Effects of replacing magnesium oxide with calcium-magnesium carbonate with or without sodium bicarbonate on ruminal fermentation and nutrient flow in vitro

The objective of this study was to evaluate the effects of replacing magnesium oxide (MgO) with calcium-magnesium carbonate [CaMg(CO₃)₂] on ruminal fermentation with or without the addition of sodium bicarbonate (NaHCO₃). Eight fermentors of a dual-flow continuous-culture system were distributed in a replicated (2) 4 × 4 Latin square design in a 2 × 2 factorial arrangement of treatments (magnesium sources × NaHCO₃). The treatments tested were 0.21% MgO [MgO; dry matter (DM) basis; 144.8 mEq of dietary cation-anion difference (DCAD)]; 0.21% MgO + 0.50% NaHCO₃ (MgO+NaHCO₃; DM basis; 205.6 mEq of DCAD); 1.00% CaMg(CO₃)₂ [CaMg(CO₃)₂; DM basis; 144.8 mEq of DCAD]; and 1.00% CaMg(CO₃)₂ + 0.50% NaHCO₃ [CaMg(CO₃)₂+NaHCO₃; DM basis; 205.6 mEq of DCAD]. Diets were formulated to have a total of 0.28% of Mg (DM basis). The experiment consisted of 40 d, which was divided into 4 periods of 10 d each, where 7 d were used for adaptation and 3 d for sampling to determine pH, volatile fatty acids (VFA), ammonia (NH₃-N), lactate, mineral solubility, N metabolism, and nutrient digestibility. The effects of Mg source [MgO vs. CaMg(CO₃)₂], NaHCO₃ (with vs. without), and the interaction were tested with the MIXED procedure of SAS version 9.4 (SAS Institute). There was no Mg source × NaHCO₃ interaction in the pH variables and mineral solubility, and Mg sources evaluated did not affect the variables related to ruminal pH and solubility of Mg. On the other hand, the inclusion of NaHCO₃ increased the pH daily average, independent of Mg source, which led to a reduced time that pH was below 5.8 and decreased area under the curve. Total VFA and lactate concentration were similar among treatments regardless of NaHCO₃ and Mg source; however, the molar proportion of isobutyrate and NH₃-N concentration were lower in diets with CaMg(CO₃)₂ compared with MgO. Moreover, NaHCO₃ inclusion increased NH₃-N, total daily NH₃-N flow, isobutyrate concentration, and acid detergent fiber digestibility. Our results showed that CaMg(CO₃)₂ leads to a lower NH₃-N concentration and isobutyrate proportion. Therefore, because most of the tested variables were not significantly different between MgO and CaMg(CO₃)₂ when combined or not with NaHCO₃, CaMg(CO₃)₂ can be a viable alternative source to replace MgO in dairy cow diets without affecting mineral solubility, ruminal pH, nutrient digestibility, total VFA, and the main ruminal VFA. Although Mg sources are known to have an alkalizing effect, NaHCO₃ inclusion in diets with Mg supplementation allowed an increase in ruminal pH, as well as an increase in isobutyrate and NH₃-N flow.     

Characterization of the Maillard reaction in bread crisps

Maillard reaction (MR) is one of the main chemical event occurring during baking. To study the reaction in bakery products, a dry model system is more useful than an aqueous one. In this work, the effects of formulation and processing conditions in a crisp bread system were investigated to test the effects of different additives on both the overall reaction and the formation of MR products such as 5-hydroxymethyl-2-furaldehyde (HMF) and acrylamide. Cylindrical dough made up of flour, water and yeast was baked at 180 °C for 35 min and the slices were toasted at different times/temperatures combinations. Browning and water content were monitored along with the kinetic of formation of chemical indicators such as HMF and acrylamide also calculating rate constants and activation energy. These parameters were also monitored in systems added with glycine and asparaginase. During toasting water content follows an exponential trend, being the rate of water loss faster in the initial stage of toasting and at higher temperature. Browning was more intense when toasting at higher temperature and a linear correlations between browning (ΔL*, ΔE*), HMF and acrylamide concentration were observed when toasting at 180 °C. HMF and acrylamide content increased with the toasting time and temperature. Their concentrations were strongly dependent on the water content of the final product, and both the addition of glycine and asparaginase are effective in reducing acrylamide content. The addition of glycine enhanced the browning of toasted bread, and slightly increased HMF content at any toasting temperature. The system characterized in this work represents a suitable tool to study the development of the MR in dry systems.     

