in the size range of 10 to 100 nm.     

Degradation of N‐(1‐Deoxy‐d‐xylulos‐1‐yl)glycine and N‐(1‐Deoxy‐d‐xylulos‐1‐yl)proline using thermal treatment

The formation and degradation of N‐(1‐Deoxy‐d ‐xylulos‐1‐yl)glycine and N‐(1‐Deoxy‐d ‐xylulos‐1‐yl)proline, derived from the secondary amine Maillard reaction in xylose‐amino acid model solutions, were detailed in this study. The identification and quantitative analysis of N‐(1‐Deoxy‐d‐ xylulos‐1‐yl)glycine and N‐(1‐Deoxy‐d‐ xylulos‐1‐yl)proline were carried out using high‐performance anion‐exchange chromatography and high‐performance liquid chromatography. The formation of intermediate and advanced products derived from N‐(1‐Deoxy‐d‐ xylulos‐1‐yl)glycine and N‐(1‐Deoxy‐d‐ xylulos‐1‐yl)proline was also tested using an UV‐Vis spectrophotometer to gain a better comparing of the degradation process of the two important Maillard reaction products using thermal treatment. Results showed that the degradation of N‐(1‐Deoxy‐d‐ xylulos‐1‐yl)glycine was more significant than N‐(1‐Deoxy‐d‐ xylulos‐1‐yl)proline. Moreover, xylose was tested in the degradation products of both N‐(1‐Deoxy‐d‐ xylulos‐1‐yl)glycine and N‐(1‐Deoxy‐d‐ xylulos‐1‐yl)proline, which indicated that the degradation of N‐substituted 1‐amino‐1‐deoxyketoses was a reversible reaction to form reducing sugar.     

Evaluation of Novel Back‐flush Filtration for Removal of Homopolymer from Starch‐g‐PMMA

Starch‐g‐poly(methyl methacrylate) (PMMA) was prepared by emulsion photopolymerization without photoinitiator. Grafting efficiency was determined for two types of starch (high‐amylose and amylopectin) and at several illumination times. Since clogging of the graft copolymer prevented vacuum filtration and Soxhlet extraction was too time‐consuming, a new method to separate the PMMA homopolymer from the graft copolymer was developed. Back‐flush filtration uses an intermittent pressure pulse to clean the filter as the homopolymer is separated from the graft copolymer. Back‐flush filtration was shown to be more efficient by reducing the separation time and solvent use, and the grafting efficiencies obtained with back‐flush filtration compare favorably with those from Soxhlet extraction for the starch‐g‐PMMA copolymer systems studied. More accurate grafting efficiencies could be obtained by applying a nonstick coating to the inside chamber of the back‐flush filtration unit.     

Event‐specific analytical methods for biotech maize MIR 604 and DAS‐59122‐7

BACKGROUND: It is very important to develop analytical methods for genetically modified organism (GMO) labelling systems and living modified organism (LMO) management. The polymerase chain reaction (PCR) is the most efficient DNA‐based analytical method for identifying and quantifying biotech crops. Qualitative PCR methods have been developed to detect the presence of biotech crops, while quantitative PCR methods have been developed to analyse the content of biotech crops. Analytical methods are now required for new biotech maize events, MIR 604 and DAS‐59122‐7. RESULTS: The event‐specific primers and probes were developed for qualitative and quantitative analysis of biotech maize MIR 604 and DAS‐59122‐7 based on the 3′ flanking regions. As a reference molecule, single standard plasmid was developed. The specificity of the qualitative primers was confirmed by the single PCR product, with limits of detection (sensitivity) of 0.01 and 0.05% respectively. In‐house validation of the quantitative methods was performed using six levels of mixing samples (0.1–10% w/w). As a result, the biases from the true value and the relative standard deviations were within the range of ± 30%. The limits of quantitation of the quantitative methods were all 0.1% for real time PCRs of MIR 604 and DAS‐59122‐7. CONCLUSION: In this study, event‐specific analytical methods were developed and applied to the qualitative and quantitative analysis of biotech maize MIR 604 and DAS‐59122‐7. Copyright © 2009 Society of Chemical Industry     

Evaluation of Potassium‐Clavulanate‐Supplemented Modified Charcoal‐Cefoperazone‐Deoxycholate Agar for Enumeration of Campylobacter in Chicken Carcass Rinse

