The Inhibitory Effects of Brans and Their Aqueous Extracts on the Lipolysis of Tributyrin Catalyzed by Calf Pregastric Lipase

The ability of red and white wheat brans and oat bran to inhibit calf pregastric lipase (CPGL)‐catalyzed hydrolysis of tributyrin has been studied in vitro at pH 6.5, 37 °C. Brans that had been presoaked in Bis‐Tris buffer at pH 6.5, 4 °C reduced CPGL activity by 13% to 20% after being soaked for 0.25 h and by 17% to 30% after being soaked for 24 h. Maximum reduction of CPGL activity was seen within 2 to 5 min incubation between coarse red wheat bran and CPGL. The results are discussed in terms of likely inhibitory components within the brans and their water‐soluble components and their roles in inhibiting pregastric and other lipase‐catalyzed hydrolysis of dietary lipids.     

Structural Characteristics of Pumpkin Pectin Extracted by Microwave Heating

Abstract: To improve extraction yield of pumpkin pectin, microwave heating was adopted in this study. Using hot acid extraction, pumpkin pectin yield decreased from 5.7% to 1.0% as pH increased from pH 1.0 to 2.0. At pH 2.5, no pectin was recovered from pumpkin flesh powder. After a pretreatment at pH 1.0 and 25 °C for 1 h, pumpkin powder was microwave‐extracted at 120 °C for 3 min resulting in 10.5% of pectin yield. However, premicrowave treatment at 60 °C for 20 min did not improve extraction yield. When microwave heating at 80 °C for 10 min was applied after premicrowave treatment, final pectin yield increased to 11.3%. When pH was adjusted to 2.0, the yield dropped to 7.7% under the same extraction conditions. Molecular shape and properties as well as chemical composition of pumpkin pectin were significantly affected depending on extraction methods. Galacturonic acid content (51% to 58%) of pumpkin pectin was lower than that detected in commercial acid‐extracted citrus pectin, while higher content of neutral sugars and acetyl esters existed in pumpkin pectin structure. Molecular weight (M_w) and intrinsic viscosity (η_w) determined for microwave‐extracted pumpkin pectins were substantially lower than acid‐extracted pectin, whereas polydispersity was greater. However, microwave‐extracted pectin at pH 2.0 had more than 5 times greater M_w than did the pectin extracted at pH 1.0. The η_w of microwave‐extracted pectin produced at pH 2.0 was almost twice that of other microwave‐extracted pectins, which were comparable to that of acid‐extracted pectin. These results indicate that extraction yield of pumpkin pectin would be improved by microwave extraction and different pectin structure and properties can be obtained compared to acid extraction. Practical Application: Pumpkin is a promising alternative source for pectin material. Pumpkin pectin has a unique chemical structure and physical properties, presumably providing different functional properties compared to conventional commercial pectin sources. Depending on the conditions to produce pumpkin pectin, diverse molecular structures can be obtained and utilized in various food applications.     

Factors Influencing the Heat Resistance of a Heat-Resistant Lipase of Pseudomonas

Pseudomonas spp. MC50 produced an extra-cellular lipase that was extremely heat resistant at 100–150°C when heated in water or emulsions. The D-values ranged from 40 min at 100°C to 84 sec at 150°C. The z_D-vallue was 36°C. When heated in water, the lipase exhibited greatest survival at pH 8.5. Below pH 6.5, survival was less than 10% of that at pH 8.5. Survival of lipase heated in emulsions was affected somewhat by the type of oil, corn oil concentration, and type of emulsifier. Type of stabilizer or stabilizer concentration had little effect on lipase survival. Lipase treated with ethylenediaminetetraacetate (EDTA) was fully heat resistant in water; survival declined when lipase was heated in CaCl₂ solutions of 3–120 mM.     

