Synthetic β-endorphin-like peptide immunorphin binds to non-opioid receptors for β-endorphin on T lymphocytes

The synthetic decapeptide H-SLTCLVKGFY-OH (termed immunorphin) corresponding to the sequence 364–373 of the CH3 domain of human immunoglobulin G heavy chain was found to compete with [¹²⁵I]β-endorphin for high-affinity receptors on T lymphocytes from the blood of healthy donors (K_i = 0.6 nM). Besides immunorphin, its synthetic fragments H-Val-Lys-Gly-Phe-Tyr-OH (K_i = 15 nM), H-Leu-Val-Lys-Gly-Phe-Tyr-OH (K_i = 8.0 nM), H-Cys-Leu-Val-Lys-Gly-Phe-Tyr-OH (K_i = 3.4 nM), H-Thr-Cys-Leu-Val-Lys-Gly-Phe-Tyr-OH (K_i = 2.2 nM), H-Leu-Thr-Cys-Leu-Val-Lys-Gly-Phe-Tyr-OH (K_i = 1.0 nM) possessed the ability to inhibit specific binding of [¹²⁵I]β-endorphin to T lymphocytes. Tests of the specificity of the receptors revealed that they are not sensitive to naloxone and Met-enkephalin, i.e. they are not opioid receptors. K_d values characterizing the specific binding of ¹²⁵I- labeled immunorphin and its fragment H-Val-Lys-Gly-Phe-Tyr-OH to the receptors have been determined to be 7.4 nM and 36.3 nM, respectively.     

Interaction of beta-cyclodextrin with cadmium(II) ions

Reduction of Cd(II) on a dropping mercury electrode was used to study interaction of β-cyclodextrin with Cd(II) ions. It was found that Cd(II) forms Cdβ-CD(OH)₂²⁻ hydroxy-complex with the anion of β-cyclodextrin in alkaline solutions (pH>11), the logarithm of stability constant being 10.4±0.1 (20 °C; I=1.0). The calculated value of the diffusion coefficient equal to 1.0×10⁻⁶ cm²/s shows a large size Cd(II) complex species formation in alkaline solutions containing β-CD.     

Enzyme markers and isolation of the microvillar and perimicrovillar membranes of Dysdercus peruvianus (hemiptera: pyrrhocoridae) midgut cells

The midgut of Dysdercus peruvianus is divided into four sections (V₁-V₄). All the cells have microvilli ensheathed by a lipoprotein membrane (perimicrovillar membrane) extending toward the lumen as narrow tubes with dead ends. Subcellular fractionation of V₁ and V₂ tissue in isotonic and hypotonic conditions showed that -glucosidase is associated with membranous structures larger than those associated with β-glucosidase. The /β-glucosidase activity ratio is 34 ± 4 in V₁ tissue and 170 ± 10 in membranes recovered from the V₁ luminal contents. These membranes are resolved in sucrose gradients into low density (1.087 ± 0.001 g/cm³) -glucosidase-carrying membranes (/β-glucosidase activity ratio of 330±30) and high density (1.132 ± 0.002g/cm³) β-glucosidase-carrying-membranes. Low-density membranes have 1090 ± 60 μg lipid/mg protein and apparently are not contaminated by high-density ones (electron micrographs). SDS-polyacrylamide gel electrophoresis (SDS-PAGE) showed that membranes recovered from V₁ luminal contents are composed mainly of a-glucosidase-rich membranes. The data suggest that -glucosidase-rich membranes are perimicrovillar membranes which may be partly lost into luminal contents on dissection, with densities and lipid/protein ratios similar to that of myelin sheaths, in accordance with previous freeze-fracture data. β-Glucosidase-rich membranes are probably microvillar membranes with densities increased by the presence of associated portasomes.     

