Denaturation and aggregation of three alpha-lactalbumin preparations at neutral pH

The denaturation and aggregation of reagent-grade (Sigmaalpha-La), ion-exchange chromatography purified (IEXalpha-La), and a commercial-grade (Calpha-La) alpha-lactalbumin were studied with differential scanning calorimetry (DSC), polyacrylamide gel electrophoresis, and turbidity measurement. All three preparations had similar thermal denaturation temperatures with an average of 63.7 degrees C. Heating pure preparations of alpha-lactalbumin produced three non-native monomer species and three distinct dimer species. This phenomenon was not observed in Calpha-La. Turbidity development at 95 degrees C (tau95 degrees C) indicated that pure preparations rapidly aggregate at pH 7.0, and evidence suggests that hydrophobic interactions drove this phenomenon. The Calpha-La required 4 times the phosphate or excess Ca2+ concentrations to develop a similar tau95 degrees C to the pure preparations and displayed a complex pH-dependent tau95 degrees C behavior. Turbidity development dramatically decreased when the heating temperature was below 95 degrees C. A mechanism is provided, and the interrelationship between specific electrostatic interactions and hydrophobic attraction, in relation to the formation of disulfide-bonded products, is discussed.     

Double phosphorylation of the myosin regulatory light chain during rigor mortis of bovine Longissimus muscle

To investigate changes in myosin light chains (MyLCs) during postmortem aging of the bovine longissimus muscle, we performed two-dimensional gel electrophoresis followed by identification with matrix-assisted laser desorption ionization time-of-flight mass spectrometry. The results of fluorescent differential gel electrophoresis showed that two spots of the myosin regulatory light chain (MyLC2) at pI values of 4.6 and 4.7 shifted toward those at pI values of 4.5 and 4.6, respectively, by 24 h postmortem when rigor mortis was completed. Meanwhile, the MyLC1 and MyLC3 spots did not change during the 14 days postmortem. Phosphoprotein-specific staining of the gels demonstrated that the MyLC2 proteins at pI values of 4.5 and 4.6 were phosphorylated. Furthermore, possible N-terminal region peptides containing one and two phosphoserine residues were detected in each mass spectrum of the MyLC2 spots at pI values of 4.5 and 4.6, respectively. These results demonstrated that MyLC2 became doubly phosphorylated during rigor formation of the bovine longissimus, suggesting involvement of the MyLC2 phosphorylation in the progress of beef rigor mortis. Keywords: Bovine; myosin regulatory light chain (RLC, MyLC2); phosphorylation; rigor mortis; skeletal muscle.     

Thermal denaturation and aggregation properties of Atlantic salmon myofibrils and myosin from white and red muscles

Thermal denaturation and aggregation abilities of salmon myofibrils and myosin were studied measuring turbidity, intrinsic fluorescence, 8-anilino-1-naphthalene sulfonic acid binding, and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide cross-linking. The thermal behaviors of protein preparation from white and red muscles were compared, and the relationship with thermal gelation properties is discussed. The low gelation ability of salmon muscle proteins was related to a limited extent of protein denaturation and aggregation upon heating. These properties seemed to be carried by myosin molecules as a similar behavior was observed for both myofibrils and myosin preparations. The higher thermal stability observed for red muscle proteins with higher transition temperatures in rheological profiles was related to a shift to higher temperature in denaturation and aggregation processes. The extent of denaturation and aggregation was very similar for both muscle types as was the final rigidity of the gels formed.     