Enzymatic Production of Ribonucleotides from Autolysates of Kluyveromyces marxianus Grown on Whey

After 20h fermentation of medium containing 5% (w/v) dehydrated whey, at 30°C, pH 4.5, yeast cells were harvested, diluted in 0.1M KH₂PO₄, and autolyzed at different pHs (6.5–7.5) and temperatures (45–55°C). Phosphodiesterase (0.2–1.0% w/v, 65°C, pH 6.5, 6h) and adenyl deaminase (0.5-1.0% w/v, 60°C, pH 5.5, 4h) were added to the autolysates. After heat treatment (100°C, 15 min), samples were analyzed by RP-HPLC and LC/MS. Production of 5′-ribonucleotides was maximized at 50°C, pH 6.5. Yields of 5′-AMP (800 μg/g of biomass) and 5′-GMP (2000 μg/g) increased considerably after addition of 1.0% phosphodiesterase. 5′-IMP increased only after addition of 1.0% adenyl deaminase.     

Kinetic behaviour and thermal inactivation of pectinlyase used in food processing

Kinetic properties and thermal inactivation of pectinlyase (PL) were assayed in commercial pectinase preparations (Rapidase C80, Pectinase CCM, Pectinex 3XL and Grindamyl 3PA) by using apple pectin as substrate. The PL activity of Rapidase C80 showed substrate inhibition, while the other enzyme preparations followed typical Michaelis–Menten kinetics. The optimum pH and temperature values for PL activity lay within the range of 5.5–6.5 and 35–40 °C, respectively. PL was heat‐inactivated with simple first order kinetics. With respect to this, thermodynamic activation parameters (ΔH^#, ΔS^# and ΔG^#) using a non‐linear Arrhenius plot were calculated. The Pectinase CCM and Pectinex 3XL PL showed a half‐life (t_1/2) at 50 °C of 2.0 min and 11.8 min, respectively.     

Effect of gelation factors on the formation and characteristics of protease‐induced whey protein gel

Whey protein gel formed at 10% (w/v) whey protein concentration, 0.5% E/S, pH 7.0, 55°C and 2.5 mM CaCl₂ concentration had an average particle size of 23.46 μm, hardness of 0.46, cohesiveness of 0.13 and adhesiveness of 1.40, and the gel showed semisolid, smooth and creamy texture. There were no distinct changes in gel textural properties after heating at 80 and 90°C for 5 min, respectively, or being kept at 4°C for 1 month. The textural properties of the gel showed no significant difference after its pH was adjusted to 4.5, 5.5 and 7.5 compared with that of pH 6.5 (control gel). However, the average particle size significantly increased after being adjusted to pH 4.5 and pH 5.5. Transmission electron micrographs showed that protease‐induced gel possessed much looser aggregate structure compared with heat‐induced compact gel, which may give support to its potential application in low‐fat foods that no need of extensive heating.     

Carbon Dioxide Inhibition of Escherichia coli and Staphylococcus aureus on a pH-adjusted Surface in a Model System

Growth of Escherichia coli and Staphylococcus aureuson the surface of Trypticase Soy Agar (TSA) packaged with various CO₂ partial pressures (0, 20, 40, 60, 80, 100%, balance N₂) was compared to the control (N₂ 100%) on TSA in which the pH was adjusted to equal that in CO₂ atmospheres at 15°C and 30°C. At 15°C, the biostatic effect was noted with all CO₂ partial pressures for both species. At 30°C, the biostatic effect of CO₂ was almost completely nullified for E. coli, but that for S. aureus was still effective. S. aureus was more sensitive to the inhibitory effects of CO₂ than E. coli at both the temperatures.     

