Grape and wine analysis: Oenologists to exploit advanced test kits.
Charnock, S. C. & McCleary, B. V. (2005). Revue des Enology, 117, 1-5.
It is without doubt that testing plays a pivotal role throughout the whole of the vinification process. To produce the best possible quality wine and to minimise process problems such as “stuck” fermentation or troublesome infections, it is now recognised that if possible testing should begin prior to harvesting of the grapes and continue through to bottling. Traditional methods of wine analysis are often expensive, time consuming, require either elaborate equipment or specialist expertise and frequently lack accuracy. However, enzymatic bio-analysis enables the accurate measurement of the vast majority of analytes of interest to the wine maker, using just one piece of apparatus, the spectrophotometer (see previous issue No. 116 for a detailed technical review). Grape juice and wine are amenable to enzymatic testing as being liquids they are homogenous, easy to manipulate, and can generally be analysed without any sample preparation.
Megazyme “advanced” wine test kits general characteristics and validation.
Charnock, S. J., McCleary, B. V., Daverede, C. & Gallant, P. (2006). Reveue des Oenologues, 120, 1-5.
Many of the enzymatic test kits are official methods of prestigious organisations such as the Association of Official Analytical Chemicals (AOAC) and the American Association of Cereal Chemists (AACC) in response to the interest from oenologists. Megazyme decided to use its long history of enzymatic bio-analysis to make a significant contribution to the wine industry, by the development of a range of advanced enzymatic test kits. This task has now been successfully completed through the strategic and comprehensive process of identifying limitations of existing enzymatic bio-analysis test kits where they occurred, and then using advanced techniques, such as molecular biology (photo 1), to rapidly overcome them. Novel test kits have also been developed for analytes of emerging interest to the oenologist, such as yeast available nitrogen (YAN; see pages 2-3 of issue 117 article), or where previously enzymes were simply either not available, or were too expensive to employ, such as for D-mannitol analysis.
Measurement of Starch: Critical evaluation of current methodology.
B. V. McCleary, L. M. J. Charmier & V. A. McKie. (2018). Starch‐Stärke, In Press.
Most commonly used methods for the measurement of starch in food, feeds and ingredients employ the combined action of α‐amylase and amyloglucosidase to hydrolyse the starch to glucose, followed by glucose determination with a glucose oxidase/peroxidase reagent. Recently, a number of questions have been raised concerning possible complications in starch analytical methods. In this paper, each of these concerns, including starch hydrolysis, isomerisation of maltose to maltulose, effective hydrolysis of maltodextrins by amyloglucosidase, enzyme purity and hydrolysis of sucrose and β‐glucans have been studied in detailed. Results obtained for a range of starch containing samples using AOAC Methods 996.11 and 2014 .10 are compared and a new simpler format for starch measurement is introduced. With this method that employs a thermostable α‐amylase (as distinct from a heat stable α‐amylase) which is both stable and active at 100°C and pH 5.0, 10 samples can be analysed within 2 h, as compared to the 6 h required with AOAC Method 2014.10.
Enhanced activity of ADP glucose pyrophosphorylase and formation of starch induced by Azospirillum brasilense in Chlorella vulgaris.
Choix. F. J., Bashan, Y., Mendoza, A. & de-Bashan, L. E. (2014). Journal of Biotechnology, 177, 22-34.
ADP-glucose pyrophosphorylase (AGPase) regulates starch biosynthesis in higher plants and microalgae. This study measured the effect of the bacterium Azospirillum brasilense on AGPase activity in the freshwater microalga Chlorella vulgaris and formation of starch. This was done by immobilizing both microorganisms in alginate beads, either replete with or deprived of nitrogen or phosphorus and all under heterotrophic conditions, using D-glucose or Na-acetate as the carbon source. AGPase activity during the first 72 h of incubation was higher in C. vulgaris when immobilized with A. brasilense. This happened simultaneously with higher starch accumulation and higher carbon uptake by the microalgae. Either carbon source had similar effects on enzyme activity and starch accumulation. Starvation either by N or P had the same pattern on AGPase activity and starch accumulation. Under replete conditions, the population of C. vulgaris immobilized alone was higher than when immobilized together, but under starvation conditions A. brasilense induced a larger population of C. vulgaris. In summary, adding A. brasilense enhanced AGPase activity, starch formation, and mitigation of stress in C. vulgaris.