Isolation and partial characterization of two types of muscle collagen in some cephalopods

Collagen from muscle of volador (Illex coindetii), pota (Toradopsis eblanae), and white octopus (Eledone cirrhosa) was characterized in terms of anatomical location, sex, and maturity. Collagen content was higher in arms than in mantle in all three species; there were also significant differences in octopus depending on the age of the individual. Concerning sex, the largest differences in the amount of collagen were found in relation to total protein content. In volador and pota, collagen solubility was higher in the mantle than in the arms, and in the case of pota there were also sex-related differences. In octopus males, solubility was higher in the arms. Two types of collagen, I and V, were identified as the principal constituents in all three species and at both anatomical locations (mantle and arms). The electrophoretic mobility of the alpha2 chain differed in the two types of collagen, but the amino acid compositions of the collagen were similar in the mantle and arms in all three species examined.     

Role of neck region in the thermal aggregation of myosin.

Carp dorsal myosin formed oligomers that retained ATPase activity upon heating. Cleavage of the oligomeric myosin at subfragment-1 (S-1)/rod junction released monomeric S-1 and rod, indicating that ATPase retaining myosin associated near the S-1/rod junction. The digest also contained rod oligomers. Heating a mixture of S-1 and rod generated neither ATPase retaining S-1 oligomers nor rod oligomers. Electron microscopic observation of the heated myosin revealed that some oligomers were formed by associating at the S-1/rod joining region, exhibiting a recognized double head, probably ATPase retaining oligomers. No myosin oligomers associated at the tail region were observed, thus, rod aggregation would be formed at its very restricted region near the S-1/rod junction. Based on the findings, we proposed that the neck structure is important in the thermal oligomerization process of myosin.     

alpha-tocopherol oxidation in beef and in bovine muscle microsomes

The oxidation of alpha-tocopherol (TH) in beef was analyzed using a stable isotope dilution capillary gas chromatography-mass spectrometry assay. TH decreased while alpha-tocopherolquinone (TQ) and 2,3-epoxy-alpha-tocopherolquinone (TQE(2)) increased in ground longissimus lumborum (LL) and psoas major (PM) muscles during storage (P < 0.10). In LL steaks, the relative concentrations of TH decreased and TQ and TQE(2) increased in surface samples; changes were less dramatic in deep samples. Deuterated alpha-tocopherolhydroquinone (THQ) standard was not recovered and endogenous THQ was not detected in meat; THQ was measurable in microsomes isolated from PM and incubated in the presence of 2, 2'-azobis(2-amidopropane)HCl (ABAP) or myoglobin. ABAP-challenged microsomes yielded a tocopherol product profile which favored 5, 6-epoxy-alpha-tocopherolquinone (TQE(1)) and TQE(2), while the use of myoglobin as prooxidant resulted in a higher proportion of TQ and THQ. Results demonstrated that concentrations of TH decreased and TQ and TQE(2) increased in meat during storage and are consistent with the peroxy-radical scavenging function of tocopherol.     

Degradation of four phenylurea herbicides by mixed populations of microorganisms from two soil types

The metabolism of four substituted urea herbicides by microbial populations from two soils was investigated. The herbicides were two dimethylurea herbicides (fluometuron and chloroxuron) and two methoxymethylphenylurea herbicides (metobromuron and chlorbromuron), and the two soils used were Louisiana Commerce Loam and Indiana Silt Loam. The dimethyl compounds were successively N-demethylated by microorganisms from both soils. N-demethylation of the methoxymethyl herbicides, however, was significant only in the Louisiana soil, while N-demethoxylation was found in only trace amounts in all the cultures. Metabolism of these herbicides apparently was predominantly via direct hydrolysis to the aniline, which in turn underwent further transformations.     