Curtailing Oxidation‐Induced Loss of Myosin Gelling Potential by Pyrophosphate Through Shielding the S1 Subfragment

In muscle food processing, where oxidation is inevitable, phosphates are usually added to improve water binding. This present study attempted to investigate the interactive roles of protein oxidation and pyrophosphate (PP) during thermal gelation of myosin. Myosin isolated from pork muscle was solubilized in 0.5 M NaCl at pH 6.2 then oxidatively stressed with an iron‐redox cycling system that produces hydroxyl radicals with or without 1 mM PP and 2 mM MgCl₂ at 4 °C for 12 or 24 h then heated to 50 °C at 1.3 °C/min. Protein conformational stability was measured by differential scanning calorimetry, and covalent cross‐linking was examined by sodium dodecyl sulfate–polyacrylamide gel electrophoresis following chymotrypsin digestion. The binding of PP to myosin suppressed disulfide bond formation in myosin subfragments 1 and 2 and partially inhibited oxidation‐initiated cross‐linking of heavy meromyosin during myosin gelation with a lesser effect on light meromyosin. In the presence of PP, myosin exhibited less loss of conformational integrity upon oxidation than myosin without PP. Rheological analysis from 20 to 75 °C indicated up to 32% decreases (P < 0.05) in elastic modulus (G′) of myosin gels due to oxidation. However, the presence of 1 mM PP, which also lowered the gelling capacity of myosin, inhibited the oxidation‐induced G′ by nearly half (P < 0.05). These results suggest that the protection of myosin head from oxidative modification by PP can be a significant factor for the minimization of gelling property losses during cooking of comminuted meats.     

Non-enzymatic Depolymerization of Carrot Pectin: Toward a Better Understanding of Carrot Texture During Thermal Processing

ABSTRACT Pretreated carrot discs were thermally processed (90 °C to 110 °C) in closed containers and the resulting textural characteristics were analyzed. The pretreatment conditions used include conventional high‐temperature blanching (90 °C, 4 min), low‐temperature blanching (LTB = 60 °C, 40 min), LTB combined with 0.5% calcium chloride soaking, LTB combined with 2% sodium chloride soaking, high pressure pretreatment (HP = 400 MPa, 60 °C, 15 min), HP combined with 0.5% calcium chloride soaking, and control (non‐pretreated sample). Alcohol insoluble residues (AIR) from the pretreated carrot discs were characterized in terms of degree of methoxylation (DM). The AIR samples were further subjected to fractionation into water‐soluble pectin (WSP), chelator‐soluble pectin (CSP), and sodium carbonate‐soluble pectin (NSP). Heat depolymerization patterns and β‐elimination kinetics were investigated on the different pectin fractions. Thermal texture degradation was strongly influenced by the pretreatment condition used and the processing temperature during subsequent thermal treatment. Pretreatment conditions that showed a significant reduction in DM exhibited decreased WSP content, reduced β‐elimination, and consequently superior textural characteristics. β‐elimination was markedly pronounced in the highly methoxylated WSP fractions. CSP and NSP fractions were insensitive to β‐elimination. A strong correlation (r> 0.95) between thermal texture loss of carrots and β‐elimination kinetics exists. Overall, the benefits of controlled pectinmethylesterase activity in carrot processing were pointed out.     

Novel extracellular hyper acidophil and thermostable α‐amylase from Micrococcus sp.NS 211

Among several bacteria examined in this study, a hyper acidophil and thermostable Micrococcus sp.NS 211 designated as M.Amy NS 211 was selected for production of amylase using starch agar plates with following incubation at 85°C. Identification by 16SrRNA on selected bacterium disclosed the highest similarity for protean regions of this gene, 27 F and 1492R as Micrococcus sp.NS 211. Although activity of M.Amy NS 211 was established at temperatures between 70 and 110°C and pH ranges 1.2–8.0, the optimum temperature and pH was achieved at 85°C and 3.5 in sodium citrate buffer system respectively. Two‐step chromatography was performed using (CM Bio‐Gel A) and (Bio‐Gel A‐150) columns to purify 84 kDa hyper acidophil and thermostable α‐amylase. SDS‐PAGE analysis showed molecular mass and amylolytic activity as single band. Enhancement of enzyme activity was obtained in presence of 5 mM MnCl₂ (298%), CaCl₂ (347%), FeCl₂ (211%), MgCl₂ (253%), ZnCl₂ (146%), NiCl (142%), NaCl (141%), Na‐sulfate (153%) while inhibition was observed with (5 mM) EDTA, PMSF (3 mM), urea (8 M), and SDS (1%) at 143, 134, 43, and 119%, respectively. M.Amy NS 211 can be applied in laundry detergents, textile, and modern relevant industrial processes at extreme temperatures and under acidic conditions.     