In vitro hypoglycemic effects of different insoluble fiber-rich fractions prepared from the peel of Citrus sinensis L. cv. Liucheng
Chau, C. F., Huang, Y. L. & Lee, M. H. (2003). Journal of Agricultural and Food Chemistry, 51(22), 6623-6626.
Insoluble fiber-rich fractions (FRFs), including insoluble dietary fiber, alcohol-insoluble solid, and water-insoluble solid, were isolated from the peel of Citrus sinensis L. cv. Liucheng. We found that these three FRFs could effectively adsorb glucose, retard glucose diffusion, and inhibit the activity of α-amylase to different extents. These mechanisms might create a concerted benefit in decreasing the rate of glucose absorption and eventually lower the concentration of postprandial serum glucose. The potential hypoglycemic effects of these FRFs suggested that they could be incorporated as low-calorie bulk ingredients in high-fiber foods to reduce calorie level and control blood glucose level.
Dietary fibers from mushroom sclerotia: 3. In vitro fermentability using human fecal microflora.
Wong, K. H., Wong, K. Y., Kwan, H. S. & Cheung, P. C. K. (2005). Journal of Agricultural and Food Chemistry, 53(24), 9407-9412.
The in vitro fermentability of three novel dietary fibers (DFs) prepared from mushroom sclerotia, namely, Pleurotus tuber-regium, Polyporous rhinocerus, and Wolfiporia cocos, was investigated and compared with that of the cellulose control. All DF samples (0.5 g each) were fermented in vitro with a human fecal homogenate (10 mL) in a batch system (total volume, 50 mL) under strictly anaerobic conditions (using oxygen reducing enzyme and under argon atmosphere) at 37°C for 24 h. All three novel sclerotial DFs exhibited notably higher dry matter disappearance (P. tuber-regium, 8.56%; P. rhinocerus, 13.5%; and W. cocos, 53.4%) and organic matter disappearance (P. tuber-regium, 9.82%; P. rhinocerus, 14.6%; and W. cocos, 57.4%) when compared with those of the cellulose control. Nevertheless, only the W. cocos DF was remarkably degraded to produce considerable amounts of total short chain fatty acids (SCFAs) (5.23 mmol/g DF on organic matter basis, with a relatively higher molar ratio of propionate) that lowered the pH of its nonfermented residue to a slightly acidic level (5.89). Variations on the in vitro fermentability among the three sclerotial DFs might mainly be attributed to their different amounts of interwoven hyphae present (different amounts of enzyme inaccessible cell wall components) as well as the possible different structural arrangement (linkage and degree of branching) of their β-glucans.
Potential hypoglycaemic effects of insoluble fibres isolated from foxtail millets [Setaria italica (L.) P. Beauvois].
Bangoura, M. L., Nsor‐Atindana, J., Zhu, K., Tolno, M. B., Zhou, H. & Wei, P. (2013). International Journal of Food Science & Technology, 48(3), 496-502.
Insoluble fibres were isolated from the two varieties of foxtail millet (white and yellow) grains and evaluated for their hypoglycaemic effects by in vitro studies. The hypoglycaemic effects of these fibres were compared with those of commercial soy insoluble fibre. The results revealed that minimum and maximum amounts of glucose were adsorbed on each sample at 10 and 200 μmol g-1 glucose concentrations respectively, indicating that the glucose adsorption capacity (GAC) of the fibre materials was proportional to glucose concentration for all samples. There was significant (P< 0.05) difference among all the fibre materials in relation to their GAC values. In the case of the effects of the fibres on glucose diffusion, the millets' insoluble fibres performed better than that of the commercial soy insoluble fibres. The glucose dialysis retardation indexes at the end of the maximum dialysis time were 1.1%, 27.4% and 22.6% for soy bean insoluble fibre, white foxtail millet insoluble fibre and yellow foxtail millet insoluble fibre in that order. The study showed that hypoglycaemic effects of yellow and white foxtail millet fibres were comparable to the commercial soy insoluble fibre.
A high-throughput platform for screening milligram quantities of plant biomass for lignocellulose digestibility.