Effect of two types of lime on some soil physical attributes of an oxisol from Brazil

Abstract

Liming is a very important practice, especially in acid soils such as some Oxisols from Brazil, in which the chemical effects of liming are reasonably known. However, the effects of liming on soil physical properties are either not well known or, at least, controversial. In order to verify, under field conditions, the effect of liming on some soil physical attributes such us the aggregate stability, the water dispersible clay, the water retention were evaluated in a citrus liming experiment on a Dark Red Latosol (Typic Haplorthox) clay texture, from Cordeirópolis, São Paulo State, Brazil. The soil samples were collected in 0–10 cm and 10–20 cm depths, three years after the application of lime in the experiment, which had two replications per treatment. Calcitic and dolomitic limes were used with application rates of 0, 3, and 9 Mg ha^‐1, with a total of five treatments, i.e., one control and two rates of application of each kind of lime. The results showed an increase in the aggregate stability. As a consequence, a significant linear correlation was found between aggregate mean weighted diameter (MWD) and calcium content and other indices affected by liming such as pH, sum of bases, base saturation and aluminum content. An increase in the stability indices (MWD) resulted in an increase in the water retained at 0.03 MPa and 0.1 MPa.     

Relationship between nitrogen and phosphate surplus from agricultural production and river water quality in two types of production structure

Abstract

We examined the relationship between nitrogen (N) and phosphate (P₂O₅) surplus derived from agriculture and river water quality. We selected two river basins; one was a paddy farming area (Omoigawa) and the other was an intensive livestock husbandry area (Nakagawa). Nitrogen and P₂O₅ surpluses, defined as the difference between their input and output on regional farmland, from farmland in Omoigawa were twice those of Nakagawa; the surpluses came mainly from chemical fertilizer use. Although N and P₂O₅ surpluses in Nakagawa were lower, Nakagawa had a large amount of non-utilized livestock excreta, twice the N surplus on farmland in Nakagawa. Residual N and P₂O₅ in the river basin, caused by surplus and non-utilized livestock excreta, were approximately 20% and 40%, respectively, higher in Nakagawa than in Omoigawa. Outflows of N and P₂O₅ to river water were higher in Omoigawa than Nakagawa. By excluding domestic sewage N and P₂O₅ in river water, we calculated the loads of non-point source N and P₂O₅. Non-point source N and P₂O₅ were higher in Omoigawa than Nakagawa. This inverse result might be caused by the different source of residual N (i.e. chemical N fertilizer or livestock excreta).     

供体细胞种类对牛核移植效果的影响

供体细胞种类对牛卵母细胞核移植效果影响的研究结果表明,颗粒细胞的核移植效果与成年牛成纤维细胞无显著差异(19.8%vs 17.3%,P>0.05);胎牛成纤维细胞核移植囊胚发育率高于成年牛成纤维细胞(24.2%vs 14.0%,P<0.05);血清饥饿处理的胎牛成纤维细胞(休眠细胞)与高血清培养(非休眠细胞)的胎牛成纤维细胞核移植效果差异不显著(18.8%vs 18.6%,P>0.05),表明胎牛成纤维细胞血清饥饿处理没有必要。     

Relationship between the metabolite profile and technological properties of bovine milk from two dairy breeds elucidated by NMR-based metabolomics

The aim of the present study was to investigate the relationship between the metabolite profile of milk and important technological properties by using nuclear magnetic resonance (NMR)-based metabolomics. The metabolomics approach was introduced for the metabolic profiling of a set of milk samples from two dairy breeds representing a wide span in coagulation properties. The milk metabolite profiles obtained by proton and carbon NMR spectroscopy could be correlated to breed and, more interestingly, also with the coagulation profile, as established by traditional methods by using principal component analysis (PCA). The metabolites responsible for the separation into breed could mainly be ascribed to carnitine and lactose, whereas the metabolites varying in the samples with respect to coagulation properties included citrate, choline, carnitine, and lactose. The results found in the present study demonstrated a promising potential of NMR-based metabolomics for a rapid analysis and classification of milk samples, both of which are useful for the dairy industry.     