Estradiol inhibits the estrone sulfatase activity in normal and cancerous human breast tissues

It is well accepted that estradiol (E₂) plays an important role in the genesis and evolution of breast cancer. Quantitative evaluation indicates that in human breast tumor, estrone sulfate (E₁S) ‘via sulfatase’ is a much more likely precursor for E₂ than is androstenedione ‘via aromatase’. In previous studies, it was demonstrated that in isolated MCF-7 and T-47D breast cancer cell lines, estradiol can block estrone sulfatase activity. In the present study, the effect of E₂ was explored using total normal and cancerous breast tissues. This study was carried out with post-menopausal patients with breast cancer. None of the patients had a history of endocrine, metabolic or hepatic diseases or had received treatment in the previous 2 months. Each patient received local anaesthetic (lidocaine 1%) and two regions of the mammary tissue were selected: (A) the tumoral tissue and (B) the distant zone (glandular tissue) which was considered as normal. Samples were placed in liquid nitrogen and stored at –80 °C until enzyme activity analysis. Breast cancer histotypes were ductal and post-menopausal stages were T2. Homogenates of tumoral or normal breast tissues (45–75 mg) were incubated in 20 mM Tris–HCl, pH 7.2 with physiological concentrations of [³H]-E₁S (5 × 10⁻⁹ M) alone or in the presence of E₂ (5 × 10⁻⁵ to 5 × 10⁻⁷ M) during 30 min or 3 h. E₁S, E₁ and E₂ were characterized by thin layer chromatography and quantified using the corresponding standard. The sulfatase activity is significantly more intense with the breast cancer tissue than normal tissue, since the concentration of E₁ was 3.20 ± 0.15 and 0.42 ± 0.07 pmol/mg protein, respectively after 30 min incubation. The values were 27.8 ± 1.8 and 3.5 ± 0.21 pmol/mg protein, respectively after 3 h incubation. Estradiol at the concentration of 5 × 10⁻⁷ M inhibits this conversion by 33% and 31% in cancerous and normal breast tissues, respectively and by 53% and 88% at the concentration of 5 × 10⁻⁵ M after 30 min incubation. The values were 24% and 18% for 5 × 10⁻⁷ M and 49% and 42% for 5 × 10⁻⁵ M, respectively after 3 h incubation. It was observed that [³H]-E₁S is only converted to [³H]-E₁ and not to [³H]-E₂ in normal or cancerous breast tissues, which suggests a low or no 17β-hydroxysteroid dehydrogenase (17β-HSD) Type 1 reductive activity in these experimental conditions. In conclusion, estradiol is a strong anti-sulfatase agent in cancerous and normal breast tissues. This data can open attractive perspectives in clinical trials using this hormone.     

Properties of endogenous β-glucosidase of a Saccharomyces cerevisiae strain isolated from Sicilian musts and wines

Three hundred sixty-one yeast strains (80 of which ascribable to Saccharomyces cerevisiae) were isolated from Sicilian musts and wines with the purpose of looking for β-glucosidase (βG, EC 3.2.1.21) activity. Of these, the AL 41 strain had highest endogenous βG activity and was identified as belonging to the species S. cerevisiae by biochemical and molecular methods. This enzyme was subsequently characterized. It had optimum effect at pH 3.5–4.0, whilst optimum temperature was 20 °C, compatible with typical wine-cellar conditions; it was not inhibited by ethanol, at concentrations of 12–14%, or fructose and glucose. The βG was also characterised in terms of the kinetic parameters K_m (2.55 mM) and V_max (1.71 U mg⁻¹ of protein). Finally, it remained stable for at least 35 days in model solutions of must and wine.     

Angiotensin II and angiotensin-(1-7) inhibit the inner cortex Na+ -ATPase activity through AT2 receptor