Reduction of Acrylamide Formation in Sweet Bread with l-Asparaginase Treatment

Acrylamide, 2-propenamide, has the chemical formula $ \mathrm{CH}2=\mathrm{CH}-\mathrm{CO}-\mathrm{NH}2 $ . It is produced at elevated levels in high temperature fried and baked foods. It has adverse effects on human health and is proven to be neurotoxic, genotoxic, carcinogenic, and toxic to reproductive system. The aim of this paper was to reduce acrylamide formation in bakery products such as sweet bread by enzyme treatment. l-Asparaginase produced from Cladosporium sp. was treated to wheat-based dough at different concentrations (50–300 U). There was no change in the rheological properties of wheat flour and physico-sensory characteristics of bread with l-asparaginase treatment. Moisture, sugars, l-asparagine, acrylamide, and some indicators of Millard reaction (hydroxymethylfurfural (HMF), color, browning) were estimated. With increase in l-asparaginase level the acrylamide formation was reduced. At 300 U, there was 97 % and 73 % reduction of acrylamide formation in the crust and crumb regions of bread, respectively. HMF, a common intermediate product in the Maillard reaction and a genotoxic compound via 5-sulfoxymethylfurfural, also decreased in l-asparaginase-treated bread samples. These results indicated the potential of l-asparaginase enzyme for industrial and domestic applications in reducing harmful Maillard reaction compounds.     

Formulation of Culture Medium with Agroindustrial Waste for β-Galactosidase Production from <Emphasis Type="Italic">Kluyveromyces marxianus</Emphasis> ATCC 16045

Seven strains of the genus Kluyveromyces were screened for β-galactosidase activity and Kluyveromyces marxianus ATCC 16045 was selected as the best enzyme producer for culture medium optimization. The production of β-galactosidase by submerged cultivation was evaluated using a factorial design and response surface methodology. The culture medium containing whey and parboiled rice effluent was formulated to maximize the production of β-galactosidase. The effects of the initial pH and the concentrations of whey lactose, peptone, (NH₄)₂SO₄, yeast extract, and parboiled rice effluent on enzyme production were studied using a 2 _IV ^6-2 fractional design. A CCRD (24 trials plus axial and central points) was used for the four variables selected from the fractional design (lactose, peptone, (NH₄)₂SO₄ and yeast extract), with β-galactosidase activity as the response. The optimum conditions established for production were a whey (lactose) concentration of 120 g/L, a yeast extract concentration of 5 g/L, a peptone concentration of 15 g/L, a (NH₄)₂SO₄ concentration of 15 g/L, a parboiled rice effluent concentration of 30 g/L, and a pH value of 4.0. Under these conditions, the highest enzymatic activity of 10.4 U/mL was measured, being 9.5–9.7 as the values predicted by the proposed model, showing an enzymatic activity increase of 30% using alternative sources of lactose and nitrogen for β-galactosidase production.     

Rate-Controlling Mechanisms in the Photo-degradation of 5-Hydroxymethylfurfural

Rate-controlling mechanisms in the photo-degradation of 5-hydroxymethylfurfural (HMF) were studied applying kinetic and thermodynamic compensations. Aqueous solutions of HMF were prepared at a concentration of 100 mg L^?1 and at pH values of 3, 3.4, 4, and 5. The UV irradiation of samples was performed in an installation consisting of a black chamber containing the reactor and a mid-pressure mercury lamp with emission wavelengths between 250 and 740 nm. Every sample was irradiated at 12, 25, 35, and 45 °C for 120 min, analyzing their HMF content each 10 min. The photo-degradation data fitted well to zero-order kinetic model, and the constant values were used to study whether the kinetic and thermodynamic compensation could be applied. The isokinetic temperature was very similar for kinetic compensation (T_B?=?278.0 K) and thermodynamic compensation (T_B?=?277.8 K). Applying the Leffler’s criterion, the HMF photo-degradation was entropically controlled, probably as a consequence of hydrophobic interactions. In order to check the entropical control, two experiments were repeated at pH 3 but avoiding agitation. As the new obtained kinetic constants were highly different from the values previously obtained using agitation, it can be concluded that the HMF photo-degradation is an entropy-controlled process and can be speeded up by changing non-thermal parameters, like agitation.     