High-pressure effects on lysosome integrity and lysosomal enzyme activity in bovine muscle

This study was conducted to determine whether the application of high hydrostatic pressure could modify the enzymatic activity and membrane integrity of lysosomes in muscle. Several combinations of pressure (0-600 MPa) and time (0-300 s) were applied to two types of samples: purified enzymes (cathepsin D and acid phosphatase) in buffer solution and intact muscle (biceps femoris). The enzymes studied showed varying degrees of susceptibility depending on the level of pressure, holding time, and environment. Acid phosphatase activity was minimally affected by pressure in buffer solution, whereas cathepsin D was modulated significantly by the pressure and time applied. The activities of the enzymes extracted from meat increased with pressure. The cytochemical observations showed the presence of primary and secondary lysosomes in muscles. After pressurization, the membrane integrity of the lysosomes was modified. A correlation could be established between lysosomal enzymatic activities and the lysosome membrane breakdown.     

两种枣树矿质营养元素累积特性研究

为探明枣树矿质营养元素的累积分配特征,以3年生骏枣树和灰枣树为试材,采用彻底刨根、分解取样的方法,研究了生物量的构成特点、各器官矿质元素含量和累积分配特性。结果表明,骏枣树总干质量为2694.3 g/plant,其中营养器官占68.0%,分别比灰枣高27.6%和21.9%。其N、P、K、Ca、Mg总累积量为33.91、3.43、22.20、31.25和5.53 g/plant,分别比灰枣树高50.1%、22.5%、24.7%、51.0%和88.7%。其中,N主要分配到叶片和果实,P、K主要分配到果实和叶片,Ca、Mg主要分配到叶片和主干;新生营养器官N、P、K的吸收比例为1: 0.063～0.083: 0.41～0.46,果实N、P、K的吸收比例为1: 0.19～0.20: 1.34～1.48。每生产1000 kg干质量骏枣需吸收N 32.83、P 3.41、K 23.14、Ca 29.06、Mg 5.32 kg;灰枣需吸收N 20.53、P 2.66、K 17.71、Ca 18.01、Mg 2.49 kg。骏枣生产单位干质量果实需吸收的养分比灰枣多,养分利用效率比灰枣低。骏枣树养分在叶片中的分配率显著高于灰枣树,在果实中的分配率则显著低于灰枣树。     

Trace element concentrations in the dorsal muscle of white suckers and brown bullheads from two acidic Adirondack lakes

Concentrations of 14 elements (Ag, As, Cd, Cr, Cu, Hg, Ni, Pb, Sb, Se, Sn, Tl, V, and Zn) were measured in the dorsal muscle of omnivorous white suckers and brown bullheads, fish likely to ingest sediment while feeding. The fish were collected in two acidic Adirondack lakes known to have elevated concentrations of several of these elements in their surface sediment. Trace element concentrations in the muscle of the white suckers and brown bullheads were compared with government guidelines for edible fish and survey data for concentrations present in commercial species. Only the largest white sucker exceeded the U.S. Food and Drug Administration's guidelines for Hg in food of 1 ppm (wet wt). Concentrations of all the other elements measured were at low or innocuous levels in the edible muscle. A significant positive correlation was found between body weight and Hg concentrations in both species. Zinc was negatively correlated for the same parameters in white suckers. With the possible exception of Hg, our results indicate that elevated levels of potentially toxic elements in the surface sediment of the two acidic Adirondack lakes are not reflected in the edible muscle of the omnivorous bottom feeding fish present in these systems.     

Influence of two insecticides, chlorpyrifos and quinalphos, on arginine ammonification and mineralizable nitrogen in two tropical soil types