Influence of NaCl and CaCl2 on Cold-Set Gelation of Heat-denatured Whey Protein

Heat‐denatured whey‐protein isolate (HD‐WPI) solutions were prepared by heating a 10 wt% WPI solution (pH 7) to 80 °C for 10 min and then cooling it back to 30 °C. Cold‐set gelation was initiated by adding either NaCl (0 to 400 mM) or CaCl₂ (0 to 15 mM). Both salts increased the turbidity and rigidity of the HD‐WPI solutions. Gelation rate and final gel strength increased with salt concentration and were greater for CaCl₂ than NaCl at the same concentration because the former is more effective at screening electrostatic interactions and can form salt bridges.     

Isolation and Characterization of Pectin Methylesterase from Apple Fruit

Two forms of the enzyme pectin methylesterase are evidenced in the apple (Malus communis). They differ both in their charge and molecular weight. The two enzymes were separated by DEAE-cellulose chromatography. Their molecular weights, determined by gel-filtration, were 55000 and 28000 daltons. The heavier form has been purified at homogeneity and subjected to investigations regarding its activity as a function of the pH and temperature, and determination of its kinetic parameters. The enzyme has a Km value of 1.05 mg/mL for citrus pectin and an optimum activity in the pH range between 6.5-7.5. The enzyme was stable up to 40°C. Incubation for 1 min at 90°C leads to its complete inactivation.     

The effect of thermal treatment on β‐glucosidase inactivation in vanilla bean (Vanilla planifolia Andrews)

The conditions for enzyme activity (pH and temperature) and kinetic parameters for the thermal inactivation of β‐glucosidase enzyme in vanilla beans have been investigated. The maximum enzyme activity was detected at pH 6.5 and 38 °C. The values obtained for V_max and K_m were 62.05 units and 2.07 mm, respectively. When hot water treatment (the most practical method of vanilla bean killing) was applied, β‐glucosidase treated at pH 6.0 and 60 °C for 3 min lost 51% of activity, while at 70 °C for 90 s the enzyme lost 60% of activity and at 80 °C for 30 s the enzyme lost 48% of its activity. When vanilla beans were cured in an oven at 60 °C for 36 to 48 h all β‐glucosidase activity was lost.     

Production of Cyclodextrins by Simultaneous Actions of Two CGTases from Three Strains of Bacillus

A moderate thermophile, which produces a large amount of extracellular and thermostable cyclomaltodextrin glucanotransferase, was isolated from soil samples at hot spring areas in Japan. The isolate (A-147), that grew at 30–60°C with an optimum at 45–50°C, was identified as a strain of Bacillus coagulans. The enzyme, which was most active at pH 6.5 and 60°C, was stabilized by 2–3 mM Ca²⁺, and degraded about 40% (as dry basis) of 13% (w/w) potato starch solution into cyclodextrins composed of α- and β-cyclodextrins dominantly at 74°C and pH 6.5 in the presence of 3 mM Ca²⁺. The ratio of α-, β- and γ-cyclodextrins was almost constant (2:2.6:1) during all stages of the reaction. By simultaneous actions of two enzymes from the bacterium and other 2 kinds of Bacillus, starch hydrolysates containing various amounts and rations of cyclodextrins were produced from starch. Ein mäßiges Thermophiles, das eine große Menge an extracellulärer und thermostabiler Glucanotransferase produziert, wurde aus Bodenproben in Heißquellengebieten von Japan isoliert. Das Isolat (A-147), das bei 30–60°C mit einem Optimum von 45–50°C wuchs, wurde als ein Stamm von Bacillus coagulans identifiziert. Das bei pH 6,5 und 60°C aktivste Enzym wurde durch 2–3 mM Ca²⁺ stabilisiert und baute etwa 40% (TS) einer 30%igen (w/w) Kartoffelstärkelösung zu Cyclodextrin ab, bestehend aus α- und β-Cyclodextrin, vorwiegend bei 74°C und pH 6,5 in Gegenwart von 3 mM Ca²⁺. Das Verhältnis von α-, β- und γ-Cyclodextrinen war nahezu konstant (2:2,6:1) während sämtlicher Reaktionsschritte. Durch gleichzeitige Einwirkung von zwei Enzymen aus dem Bakterium und zwei anderen Arten von Bacteria wurden Stärkehydrolysate hergestellt, die verschiedene Mengen und Verhältnisse an Cyclodextrinen enthielten.     