Enzymatic mitigation of acrylamide in fried potato chips using asparaginase from Aspergillus terreus

Acrylamide in foods is declared as carcinogen. In the present work, the effect of enzymatic pretreatment and other parameters like enzyme concentration, frying conditions with respect to temperature and time, size of potato chips, and effect of sodium chloride and citric acid on mitigation of acrylamide were studied. The concentration of acrylamide in fried potatoes after the pretreatment was found to be 815.63 μg kg^?1. The optimised asparaginase concentration for the mitigation of acrylamide was calibrated as 4 U mL^?1, and optimised frying time and temperature were 15 min and 170 °C, respectively. An in‐depth kinetic relationship for the effect of asparaginase on the mitigation of acrylamide was studied. The prime novelty of the project is focused on the immobilisation of asparaginase to nanomagnetic particles for redundant usage with stabilised enzyme activity. The work projected three stables cycles of asparaginase activity and on further usage of the immobilised enzyme resulted in decreased activity. The repeated use of immobilised asparaginase provides the advantage of decreasing cost in processing.     

Tolerance of acetate,propionate and sorbate by Saccharomyces cerevisiae and Torulopsis holmii

Several strains of Saccharomyces cerevisiae and one of Torulopsis holmii were investigated with regard to their tolerance of acetate, propionate and sorbate. The S. cerevisiae strains had poor acetic acid tolerance in sour dough pH 4·3, the dough raising power decreasing on average by 67% in the presence of 0·34% sodium acetate, while T. holmii was not affected by this concentration. Propionate (0·16%) and sorbate (0·08%) were more inhibitory than acetate, inhibiting the dough raising power even in normal dough at pH 5·9. Torulopsis holmii also had a good tolerance of propionate and sorbate and seems suitable for raising sour doughs.     

Effects of dough formula and baking conditions on acrylamide and hydroxymethylfurfural formation in cookies

The effects of dough formula and baking conditions on the formations of acrylamide and hydroxymethylfurfural (HMF) were studied in a cookie model system. Increasing the sugar concentration in the dough formula increased acrylamide formation during baking at 205 °C for 11 min. The effect of sugar on acrylamide formation was more pronounced for glucose than for sucrose, expectedly. Addition of citric acid into dough formula comprising sucrose increased the susceptibility of acrylamide formation, while it decreased acrylamide formation in the dough formula comprising glucose. Decreasing the pH of dough formula increased the tendency to surface browning and the formation of hydroxymethylfurfural in cookies during baking. The results suggest that a cookie with acceptable texture and colour, but having less than 150 ng/g of acrylamide, can be manufactured by lowering the baking temperature and avoiding reducing sugars in the recipe.     

Quantification of 5-Hydroxymethylfurfural in Coated Deep-Fried Products: Optimization of the Extraction Procedure by Using Statistical Design

This study describes for the first time the development and validation of an extraction procedure for the quantification of 5-hydroxymethylfurfural (HMF) in coated deep-fried products by liquid chromatography with photodiode array detection. The method entailed the extraction of HMF with ethyl acetate/hexane (4:1) followed by a concentration step with 40 mM sodium formate (pH?=?3)/methanol (1:1). The optimum combination of the extraction variables was achieved by response surface methodology. Sample amount and concentration solvent volume showed a notable influence on HMF yield, while the effect of extraction solvent volume seemed to be less marked. From experimental results, 5 g of sample, 10 ml of the extraction solvent, and 550 μl of the concentration solvent were selected as optimal combination. The consistency between predicted and experimented values as well as in the quality parameters was observed. Quantities of HMF in coated deep-fried fish products were 1.25?±?0.21 μg g^?1.     

