Identification of discrepancies in grain quality and grain protein composition through avenin proteins of oat after an effort to increase protein content

Background: Oat prolamin (avenin) is a family of proteins that contain several polymorphic components. The highinterspecific variability of avenin among cultivars, electrophoretic patterns have been used for grain protein comparisonand species relationships. Therefore, the present study was carried out to evaluate the effect of environment onavenin constitution following an effort to increase protein content of oat using different approaches.Results: The avenin electrophoretic patterns of 25 samples resulting from 20 oat cultivars and 5 mutated oat linesgrown or developed in different fields and greenhouses were compared. Quantification of total proteins and SDSPAGEanalysis of avenin protein extracts were carried out. Different pattern of avenin allele in electrophoretic gelallowed the comparison within and between groups. This analysis shows that the cultivars obtained from Swedishfields had differences in the number of alleles compared with the parental cultivar, so were not stable across sites. Thestudy revealed that oats treated with nitrogen and the selected mutated lines showed increase in protein concentration,with consistent avenin loci to the parental line, so these lines are now being used as potential candidates fordeveloping high-protein oat lines in the future.Conclusions: Our results provide useful information, on the stability of modifications, for breeders aiming to increasingthe content and nutritional value of oat protein. The study will increase our knowledge in the stability of differentmodifications and thus to accordingly select oat lines with stable and improved nutritional values.Keywords: Avenin, Mutation, Nitrogen, Geographical location, Electrophoresi

Review of analytical methods for measurement of oat proteins : The need for standardized methods

This review summarizes the analytical methods that have been developed for quantification and characterization of oat proteins. These include sampling, sample preparation, extraction, quantification, separation, detection, and characterization of oat proteins. The review also provides a comparison of different methods for the determination of protein fraction of oat and the efficiency thereof. We conclude that there is a need for further validation of existing data or methods and for a standard methodology to quantify oat proteins

Heat-treated high-fat diet modifies gut microbiota and metabolic markers in apoe-/- mice

BACKGROUND: High-fat diet has been known to have adverse effects on metabolic markers, as well as the gut microbiota. However, the effect of heat processing of high-fat diet, which leads to formations of advanced glycation end products (AGEs) has not been clearly distinguished from the effect of unheated fat. This study compared the effect of high-fat diet with heat-treated high-fat diet on adiposity, atherosclerosis and gut microbiota composition in the caecum of apoe (-/-) mice.METHOD: Male apoe (-/-) mice were fed either low-fat (LF) control diet, high-fat (40 E% saturated fat, HF) control diet, or heat-treated high-fat (200 °C for 10 min, HT) diet, for 8 weeks. The plasma samples were used in the analysis of Nε-carboxy-methyl-lysine (CML) and Nε-carboxy-ethyl-lysine (CEL). The heart samples were analysed for atherosclerotic plaques, and the DNA from caecum was extracted and analysed for microbiota composition using 16S rRNA gene sequencing on a Miseq instrument. Additionally, the functions of microbial communities were also predicted based on the bacterial 16S rRNA gene sequence using Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt).RESULTS: Here we found that HT modifies gut microbiota composition and host adiposity. Prediction of bacterial gene functions based on 16S rRNA gene sequence revealed that HF increased bacterial genera enriched in lipid metabolism genes, while HT did not. Plasma CML and CEL increased 1.7 and 2.5 times, respectively, in mice fed HT as compared to mice fed HF. Despite lower adiposity, mice fed HT maintained atherosclerosis and displayed enlarged spleens.CONCLUSIONS: The results suggested that heat processing of high-fat diet modifies the substrates reaching the lower gut of apoe (-/-) mice, resulting in different effects on gut microbiota composition. AGEs seem to maintain the effect on atherosclerosis, despite lower adiposity, and causing enlarged spleens, which possibly reflect elevated levels of inflammation in the body

On the Importance of Processing Conditions for the Nutritional Characteristics of Homogenized Composite Meals Intended for Infants

The nutritional quality of infant food is an important consideration in the effort to prevent a further increase in the rate of childhood obesity. We hypothesized that the canning of composite infant meals would lead to elevated contents of carboxymethyl-lysine (CML) and favor high glycemic and insulinemic responses compared with milder heat treatment conditions. We have compared composite infant pasta Bolognese meals that were either conventionally canned (CANPBol), or prepared by microwave cooking (MWPBol). A meal where the pasta and Bolognese sauce were separate during microwave cooking (MWP_CANBol) was also included. The infant meals were tested at breakfast in healthy adults using white wheat bread (WWB) as reference. A standardized lunch meal was served at 240 min and blood was collected from fasting to 360 min after breakfast. The 2-h glucose response (iAUC) was lower following the test meals than with WWB. The insulin response was lower after the MWP_CANBol (−47%, p = 0.0000) but markedly higher after CANPBol (+40%, p = 0.0019), compared with WWB. A combined measure of the glucose and insulin responses (ISIcomposite) revealed that MWP_CANBol resulted in 94% better insulin sensitivity than CANPBol. Additionally, the separate processing of the meal components in MWP_CANBol resulted in 39% lower CML levels than the CANPBol. It was therefore concluded that intake of commercially canned composite infant meals leads to reduced postprandial insulin sensitivity and increased exposure to oxidative stress promoting agents

Application of a dye-binding method for the determination of available lysine in skim milk powders.