Further characterization of bacteriocins Plantaricin BN,bavaricin MN and pediocin A

We report here several characteristics of bacteriocins which are inhibitory to Clostridium botulinum and Listeria monocytogenes. Plantaricin BN produced by Lactobacillus plantarum BN, bavaricin MN produced by Lactobacillus bavaricus MN, and pediocin A produced by Pediococcus pentosaceus 43200 all demonstrated a bactericidal mode of action and retained some activity after heating at 60°C for ten minutes or 100°C for 5 minutes. pH and temperature optima for production on solid media were pH 7.9 and 15°C for plantaricin BN, pH 6.5 and 30°C for bavaricin MN, and pH 6.15–7.9 and 30°C for pediocin A. The molecular weight of plantaricin BN appeared greater than 10,000, that of bavaricin MN 22,600, and that of pediocin A 11,000. All of these bacteriocins were produced during the growth phase of the bacterial cultures.     

Purification,properties and heat inactivation of pectin methylesterase from apple (cv Golden Delicious)

Pectin methylesterase from apple (cv Golden Delicious) was extracted and purified by affinity chromatography on a CNBr‐Sepharose®‐PMEI column. A single pectin methylesterase peak was observed. Isoelectric points were higher than 9. Kinetic parameters of the enzyme were determined as K_m = 0.098 mg ml⁻¹ and V_max = 3.86 µmol min⁻¹ ml⁻¹ of enzyme. The optimum pH of the enzyme was above 7.5 and its optimum temperature was 63 °C. The purified PME required the presence of NaCl for optimum activity, and the sodium chloride optimum concentration increased with decreasing pH (from 0.13 M at pH 7 to 0.75 M at pH 4). The heat stability of purified PME was investigated without and with glycerol (50%), and thermal resistance parameters (D and Z values) were calculated showing that glycerol improved the heat resistance of apple PME. © 2000 Society of Chemical Industry     

Properties of Trypsin from the Pyloric Ceca of Atlantic Cod (Gadus morhua)

Trypsin (EC 3.4.21.4) was isolated from the pyloric ceca of Atlantic cod and purified to homogeneity by affinity chromatography. The enzyme catalyzed the hydrolysis of benzoyl arginine p-nitroanilide (BAPA, pH 8.2 and 25°C) such that V_max was 250 BAPA units per micromole trypsin and K_m was 1.48 mM. For the hydrolysis of tosyl arginine methyl ester (TAME, pH 8.1 and 25°C), V_max was 18.2 × 10³ TAME units/micromole trypsin, and K_m 0.22 mM. The pH and temperature optima with BAPA substrate were 7.5 and 40°C, respectively. Atlantic cod trypsin was most active and stable at alkaline pH. The enzyme was heat labile, losing more than 50% of its activity after incubation at 50°C for 30 min. Amino acid analysis of Atlantic cod trypsin revealed that the enzyme was rich in residues such as serine, glycine, glutamate and aspartate, but poor in basic amino acid residues compared to trypsins from warm blooded animals.     