A novel potentiometric pH electrode based on sulfated natural Fe<Subscript>3</Subscript>O<Subscript>4</Subscript> and analytical application in food samples

A novel solid state potentiometric pH electrode based on sulfated natural Fe₃O₄ silicone was fabricated. The optimum potentiometric performances such as Nernstian response, response time, selectivity, life-time and reproducibility of pH electrodes were investigated by using a computer-controlled potentiometric device. Moreover, the potentiometric performance of the solid state pH electrodes was studied with different mixtures sulfated natural Fe₃O₄, silicone and graphite powder. The best potentiometric behavior of proposed pH electrode was obtained with a composition of 20% (w/w) sulfated natural Fe₃O₄, 40% (w/w) graphite powder and 40% (w/w) silicone. The sub-Nernstian response for pH electrode was exhibited with a slope of 30.8?±?1.4 mV/pH (r?=?0.9963) from pH 2 to pH 12. In addition, the dynamic response time was found as 10 s in acidic medium and further the proposed pH electrode can be used for at least 1 year without any significant slope of the pH–potential curve. The selectivity coefficient of pH electrode was interpreted according to fixed interference method in the presences of Na⁺, Li⁺, K⁺, NH₄⁺, SO₄^2?, CH₃COO^? and NO₃^? ions. The reproducibility of pH electrode was calculated in pH 4 and pH 6 phosphate buffers and it was found as 0.24 RSD (%) and 0.27 RSD (%) respectively. The proposed pH electrode was used to determine of pH in acid?bases titration compared with glass pH electrode and is highly stable in corrosive systems including HF solution. Terminally, the pH value was successfully determined in some soft drinks and milk samples by proposed solid state pH electrode at 95% confidence level with satisfactory agreement compared with glass pH electrode.     

Acrylamide formation and antioxidant level in biscuits related to recipe and baking

Heated plant foods may contain compounds with adverse health effects (e.g. acrylamide). On the other hand, health-promoting compounds (e.g. antioxidants) have also been identified in such foods. Therefore, a baking experiment with biscuits was carried out to study the potential impact of both acrylamide and antioxidants in that food. Two different wheat flour types – wholemeal (WMF) and white flour type 550 (T550; 0.55% mineral content) – as well as recipe (fat and leavening agent) and thermal input (temperature?×?time) were changed. Furthermore, the effect of an enzymatic asparagine hydrolysation was tested. Antioxidants were determined with two independent procedures ABTS – (2,2′-azinobis-(3-ethylbenzothiazoline-6-sulfonate)) and FRAP-assay (ferric reducing ability of plasma). WMF samples resulted in an unchanged acrylamide level, but in a significantly higher antioxidant concentration when compared with T550 samples (149 and 141?µg?kg^?1 acrylamide and 9.1 and 5.1?mmol?TE?kg^?1 FW ABTS for WMF and T550, respectively). A reduced fat content yielded in an increased volume. Raising agents had no effect on acrylamide levels, but antioxidants were higher in samples with sodium bicarbonate (SBC) than with ammonium bicarbonate (ABC). Thermal input (temperature?×?time; 150°C?×?25?min to 240°C?×?9?min) indicated an exponential acrylamide increase especially at higher temperatures (from 75 to 236?µg?kg^?1), whereas antioxidant increase was linear (from 7.0 to 7.7?mmol?TE?kg^?1 FW, ABTS). FRAP and ABTS values were correlated on a low level, whereas acrylamide content of biscuits was correlated with FRAP and lightness (R ^2?=?0.62 and 0.47, and 0.71 and 0.85 for WMF and T550, respectively). The enzyme asparaginase reduced acrylamide formation by about 50% for both raising agents (SBC and ABC, respectively), whereas antioxidant levels were not affected. An evaluation of recipe variants with low acrylamide and high antioxidants indicated the advantage of wholemeal biscuits.     

Mode of action of yeast culture (YEA-SACC 1026) for stimulation of rumen fermentation in buffalo calves