Santoro, N., Cantu, S. L., Tornqvist, C. E., Falbel, T. G., Bolivar, J. L., Patterson, S. E., Pauly, M. & Walton, J. D. (2010). BioEnergy Research, 3(1), 93-102.
The development of a viable lignocellulosic ethanol industry requires multiple improvements in the process of converting biomass to ethanol. A key step is the improvement of the plants that are to be used as biomass feedstocks. To facilitate the identification and evaluation of feedstock plants, it would be useful to have a method to screen large numbers of individual plants for enhanced digestibility in response to combinations of specific pretreatments and enzymes. This paper describes a high-throughput digestibility platform (HTDP) for screening collections of germplasm for improved digestibility, which was developed under the auspices of the Department of Energy-Great Lakes Bioenergy Research Center (DOE-GLBRC). A key component of this platform is a custom-designed workstation that can grind and dispense 1–5 mg quantities of more than 250 different plant tissue samples in 16 h. The other steps in the processing (pretreatment, enzyme digestion, and sugar analysis) have also been largely automated and require 36 h. The process is adaptable to diverse acidic and basic, low-temperature pretreatments. Total throughput of the HTDP is 972 independent biomass samples per week. Validation of the platform was performed on brown midrib mutants of maize, which are known to have enhanced digestibility. Additional validation was performed by screening approximately 1,200 Arabidopsis mutant lines with T-DNA insertions in genes known or suspected to be involved in cell wall biosynthesis. Several lines showed highly significant (p < 0.01) increases in glucose and xylose release (20–40% above the mean). The platform should be useful for screening populations of plants to identify superior germplasm for lignocellulosic ethanol applications and also for screening populations of mutant model plants to identify specific genes affecting digestibility.
Effects of gamma irradiation on starch digestibility of rice with different resistant starch content.
Shu, X., Xu, J., Wang, Y., Rasmussen, S. K. & Wu, D. (2013). International Journal of Food Science & Technology, 48(1), 35-43.
Three rice cultivars (RS3M, RS4H and RS5L) differing in resistant starch contents but similar in genetic background were chosen to study the effects of gamma irradiation on starch physicochemical properties and digestibility. Irradiation increases the resistant starch content in all the three cultivars and in a dose-dependent manner in rice with low-resistant starch content (RS5L). Irradiation decreases apparent amylose content and gelatinisation temperature and changed the starch granule structure, while increasing V-type crystallinity. Starch enzymatic hydrolysis rate was reduced following irradiation, and the effect of irradiation on reducing starch digestibility was negatively correlated with resistant starch content. Treatment with gamma irradiation has therefore a potential for increasing resistant starch content and producing low digestibility of starch in common rice.
Anatomical, chemical, and biochemical characterization of cladodes from prickly pear [Opuntia ficus-indica (L.) Mill.].
Ginestra, G., Parker, M. L., Bennett, R. N., Robertson, J., Mandalari, G., Narbad, A., Lo Curto, R. B., Bisignano, G., Faulds, C. B. & Waldron, K. W. (2009). Journal of Agricultural and Food Chemistry, 57(21), 10323-10330.
Opuntia ficus-indica cladodes represent the green stem of the plant and are generally used as animal feed or disposed of in landfills. The present work investigated the anatomical and chemical composition of Opuntia cladodes, which form the basis of their pharmacological effects. Glucose and galacturonic acid were the main sugars of Opuntia cladodes, whereas high-performance liquid chromatography (HPLC) analysis showed the presence of mainly kaempherol and isorhamnetin glycosides (glucoside and rhamnoside). The presence of high amounts of calcium oxalate crystals was demonstrated by light microscopy on fresh and lyophilized cladodes. No antimicrobial activity was observed even after enzymatic treatment. O. ficus-indica cladodes may retain material tightly associated with cell-wall components, and this property will have the potential to greatly reduce the bioavailability of bioactive compounds.
Structure and digestibility of endosperm water-soluble α-glucans from different sugary maize mutants.
Miao, M., Li, R., Jiang, B., Cui, S. W., Lu, K. & Zhang, T. (2014). Food Chemistry, 143, 156-162.