Potassium‐clavulanate‐supplemented modified charcoal‐cefoperazone‐deoxycholate agar (C‐mCCDA) that was described in our previous study was compared with original mCCDA for the enumeration of Campylobacter in pure culture and chicken carcass rinse. The quantitative detection of viable Campylobacter cells from a pure culture, plated on C‐mCCDA, is statistically similar (P > 0.05) to mCCDA. In total, 120 chickens were rinsed using 400 mL buffered peptone water. The rinses were inoculated onto C‐mCCDA and mCCDA followed by incubation at 42 °C for 48 h. There was no statistical difference between C‐mCCDA (45 of 120 plates; mean count, 145.5 CFU/mL) and normal mCCDA (46 of 120 plates; mean count, 160.8 CFU/mL) in the isolation rate and recovery of Campylobacter (P > 0.05) from chicken carcass rinse. The Pearson correlation coefficient value for the number of Campylobacter cells recovered in the 2 media was 0.942. However, the selectivity was much better on C‐mCCDA than on mCCDA plates (P < 0.05). Significantly fewer C‐mCCDA plates (33 out of 120 plates; mean count, 1.9 CFU/mL) were contaminated with non‐Campylobacter cells than the normal mCCDA plates (67 out of 120 plates; mean count, 27.1 CFU/mL). The C‐mCCDA may provide improved results for enumeration of Campylobacter in chicken meat alternative to mCCDA with its increased selectivity the modified agar possess.     

Sensory and compositional attributes of melting‐ and non‐melting‐flesh peaches for the fresh market

A study was conducted to determine differences in sensory and compositional characteristics of melting‐flesh (MF) and non‐melting flesh (NMF) fresh market peach genotypes. Sensory results showed that the NMF fruit (‘Oro A’ and FL 86‐28C) were ‘harder’, less ‘juicy’ and more ‘rubbery’ than their MF (FL 90‐20 and ‘TropicBeauty’) counterparts. A principal component analysis of the sensory data showed a clear distinction between the textural aspects of MF and NMF fruit, but not between their flavour aspects. Likewise, chemical analysis showed that while differences in pH, titratable acidity, and soluble solids were detected among the four genotypes, no consistent grouping could be made based on the MF/NMF nature of the fruit. © 1999 Society of Chemical Industry     

Puromycin‐ and methotrexate‐resistance cassettes and optimized Cre‐recombinase expression plasmids for use in yeast

Here we expand the set of tools for genetically manipulating Saccharomyces cerevisiae. We show that puromycin‐resistance can be achieved in yeast through expression of a bacterial puromycin‐resistance gene optimized to the yeast codon bias, which in turn serves as an easy‐to‐use dominant genetic marker suitable for gene disruption. We have constructed a similar DNA cassette expressing yeast codon‐optimized mutant human dihydrofolate reductase (DHFR), which confers resistance to methotrexate and can also be used as a dominant selectable marker. Both of these drug‐resistant marker cassettes are flanked by loxP sites, allowing for their excision from the genome following expression of Cre‐recombinase. Finally, we have created a series of plasmids for low‐level constitutive expression of Cre‐recombinase in yeast that allows for efficient excision of loxP‐flanked markers. Copyright © 2015 John Wiley & Sons, Ltd.     

Lycopene‐derived bioactive retinoic acid receptors/retinoid‐X receptors‐activating metabolites may be relevant for lycopene's anti‐cancer potential

Dietary consumption of tomato products and especially the red tomato pigment lycopene has been associated with lower risk of cancer. New evidence is emerging toward metabolic pathways mediating the anti‐cancer activities of lycopene. In this review, we explore associations between tomatoes and lycopene intake and cancer and relate this to the metabolic activation pathways of lycopene via carotene oxygenases and further carotenoid/retinoid‐metabolizing enzymes to apo‐lycopenoids. Several of these apo‐lycopenoids have already been identified but up to date no direct connection between lycopene metabolism and apo‐lycopenoids mediated receptor activation pathways has been established. Retinoic acid receptors/retinoid‐X receptors activation pathways in particular, may be mediated via lycopene metabolites that are related to retinoic acids. Various studies have shown an association between lower concentration of insulin‐like growth factor‐1 upon lycopene treatment, cancer incidences, and retinoid‐mediated signaling. In this review, we interrelate tomato/lycopene ingestion and cancer incidence, with metabolic activation of lycopene and retinoid‐mediated signaling. The aim is to discuss a potential mechanism to explain lycopene related anti‐cancer activities by modulation of insulin‐like growth factor‐1 concentrations via lycopene metabolite activation of retinoid‐mediated signaling.