The effect of daily supplementation of 5 g Saccharomyces cerevisiae yeast culture (YC, YEA-SACC 1026), 30 g NaHCO₃, supernatant from 5 g YC (YCS), 5 g autoclaved YC (YCH) or 5 g γ-irradiated YC (YCR) to the diet of buffalo calves on rumen microbial populations and fermentation pattern was examined. Addition of 30 g NaHCO₃ increased the rumen pH to the level observed with YC group. The pH and the concentrations of total, total viable and cellulolytic bacteria and total volatile fatty acids (VFA) were significantly higher while that of lactic acid, hexose-unit oligosaccharides and NH₃-N were significantly lower in the rumen fluid of YC compared with the control group. The effect of NaHCO₃ was 39·5 and 59·5% in decreasing the concentrations of lactic acid and hexose-unit oligosaccharides, 48·1, 47·2 and 45·5% in increasing the numbers of total, total viable and cellulolytic bacteria, 50·0 and 58·1% in increasing the concentrations of total VFA and protein and 51·3% in decreasing the concentration of NH₃-N of YC. The corresponding values for YCR addition in the diet were 38·6, 45·7, 48·5, 44·4, 51·5, 39·1, 48·1 and 46·5%. The effect of YCS and YCH was only marginal, but conspicuous up to 2 h after feeding, in changing the above rumen variables when compared with the YC group. The results indicated that contribution of increase in pH in changing the rumen variables was approximately 50% of YC and almost all the stimulatory activity was associated with live yeast cells. Autoclaving of YC destroyed almost all and γ-irradiation of YC retained about 50% of stimulatory activity of YC. The effect of YC on rumen fermentation, which was maximum up to 2 to 4 h after feeding, decreased with time. © 1998 SCI.     

Immobilized alcalase alkaline protease on the magnetic chitosan nanoparticles used for soy protein isolate hydrolysis

An efficient immobilization of Alcalase 2.4L alkaline protease has been developed by using chitosan-coated magnetic nanoparticles as support via glutaraldehyde cross-linking reaction. The Fe₃O₄ nanoparticles, Fe₃O₄-chitosan, and immobilized Alcalase 2.4L alkaline protease were characterized by X-ray diffraction, transmission electron microscope, Fourier transform infrared spectroscopy, electron spin resonance, and vibrating sample magnetometry. Results showed that the binding of chitosan and Alcalase 2.4L alkaline protease on Fe₃O₄ through cross-linking was successful. In addition, the Alcalase 2.4L alkaline protease immobilized with chitosan-coated magnetic nanoparticles enhanced the activity, the optimum reaction temperature and pH value for the immobilized enzyme were 55 °C and 10, respectively, compared with the free enzyme, and the optimal temperature and pH profile range were considerably broadened. Similarly, the thermal stability was enhanced by immobilization, and the kinetic parameters of free and immobilized Alcalase 2.4L alkaline protease were determined. Then, from our hydrolysis experiments, we found that immobilized Alcalase 2.4L alkaline protease uses Fe₃O₄-chitosan had a greatest hydrolytic activity, and the DH of soy protein isolate (SPI) can reach to 18.38 %, against 17.50 % with the free enzyme after 140 min. Furthermore, the immobilized Alcalase 2.4L alkaline protease could maintain about 86 % of its original activity after ten consecutive operations. Thus, Fe₃O₄-chitosan immobilized Alcalase 2.4L alkaline protease a good candidate for the continuous hydrolysis of SPI.     

Acrylamide, 5-hydroxymethylfurfural and Nε-carboxymethyl-lysine in coffee substitutes and instant coffees

Sensitive analytical methods were developed and validated for the quantification of acrylamide, N^ε-carboxymethyl-lysine (CML) and 5-hydroxymethylfurfural (HMF) in 24 commercial coffee substitutes (CSs) and 12 instant coffees (ICs). Acrylamide levels varied widely from 200 to 4940 µg kg^–1 with higher levels in CSs. Only two out of 24 CSs had a level of acrylamide above the indicative value set for this food category by the European Commission (4000 µg kg^–1). None of the ICs tested in this study exceeded the indicative value set for this foodstuff (900 µg kg^–1). CML ranged from 0.17 to 47 mg kg^–1 and it increased in proportion to the protein content of the samples. The highest concentrations were found in IC partly due to the relatively high protein content of this food group. HMF was the most abundant neoformed compound (NFC) found in the tested commercial samples. It was found between 0.59 and 13 g kg^–1. Among other food categories IC and CS could appear to be major contributors to the exposure to NFCs if consumed on a daily basis. Further investigations are needed to elucidate the acrylamide formation during processing and to determine the daily intake level of frequent consumers of these products.     