The structure and digestibility of endosperm water-soluble α-glucans from different sugary-1 maize mutants (Zhongtian 8#, Zhongtian 2# and Pintian 8#) were investigated. The yield of pure glucan was in the range of 25.91–34.38%. The α-glucan belonged to a typical native nano-scale particle and the average particle size was in the following order: Zhongtian 8# >Pintian 8# >Zhongtian 2#. The weight-average molar mass of glucans ranged from 1.69 to 2.08 × 107 g/mol. The branch densities and α-1,6 linkages of Zhongtian 8#, Zhongtian 2# and Pintian 8#, were 8.60%, 8.77% and 9.51%, 7.71%, 6.58% and 6.81%, respectively. The resistant starch (10.06%) of Pintian 8# was lower than other two cultivars. The study showed that water-soluble glucan exhibited α-1,4-linked backbone with α-1,6 branch sites and digestibility was influenced by granule size, ratio of α-1,4 to α-1,6 linkages, molecular fine structure in this set of sugary maize mutants.
Enhanced accumulation of starch and total carbohydrates in alginate-immobilized Chlorella spp. induced by Azospirillum brasilense: II. Heterotrophic conditions.
Choix, F. J., de-Bashan, L. E. & Bashan, Y. (2012). Enzyme and Microbial Technology, 51(5), 300-309.
The effect of the bacterium Azospirillum brasilense jointly immobilized with Chlorella vulgaris or C. sorokiniana in alginate beads on total carbohydrates and starch was studied under dark and heterotrophic conditions for 144 h in synthetic growth medium supplemented with either D-glucose or Na-acetate as carbon sources. In all treatments, enhanced total carbohydrates and starch content per culture and per cell was obtained after 24 h; only jointly immobilized C. vulgaris growing on D-glucose significantly increased total carbohydrates and starch content after 96 h. Enhanced accumulation of carbohydrate and starch under jointly immobilized conditions was variable with time of sampling and substrate used. Similar results occurred when the microalgae was immobilized alone. In both microalgae growing on either carbon sources, the bacterium promoted accumulation of carbohydrates and starch; when the microalgae were immobilized alone, they used the carbon sources for cell multiplication. In jointly immobilized conditions with Chlorella spp., affinity to carbon source and volumetric productivity and yield were higher than when Chlorella spp. were immobilized alone; however, the growth rate was higher in microalgae immobilized alone. This study demonstrates that under heterotrophic conditions, A. brasilense promotes the accumulation of carbohydrates in two strains Chlorella spp. under certain time–substrate combinations, producing mainly starch. As such, this bacterium is a biological factor that can change the composition of compounds in microalgae in dark, heterotrophic conditions.
In vitro starch digestibility, estimated glycemic index and antioxidant potential of Taro (Colocasia esculenta L. Schott) corm.
Simsek, S. & El, S. N. (2015). Food Chemistry, 168, 257-261.
The purpose of this study was to determine some functional properties of taro (Colocasia esculenta L. Schott) corm, which can be a good alternative to the other dietary carbohydrate sources with its high starch content. The total phenolic and flavonoid content of taro corm was found as 205 ± 53 mg CAE/100 g and 61 ± 9 mg CAE/100 g, respectively. The antioxidant capacity of corm was determined as 452 ± 72 mM TEAC/100 g and 244 ± 73 mM TEAC/100 g, by the scavenging activity against ABTS and DPPH radicals, respectively. The free glucose content of corms was less than 1%, whereas the 60% of dry matter was composed of starch. According to the results, the taro corms’ starch was highly digestible and higher than the 50% of the starch was composed of rapidly digestible starch (RDS) fractions. The estimated glycemic index (eGI) of taro corm was 63.1 ± 2.5, indicating taro corm as a medium GI food and a good dietary carbohydrate alternative especially for diabetic people.
Hydrothermal treatment of oleaginous yeast for the recovery of free fatty acids for use in advanced biofuel production.
Espinosa-Gonzalez, I., Parashar, A. & Bressler, D. C. (2014). Journal of Biotechnology, 187, 10-15.