Effects of seed treatments with chlorpyrifos [5 g of active ingredient (ai) kg(-1) of seed] and quinalphos (6.25 g of ai kg(-1) of seed) and standing crop treatments with chlorpyrifos (800 g of ai ha(-1)) and quinalphos (1000 g of ai ha(-1)) on arginine deamination and mineralizable nitrogen were monitored, in the sandy loam and loamy sand soils of two tropical semiarid fields, for three consecutive crop seasons. The arginine ammonification activity of rhizospheric microbes was inhibited after seed treatment with chlorpyrifos and quinalphos and their principal metabolites, 3,5,6-trichloro-2-pyridinol (TCP) and 3,5,6-trichloro-2-methoxypyridine (TMP) and 2-hydroxyquinoxaline and quinoxaline-2-thiol, respectively. Quinalphos produced transient inhibitions, whereas chlorpyrifos and its metabolites (TCP and TMP) exerted a greater inhibition in both loamy sand and sandy loam soils. Arginine ammonification by nonrhizospheric microbes was stimulated by standing crop treatments with both pesticides. In the loamy sand soil, the parent compounds stimulated rhizospheric N-mineralization, whereas the metabolites were inhibitory. However, nonrhizospheric N-mineralization was inhibited by both chlorpyrifos and quinalphos and stimulated by their metabolites. A higher magnitude of inhibition of arginine deamination in the loamy sand than in the sandy loam soil could be due to greater bioavailability of the pesticides in the former, resulting from lesser sorption of the pesticides due to alkalinity of the soil and its low content of clay and organic carbon. Although both pesticides affected mineralizable nitrogen, seed treatment with quinalphos and standing crop treatment with quinalphos and chlorpyrifos produced the most significant effects. The recommended doses of the pesticides not only efficiently controlled whitegrubs, which increased pod yields, but also left no residues in harvested kernels. They also caused no long-term inhibition of ammonification, which could have been of significant concern during the short crop period in semiarid areas where nitrogen determines plant productivity.     

两类温带森林下土壤氮的有效性指标比较

To evaluate the validity of different indices in estimating soil readily mineralizable N, soil microbial biomass （Nmic）, soil active N （SAN）, soluble organic N （SON）, net N mineralization rate （NNR） and gross N mineralization rate （GNR） in mineral soils （0-10 cm） from six forest stands located in central Germany were determined and compared with two sampling times： April and November. Additionally, soil density fractionation was conducted for incubated soils （with addition of ^15NH4-N and glucose, 40 days） to observe the sink of added ^15N in different soil fractions. The study showed that Nmic and NNR in most stands differed significantly （P 〈 0.05） between the two sampling times, but not GNR, SAN and SON. In November, no close relationships were found between GNR and other N indices, or between Nrnic, SON, and SAN and forest type. However, in April, GNR was significantly correlated （P 〈 0.05） with Nmic, SAN, and NNR along with Nmlc under beech being significantly higher （P 〈 0.05） than under conifers. Furthermore, density fractionation revealed that the light fraction （LF, 0.063-2 mm, 〉 1.7 gcm^-3） was not correlated with the other N indices. In contrast, results from the incubation study proved that more 15N was incorporated into the heavy fraction （HF 〈 0.063 ram, 〉 1.7 g cm^-3） than into LF, indicaing that more labile N existed in HF than in LF. These findings suggested that attention should be paid to the differences existing in N status between agricultural and forest soils.     

Competitiveness of native Bradyrhizobium japonicum strains in two different soil types

Interstrain competitiveness is a key factor affecting the performance of rhizobium inoculant. In the present study five native strains of Bradyrhizobium japonicum, namely SSF 4, SSF 5, SSF 6, SSF 7 and SSF 8, were assessed for their competitiveness in nodulating soybean using serological methods. The strains were inoculated individually or with the type strain USDA 110 at a 1:1 ratio. Nodule occupancy determined by immunofluorescence and dot immunoblot assay revealed that under in vitro conditions SSF 8 is more competitive than USDA 110 whereas the others were less competitive. The competitive ability of these strains was also estimated in pot culture in the field. In red soil both SSF 8 and USDA 110 were equally competitive whereas in black soil SSF 8 competed better than USDA 110 and produced more nodules. In a black soil field trial using a randomized block design, USDA 110 or SSF 8, when inoculated alone, occupied the majority of the nodules and enhanced nodule dry weight and shoot biomass. SSF 8 was more competitive when the strains were co-inoculated. Received: 1 November 1996