A dye-binding method using Acid Orange 12 was investigated regarding its suitability for the quantification of available lysine, as a means of monitoring the Maillard reaction in skim milk powders. The method was evaluated by analyzing a wide range of milk powders produced by three different drying methods and stored under various conditions. A pilot-scale freeze-dryer, spray-dryer and drum-dryer were used to produce skim milk powders and the samples were stored at two temperatures (20°C and 30°C) and two relative humidities (33% and 52%) under strictly controlled conditions. Moreover to validate the method, two protein isolates; bovine serum albumin and casein were investigated for their available lysine content. The results demonstrate the suitability of this method for measuring the available lysine in skim milk powders with good precision and high reproducibility. The relative standard deviations obtained from the 125 freeze-dried powders were 1.8%, and those from the 100 drum-dried samples were all 1.9%. The highest variation was found for the spray-dried powders, which showed relative standard deviations between 0.9% and 6.7%

The Impact of Different Drying Techniques and Controlled Storage on the Development of Advanced Glycation End Products in Skim Milk Powders Using Isotope Dilution ESI-LC-MS/MS

The impact of different drying techniques and storage conditions on the formation of carboxymethyl lysine (CML) as an advanced glycation end product (AGE) was investigated in various pilot-scale produced skim milk powders (SMP). CML, an indicator of advanced stages of the Maillard reaction, was analyzed by isotope dilution ESI-LC-MS/MS after drying and at intervals during storage for 200 days. The aim of this study is to increase the awareness that the development of AGEs can be hindered by controlled storage, which is mostly neglected at the domestic level by the consumers. Pilot-scale freeze-dryer, spray-dryer, and drum-dryer were utilized for manufacturing SMPs. The storage was designed in such a way that it simulated how an average consumer keeps the dried products at home, specifically after opening the packages and how the products may be exposed to unfavorable conditions. To do so, four storage conditions including two temperatures (20 °C, 30 °C) and two relative humidities RH (33%, 52%) were studied and CML formation was monitored using LC-MS/MS. Our results show that the spray-dried samples developed two times higher CML after 200 days, compared to the freeze-dried samples, and this value was 1.6 times for the drum-dried samples. Storage at 52% RH developed 9.8–10 times more CML in the samples, in comparison with 33% RH, at the same storage temperature (P < 0.05). Our findings demonstrate that AGEs, molecules suspected to be involved in risks factors for auto-immune diseases, may be formed during storage of SMPs after opening the packages by the consumers

Early and advanced stages of Maillard reaction in infant formulas : Analysis of available lysine and carboxymethyl-lysine

Although the literature on the Maillard reaction in infant formulas is extensive, most studies have focused on model systems, and in only a few cases on real food systems. Therefore, the objective of the present study was to determine the status of the Maillard reaction, both the early and advanced phases, in a variety of commercial infant formulas available on the Swedish market. Ten powder and liquid milk-based infant formulas from three manufacturers were selected to determine available lysine and CML contents, the two established indicators of the reaction. The products were also characterized with respect to protein content, carbohydrates composition, water content and water activity. In order to be able to compare the impact of different processing steps applied on powder and liquid formulas, the solid formulas contained similar ingredients as their corresponding liquid ones. Our findings showed that powder and liquid formulas contained similar available lysine concentrations regardless of the manufacturer, showing 27.14–36.57% decrease in the available lysine, compared to the reference skim milk powder in this study. The CML concentrations were in a broad range of 68.77–507.99 mg/kg protein. In the case of one manufacturer, liquid infant formulas had significantly higher CML content, compared to the powder products (p < 0.05). The results from this study are a step taken towards better understanding of the extent of the Maillard reaction in real complex systems of infant formulas

Isotope dilution ESI-LC-MS/MS for quantification of free and total Nε-(1-Carboxymethyl)-l-Lysine and free Nε-(1-Carboxyethyl)-l-Lysine: Comparison of total Nε-(1-Carboxymethyl)-l-Lysine levels measured with new method to ELISA assay in gruel samples.

ESI-LC-MS/MS method with isotope dilution and SPE based on cation-exchange was developed for determination of free and total Nε-(1-Carboxymethyl)-l-Lysine (CML) and free Nε-(1-Carboxyethyl)-l-Lysine (CEL). The use of nonafluoropentanoic acid in mobile phase was omitted, SPE recoveries of 82±3% and 91±10% (n=6) for CML and CEL respectively and, calibration curves (R(2)>0.9985) were attained. The method was applied to gruel samples and LoQ for the method was 5ng/ml, RSD <10% and accuracy was 115%. Total CML levels in the gruel samples varied from 103-408mg/kg protein. Free CML levels which were 1000 times lower than total CML were three times higher than free CEL levels. CML in a gruel sample was 127±7, 84±9 and 253±28mg/kg using the current ESI-LC-MS/MS, ELISA and GC-MS respectively. The described method has advantages over ELISA with respect to reproducibility and specificity and over GC-MS with respect to reproducibility

Additional file 2: Figure S2. of Heat-treated high-fat diet modifies gut microbiota and metabolic markers in apoe−/− mice

Cladogram plot showing bacterial taxa with LDA scores higher than two using strict version of LEfSe. Different colors represent the most abundant taxa in different groups of mice (yellow indicating LF, red indicating HF and blue indicating HT). Circles represent phylogenetic levels from phylum to genus. The sizes of the circle are proportional to the taxon’s abundance. (PDF 436 kb