EFFECT OF SODIUM METABISULFITE ADDITION AND BAKING TEMPERATURE ON MAILLARD REACTION IN BREAD

ABSTRACT

In this study, the effect of sodium metabisulfite (SMBS) doses (0, 25, 50, 100 mg/kg dough) and baking temperatures (200, 230 and 250C) on the physical, chemical and sensory properties of bread were researched to reduce 5‐hydroxymethyl‐2‐furfural (HMF) and acrylamide contents. HMF and acrylamide contents of bread crust were decreased significantly by increasing SMBS dose and decreasing baking temperature. The HMF (137.29 mg/kg) and acrylamide (671.44 µg/kg) contents of bread crust were decreased by 33 and 67%, respectively by addition of 100 mg/kg SMBS. The maltol content of bread crusts were significantly affected by baking temperature, and were 7.19, 10.23 and 22.69 mg/kg in breads baked at 200, 230 and 250C, respectively. No HMF, acrylamide and maltol were detected in the bread crumb. The sulfur dioxide content of the crust and crumb of control bread was 6.99 and 10.69 mg/kg, and increased by 49 and 59%, respectively at 100 mg/kg SMBS dose. All breads were evaluated as acceptable by a sensory panel.

PRACTICAL APPLICATIONS

Since Maillard reaction products such as acrylamide and 5‐hydroxymethyl‐2‐furfural (HMF) are known as toxic compounds, mitigation of these compounds is important subject for health and nutrition. There is not an efficient method to prevent the formation of acrylamide and HMF in bread crusts comparison with potato crisps. The purpose of this research is to slow down Maillard reaction by addition of sodium metabisulfite in bread‐making process. As a result of this research, acrylamide and HMF content of bread crusts decreased by 33 and 67%, respectively with acceptable sensory evaluation.     

Occurrence of patulin and 5-hydroxymethylfurfural in apple sour,which is a traditional product of Kastamonu,Turkey

Apple sour is a traditional product of Kastamonu, Turkey. It is consumed by spreading on bread or drinking after diluting with water. The aim of this study was to determine patulin (PAT) and 5-hydroxymethylfurfural (HMF) in apple sour. This study is the first to evaluate the occurrence of PAT and HMF in apple sour. The samples were extracted with ethyl acetate using liquid-liquid extraction technique. PAT and HMF were determined by HPLC with UV detection. PAT was detected in all samples, and the PAT level in 94.9% of samples was found to be equal or greater than the legal limit for juice concentrates. The mean value for PAT was found to be 284 ± 307 μg kg^?1. PAT levels in 13 of 39 samples were in the range of 100 ≤ x < 200 μg kg^?1, two samples were in the range of 0 ≤ x < 50 μg kg ?1 and two samples were in the range of 1000 ≤ x < 1500 μg kg¹. HMF levels of all samples were above the legal limit for solid molasses. The mean value for HMF was found to be 16215 ± 13317 mg kg^?1. HMF levels of 10 of 39 samples were determined to be in the range of 10000 ≤ x < 20000 mg kg^?1, eight samples were in the range of 20000 ≤ x < 30000 mg kg^?1 and only three samples were in the range of 100 ≤ x < 1000 mg kg^?1. There was a significant and inverse relationship between HMF and pH of the samples. These results indicate that consumption of apple sour is a considerable risk in terms of HMF and PAT toxicity.     

Purification and characterisation of thermo-alkaline pectinase enzyme from Hylocereus polyrhizus

The thermo-alkaline pectinase enzyme from Hylocereus polyrhizus was purified 232.3-fold with a 73.3 % recovery through ammonium sulphate precipitation, gel filtration, and ion exchange chromatography. Ion exchange chromatography combined with sodium dodecyl sulphate gel electrophoresis (SDS-PAGE) revealed that the enzyme was monomeric with a molecular weight of 34.2 kDa. The pectinase exhibited broad specificity towards polygalacturonic acid, arabinan, oat spelt xylan, and pNP-α-glucopyranoside. The optimum pH and temperature were 8.0 and 75 °C, respectively. This enzyme was stable over a wide pH range (3.0–11.0) and at relatively high temperature (85 °C for 1 h). The K_m and V_max values of pectinase towards polygalacturonic acid were 2.7 mg/ml and 34.30 U/mg proteins, respectively. In addition, the enzyme activity was inhibited by Ni²⁺, Al³⁺, and Fe²⁺ and was increased in the presence of Ca²⁺ and Mg²⁺ by 120 and 112 %, respectively. The purified pectinase demonstrated robust stability in response to surfactants and oxidising agents. EDTA, which is a powerful chelating agent, did not exert any significant effect on the enzyme stability. Thus, enzymes with these unique properties may be widely used in different types of industries and biotechnological applications.     