Microbial oils hold great potential as a suitable feedstock for the renewable production of biofuels. Specifically, the use of oleaginous yeasts offers several advantages related to cultivation and quality of lipid products. However, one of the major bottlenecks for large-scale production of yeast oils is found in the lipid extraction process. This work investigated the hydrothermal treatment of oleaginous yeast for hydrolysis and lipid extraction resulting in fatty acids used for biofuel production. The oleaginous yeast, Cryptococcus curvatus, was grown in 5 L bioreactors and the biomass slurry with 53 ± 4% lipid content (dry weight basis) was treated at 280°C for 1 h with initial pressure 500 psi in batch stainless steel reactors. The hydrolysis product was separated and each of the resulting streams was further characterized. The hexane soluble fraction contained fatty acids from the hydrolysis of yeast triacylglycerides, and was low in nitrogen and minerals and could be directly integrated as feedstock into pyrolysis processing to produce biofuels. The proposed hydrothermal treatment addresses some current technological bottlenecks associated with traditional methodologies such as dewatering, oil extraction and co-product utilization. It also enhances the feasibility of using microbial biomass for production of renewable fuels and chemicals.
Effects of heat treatment and moisture contents on interactions between Lauric acid and starch granules.
Chang, F., He, X., Fu, X., Huang, Q. & Jane, J. L. (2014). Journal of Agricultural and Food Chemistry, 62(31), 7862-7868.
This study aimed to understand the effects of the moisture content of granular normal cornstarch (NC), heat treatment at 80°C, and order of adding lauric acid (LA) to starch before or after the heat treatment on the physicochemical properties and digestibility of the starch. LA was added to NC priority heated with different moisture contents (10, 20, 30, 40, and 50%) or added to dried NC and then heated with different moisture contents. The hydrothermal/LA treatments increased the pasting temperature but decreased the peak viscosity of the NC. Light and scanning electron microscopy revealed that the addition of LA retarded gelatinization. The hydrothermal/LA treatments changed the X-ray pattern of the NC to a mixture of A- and V-type patterns. The thermal property and digestibility analysis showed that 40% was the optimum moisture content for the formation of the amylose–LA complex and adding LA prior to heating the NC favored the formation of slowly digestible starch.
In vitro starch digestibility and in vivo glucose response of gelatinized potato starch in the presence of non‐starch polysaccharides.
Sasaki, T., Sotome, I. & Okadome, H. (2015). Starch‐Stärke, 67(5-6), 415-423.
Blending non-starch polysaccharides with starch is expected to change the rate and extent of starch digestion and has a potential effect on glucose response. The objective of this study was to investigate the effects of four water-soluble non-starch polysaccharides: xanthan gum, guar gum, pectin, and konjac-glucomannan, on in vitro starch digestibility and in vivo glucose response of gelatinized potato starch. Gelatinized potato starch was digested rapidly, and the added polysaccharides significantly reduced the digestion of gelatinized potato starch even over a short reaction time. Postprandial blood glucose response in rats was reduced by the addition of polysaccharides to gelatinized potato starch. The effects on the blood glucose level after ingestion were dependent on the type of polysaccharide. The addition of xanthan gum significantly reduced the blood glucose levels at 30 min, whereas guar gum and pectin reduced the levels at 60 min. The interaction between gelatinized potato amylopectin and polysaccharides was investigated using a quartz crystal microbalance. Xanthan gum was found to be immobilized on the layer of gelatinized amylopectin, suggesting that interaction between starch and xanthan gum is one of the major factors in starch hydrolysis inhibition.
Bioprocess automation on a Mini Pilot Plant enables fast quantitative microbial phenotyping.
Unthan, S., Radek, A., Wiechert, W., Oldiges, M. & Noack, S. (2015). Microbial Cell Factories, 14(1), 32.
Background: The throughput of cultivation experiments in bioprocess development has drastically increased in recent years due to the availability of sophisticated microliter scale cultivation devices. However, as these devices still require time-consuming manual work, the bottleneck was merely shifted to media preparation, inoculation and finally the analyses of cultivation samples. A first step towards solving these issues was undertaken in our former study by embedding a BioLector in a robotic workstation. This workstation already allowed for the optimization of heterologous protein production processes, but remained limited when aiming for the characterization of small molecule producer strains. In this work, we extended our workstation to a versatile Mini Pilot Plant (MPP) by integrating further robotic workflows and microtiter plate assays that now enable a fast and accurate phenotyping of a broad range of microbial production hosts. Results: A fully automated harvest procedure was established, which repeatedly samples up to 48 wells from BioLector cultivations in response to individually defined trigger conditions. The samples are automatically clarified by centrifugation and finally frozen for subsequent analyses. Sensitive metabolite assays in 384-well plate scale were integrated on the MPP for the direct determination of substrate uptake (specifically D-glucose and D-xylose) and product formation (specifically amino acids). In a first application, we characterized a set of Corynebacterium glutamicum L-lysine producer strains and could rapidly identify a unique strain showing increased L-lysine titers, which was subsequently confirmed in lab-scale bioreactor experiments. In a second study, we analyzed the substrate uptake kinetics of a previously constructed D-xylose-converting C. glutamicum strain during cultivation on mixed carbon sources in a fully automated experiment. Conclusions: The presented MPP is designed to face the challenges typically encountered during early-stage bioprocess development. Especially the bottleneck of sample analyses from fast and parallelized microtiter plate cultivations can be solved using cutting-edge robotic automation. As robotic workstations become increasingly attractive for biotechnological research, we expect our setup to become a template for future bioprocess development.