In the present paper, the modulation of the basolateral membrane (BLM) Na⁺-ATPase activity of inner cortex from pig kidney by angiotensin II (Ang II) and angiotensin-(1–7) (Ang-(1–7)) was evaluated. Ang II and Ang-(1–7) inhibit the Na⁺-ATPase activity in a dose-dependent manner (from 10⁻¹¹ to 10⁻⁵ M), with maximal effect obtained at 10⁻⁷ M for both peptides. Pharmacological evidences demonstrate that the inhibitory effects of Ang II and Ang-(1–7) are mediated by AT₂ receptor: The effect of both polypeptides is completely reversed by 10⁻⁸ M PD 123319, a selective AT₂ receptor antagonist, but is not affected by either (10⁻¹²–10⁻⁵ M) losartan or (10⁻¹⁰–10⁻⁷ M) A779, selective antagonists for AT₁ and AT_(1–7) receptors, respectively. The following results suggest that a PTX-insensitive, cholera toxin (CTX)-sensitive G protein/adenosine 3′,5′-cyclic monophosphate (cAMP)/PKA pathway is involved in this process: (1) the inhibitory effect of both peptides is completely reversed by 10⁻⁹ M guanosine 5′-O-(2-thiodiphosphate) (GDPβS; an inhibitor of the G protein activity), and mimicked by 10⁻¹⁰ M guanosine 5′-O-(3-thiotriphosphate) (GTPγS; an activator of the G protein activity); (2) the effects of both peptides are mimicked by CTX but are not affected by PTX; (3) Western blot analysis reveals the presence of the G_s protein in the isolated basolateral membrane fraction; (4) (10⁻¹⁰–10⁻⁶ M) cAMP has a similar and non-additive effect to Ang II and Ang-(1–7); (5) PKA inhibitory peptide abolishes the effects of Ang II and Ang-(1–7); and (6) both angiotensins stimulate PKA activity.     

Bradykinin counteracts the stimulatory effect of angiotensin-(1-7) on the proximal tubule Na+ -ATPase activity through B2 receptor

Recently, we demonstrated that angiotensin-(1–7) (Ang-(1–7)) stimulates the Na⁺-ATPase activity through a losartan-sensitive angiotensin receptor, whereas bradykinin inhibits the enzyme activity through the B₂ receptor [Regul. Pept. 91 (2000) 45; Pharmacol. Rev. 32 (1980) 1]. In the present paper, the effect of bradykinin (BK) on Ang-(1–7)-stimulated Na⁺-ATPase activity was evaluated. Preincubation of Na⁺-ATPase with 10⁻⁹ M Ang-(1–7) increases enzyme activity from 7.9±0.9 to 14.1±1.5 nmol Pi mg⁻¹ min⁻¹, corresponding to an increase of 79% (p<0.05). This effect is reverted by bradykinin in a dose-dependent manner (10⁻¹⁴–10⁻⁸ M), reaching maximal inhibitory effect at 10⁻⁹ M. Des-Arg⁹ bradykinin (DABK), an agonist of B₁ receptor, at the concentrations of 10⁻⁹–10⁻⁷ M, does not mimic the BK inhibitory effect, and des-Arg⁹-[Leu⁸]-BK (DALBK), a B₁ receptor antagonist, at the concentrations of 10⁻¹⁰–10⁻⁷ M, does not prevent the inhibitory effect of BK on Ang-(1–7)-stimulated enzyme. On the other hand, HOE 140, an antagonist of B₂ receptor, abolishes the inhibitory effect of BK on the Ang-(1–7)-stimulated enzyme in a dose-dependent manner, reaching maximal effect at 10⁻⁷ M. Taken together, these data indicate that stimulation of B₂ receptors by BK can counteract the stimulatory effect of Ang-(1–7) on the proximal tubule Na⁺-ATPase activity.     

Hydrodynamic properties of the fractions of mannan formed by Rhodotorula rubra yeast

Aqueous solutions of fractions of an extracellular linear mannan formed by Rhodotorula rubra yeast have been investigated by hydrodynamic methods (high-speed sedimentation, translation isothermic diffusion and viscometry). The molecular weight was determined according to Svedberg ( ) and the polydispersity parameters of the initial sample were also determined (M_w/M_n = 1·20 and M_z/M_w = 1·21). Relationships between the molecular weight (M) and s_o, D_o and [η] in the range were: [η] = 2·33 × 10⁻² M^0.75, D_o = 1·65 × 10⁻⁴ M^0·58, s_o = 2·24 × 10⁻¹⁵ M^0·43. The equilibrium rigidity and hydrodynamic diameter of chains representing mannan molecules were evaluated.     