Purification and characterisation of basic ascorbate oxidase from satsuma mandarin (Citrus unshiu Marc)

Basic ascorbate oxidase of the multiple enzyme forms existing in young fruit of satsuma mandarin (Citrus unshiu Marc) has been separated and subsequently purified to electrophoretic homogeneity through (NH₄)₂SO₄ fractionation and chromatographies on DEAE-Toyopearl 650M, CM-Sephadex C-50 and Sephadex G-100. The native molecular weight was estimated to be 141 kDa by gel filtration and composed of two non-identical subunits with an apparent mass of 74 kDa and 62 kDa. The optimum pH was found to be 5.5 with reasonable stability between pH 5 and 8. The enzyme had an optimum temperature at 45°C and was stable up to 50°C upon heat treatment for 5 min. The presence of sodium diethyldithiocarbamate, metabisulphite and potassium cyanide completely inhibited the enzyme activity. Fluoride also inhibited the activity substantially at higher concentrations. Other tnonovalent and divalent metal ions did not have inhibitory effects.     

Purification, characterization, and production of β-mannanase from Bacillus subtilis TJ-102 and its application in gluco-mannooligosaccharides preparation

A novel β-mannanase-producing strain, Bacillus subtilis TJ-102, was identified and characterized. Response surface method was applied to improving and enhancing the enzyme production. The optimized media components were obtained: 45.25 g/L konjac, 9.29 g/L Na₂HPO₄·12H₂O, 2.60 g/L CaCO₃, 1.0 g/L (NH₄)₂SO₄, 0.3 g/L KH₂PO₄, 1.0 g/L NaCl, 1.0 g/L MgCl₂·6H₂O, and 0.01 g/L FeSO₄. Under these conditions, the β-mannanase activity could achieve 205.3 U/mL in a 7-L fermentor. Then, β-mannanase was 7.39-fold purified by salting out, ultrafiltration, anion-exchange, and size-exclusion preparative chromatography with a recovery of 21.41 % and a specificity of 125.36 U/mg proteins. β-Mannanase was stable below 65 °C and pH 5.0–8.0, which exhibited excellently enzymatic efficiency in the preparation of gluco-mannooligosaccharides (GMOS) by hydrolyzing konjac flour. The GMOS yield of 57.76 % has been achieved with 8.71 % of mannose and 14.49 % of glucose, demonstrating the potential use of β-mannanase in food industry.     

Effects of Feeding Synthetic Zeolite A and Sodium Bicarbonate on Milk Production Nutrient Digestion,and Rate of Digesta Passage in Dairy Cows

Four rumen-cannulated Holstein cows were fed synthetic zeolite A and NaHCO₃ to evaluate their affect on milk production, nutrient digestibility, rumen fermentation, and rate of digesta passage. Treatments were allocated in a 2 × 2 factorial arrangement within a 4 × 4 Latin-square design. Treatments consisted of control; 1.0% NaHCO₃; 2.0% zeolite; and 1.0% NaHCO₃ plus 2.0% zeolite. A total mixed ration with 50:50 concentrate to forage (80% corn silage, 20% haylage) DM was fed.Intake of DM was lower for cows receiving zeolite (18.7 vs. 20.7 kg/d). Decreases were noted in daily milk (26.3 vs. 28.9 kg/d). 4% FCM (23.6 vs. 25.6 kg/d); milk fat yield (.86 vs. .93 kg/d); milk protein yield (.85 vs. .95 kg/d); and milk protein percent (3.21 vs. 3.34) with zeolite. Digestibilities of DM, organic matter, and crude protein were also decreased by zeolite but ADF digestion was unaffected. Rumen pH was increased, ruminal propionate decreased, and acetate:propionate ratio increased by zeolite. All other VFA plus rumen NH₃ were not affected by treatment. Decreases due to zeolite were observed in liquid fractional rate of passage and liquid flow rate when measured by Cr-EDTA in the feces. No treatment differences were found in fractional rate of passage of feed particles. Addition of NaHCO₃ had no significant effects.