Responses of three tropical seagrass species to CO2 enrichment.
Ow, Y. X., Collier, C. J. & Uthicke, S. (2015). Marine Biology, 162(5), 1005-1017.
Increased atmospheric carbon dioxide leads to ocean acidification and carbon dioxide (CO2) enrichment of seawater. Given the important ecological functions of seagrass meadows, understanding their responses to CO2 will be critical for the management of coastal ecosystems. This study examined the physiological responses of three tropical seagrasses to a range of seawater pCO2 levels in a laboratory. Cymodocea serrulata, Halodule uninervis and Thalassia hemprichii were exposed to four different pCO2 treatments (442-1204 µatm) for 2 weeks, approximating the range of end-of-century emission scenarios. Photosynthetic responses were quantified using optode-based oxygen flux measurements. Across all three species, net productivity and energetic surplus (PG:R) significantly increased with a rise in pCO2 (linear models, P < 0.05). Photosynthesis-irradiance curve-derived photosynthetic parameters-maximum photosynthetic rates (Pmax) and efficiency (α)-also increased as pCO2 increased (linear models, P < 0.05). The response for productivity measures was similar across species, i.e. similar slopes in linear models. A decrease in compensation light requirement (Ec) with increasing pCO2 was evident in C. serrulata and H. uninervis, but not in T. hemprichii. Despite higher productivity with pCO2 enrichment, leaf growth rates in C. serrulata did not increase, while those in H. uninervis and T. hemprichii significantly increased with increasing pCO2 levels. While seagrasses can be carbon-limited and productivity can respond positively to CO2 enrichment, varying carbon allocation strategies amongst species suggest differential growth response between species. Thus, future increase in seawater CO2 concentration may lead to an overall increase in seagrass biomass and productivity, as well as community changes in seagrass meadows.
Jet milling effect on functionality, quality and in vitro digestibility of whole wheat flour and bread.
Protonotariou, S., Mandala, I. & Rosell, C. M. (2015). Food and Bioprocess Technology, 8(6), 1319-1329.
Jet milling is an ultragrinding process in order to produce superfine powders with increased functionalities. The effect of milling pressure, feed rate, vibration rate of feeder, and feedback of jet milling on whole wheat flour functionality and the potential of those flours for breadmaking with the goal of improving bread quality and digestibility were investigated. Increasing milling pressure (from 4 to 8 bar), decreasing feed rate (from 0.67 to 5.18 kg/h), and/or using recirculation augmented the severity of the process and reduced flour particle size from 84.15 to 17.02 µm. Breakage of aleurone particle layer and the reduction of particle size in jet milled flours were detected using scanning electron microscopy (SEM). Ash and protein content did not change after jet milling. However, total fiber content and digestible starch increased from 13.01 to 14.72 % and from 33.80 to 43.23 mg/100 mg, respectively, when subjected to jet milling at 8-bar air pressure. Mixolab ® data indicated that water absorption increased from 64.1 to 68.0 %, while pasting temperature decreased from 63.4 to 66.1°C owing to the milling intensity. Referring to bread, jet milled flour addition reduced the specific volume from 2.50 to 1.90 cm3/g, luminosity from 60.48 to 55.87, and moisture content from 35.78 to 33.49 %, and increased crumb hardness from 707 to 1808 g. Jet milled breads presented a slight decrease in estimated glycemic index (eGI) (from 86 to 81), suggesting that jet milling treatment could also have nutritional benefits.