Enhanced inactivation of pectin methyl esterase in orange juice using modified supercritical carbon dioxide treatment

The aim of the study is to quantify the effect of ethanol addition and exposure surface on the inactivation of pectin methyl esterase (PME), a juice clarifying enzyme, in orange juice using supercritical carbon dioxide (SC‐CO₂). Addition of ethanol to the SC‐CO₂ at 2% (v/v) caused greater inactivation than SC‐CO₂ alone, with a maximum reduction of PME activity of 97% at 30 MPa and 40 °C for 60 min. As the surface area to volume ratio was increased, the rate of inactivation of PME increased. Analysis of first‐order reaction kinetic data revealed that D values were greatly influenced by ethanol addition and agitation. With the addition of 2% ethanol, the D value reduced by half, that is, 56 min from 109 min. With impeller agitation of the sample at 1100 ± 100 rpm, the D value for PME was further reduced to 43 and 30 min without and with ethanol, respectively. The activity of PME treated with SC‐CO₂ remained unchanged after 14 days of storage at 4 °C. Treatment did not significantly change pH or colour, but did significantly increase the cloud values of the juice, resulting in a cloud stabilised juice with similar qualities to fresh juice.     

Multiple forms of β‐galactosidase from mango (Mangifera indica L Alphonso) fruit pulp

Mango (Mangifera indica L cv Alphonso) was found to contain three isoforms (I, II and III) of β‐galactosidase which, upon purification on Sephadex G‐200, had relative abundances of 44, 38 and 18%, respectively. The total specific activity increased from 20 to 727 µmol l^?1 upon purification, representing a ～36‐fold increase with a recovery of 0.28 U U^?1. The optimal pH for activity and stability were in the ranges 3.6–4.3 and 4–6.2, respectively. The optimal temperature for β‐galactosidase activity was between 42 and 47 °C with T_m in the range 45–51 °C. The K_m for pNP‐β‐galactopyranoside was 0.98, 1.11 and 0.95 mM , and V_max was 0.56, 0.53 and 0.35 µmol pNP min^?1, respectively for isoforms I, II and III. Hg²⁺ caused strong inhibition, whereas galacturonic acid, galactose, xylose, fucose and mannose slightly inhibited the activity of β‐galactosidase isoforms. The apparent molecular weights by GPC were 78, 58 and 91 kDa for isoforms I, II and III, respectively. The ability of these isoforms to degrade the endogenous substrate (arabinogalactan) possibly suggests a role in pectin dissolution during tissue softening/fruit ripening. Copyright © 2004 Society of Chemical Industry     

Physiochemical Properties,Microstructure, and Probiotic Survivability of Nonfat Goats’ Milk Yogurt Using Heat‐Treated Whey Protein Concentrate as Fat Replacer

There is a market demand for nonfat fermented goats’ milk products. A nonfat goats’ milk yogurt containing probiotics (Lactobacillus acidophilus, and Bifidobacterium spp.) was developed using heat‐treated whey protein concentrate (HWPC) as a fat replacer and pectin as a thickening agent. Yogurts containing untreated whey protein concentrate (WPC) and pectin, and the one with only pectin were also prepared. Skim cows’ milk yogurt with pectin was also made as a control. The yogurts were analyzed for chemical composition, water holding capacity (syneresis), microstructure, changes in pH and viscosity, mold, yeast and coliform counts, and probiotic survivability during storage at 4 °C for 10 wk. The results showed that the nonfat goats’ milk yogurt made with 1.2% HWPC (WPC solution heated at 85 °C for 30 min at pH 8.5) and 0.35% pectin had significantly higher viscosity (P < 0.01) than any of the other yogurts and lower syneresis than the goats’ yogurt with only pectin (P < 0.01). Viscosity and pH of all the yogurt samples did not change much throughout storage. Bifidobacterium spp. remained stable and was above 10⁶CFU g^‐1 during the 10‐wk storage. However, the population of Lactobacillus acidophilus dropped to below 10⁶CFU g^‐1 after 2 wk of storage. Microstructure analysis of the nonfat goats’ milk yogurt by scanning electron microscopy revealed that HWPC interacted with casein micelles to form a relatively compact network in the yogurt gel. The results indicated that HWPC could be used as a fat replacer for improving the consistency of nonfat goats’ milk yogurt and other similar products.