The F1-ATPase beta-subunit is the putative enterostatin receptor

It has been suggested that the F₁-ATPase β-subunit is the enterostatin receptor. We investigated the binding activity of the purified protein with a labeled antagonist, β-casomorphin_1–7, in the absence and presence of cold enterostatin. ¹²⁵I-β-casomorphin_1–7 weakly binds to the rat F₁-ATPase β-subunit. Binding was promoted by low concentrations of cold enterostatin but displaced by higher concentrations. To study the relationship between binding activity and feeding behavior, we examined the ability of a number of enterostatin analogs to affect β-casomorphin_1–7 binding to the F₁-ATPase β-subunit. Peptides that suppressed food intake promoted β-casomorphin_1–7 binding whereas peptides that stimulated food intake or did not affect the food intake displaced β-casomorphin_1–7 binding. Surface plasmon resonance measurements show that the β-subunit of F₁-ATPase binds immobilized enterostatin with a dissociation constant of 150 nM, where no binding could be detected for the assembled F₁-ATPase complex. Western blot analysis showed the F₁-ATPase β-subunit was present on plasma and mitochondrial membranes of rat liver and amygdala. The data provides evidence that the F₁-ATPase β-subunit is the enterostatin receptor and suggests that enterostatin and β-casomorphin_1–7 bind to distinct sites on the protein.     

Piezoelectric properties of poly-β-hydroxybutyrate and copolymers of β-hydroxybutyrate and β-hydroxyvalerate

The shear piezoelectricity was observed in oriented films of poly-β-hydroxybutyrate (PHB) and copolymers of β-hydroxybutyrate (HB) and β-hydroxyvalerate (HV). The piezoelectric stress constant 3₁₄ = e′₁₄ − ie″₁₄ (polarization/strain), the piezoelectric strain constant d₁₄ = d′₁₄ − id″₁₄ (polarization/stress), the elastic constant c = c′ + ic″ and the dielectric constant = ′ − i″ were determined at a frequency of 10 Hz over a temperature range from −150° to +150°C. Piezoelectric relaxations as well as elastic and dielectric relaxations were clearly observed at the glass transition temperature of about 15°C. In order to evaluate the piezoelectric constants (e₂ and d₂) for the piezoelectric phase which consists of the crystalline region and the oriented non-crystalline region, a spherical dispersion two phase model was utilized. Assuming the appropriate fixed values for the elastic and dielectric constants in the piezoelectric phase, d₂ and d₂ were calculated as a function of temperature. For a PHB and a copolymer (17 HV/83 HB), e₂ and d₂ showed relaxations, leading to a conclusion that the instantaneous piezoelectric constant in the crystalline phase is constant independent of temperature but the piezoelectric constant in the oriented non-crystalline phase is relaxational and has the opposite sign. For a copolymer (25 HV/75 HB) and a chloroform treated copolymer (17 HV/83 HB), e₂ and d₂ were constant independent of temperature, indicating that the oriented non-crystalline phase has disappeared owing to the increased molecular flexibility due to copolymerization or annealing in chloroform vapour.     

Kinetics and mechanism of the stoichiometric oxygenation of [Cu(fla)(idpa)]ClO4 [fla=flavonolate, IDPA=3,3′-imino-bis(N,N-dimethylpropylamine)] and the [Cu(fla)(idpa)]ClO4-catalysed oxygenation of flavonol

Oxygenation of [Cu^II(fla)(idpa)]ClO₄ (fla=flavonolate; IDPA=3,3′-iminobis(N,N-dimethylpropylamine)) in dimethylformamide gives [Cu^II(idpa)(O-bs)]ClO₄ (O-bs=O-benzoylsalicylate) and CO. The oxygenolysis of [Cu^II(fla)(idpa)]ClO₄ in DMF was followed by electronic spectroscopy and the rate law −d[{Cu^II(fla)(idpa)}ClO₄]/dt=k_obs[{Cu^II(fla)(idpa)}ClO₄][O₂] was obtained. The rate constant, activation enthalpy and entropy at 373 K are k_obs=6.13±0.16×10⁻³ M⁻¹ s⁻¹, ΔH^‡=64±5 kJ mol⁻¹, ΔS^‡=−120±13 J mol⁻¹ K⁻¹, respectively. The reaction fits a Hammett linear free energy relationship and a higher electron density on copper gives faster oxygenation rates. The complex [Cu^II(fla)(idpa)]ClO₄ has also been found to be a selective catalyst for the oxygenation of flavonol to the corresponding O-benzoylsalicylic acid and CO. The kinetics of the oxygenolysis in DMF was followed by electronic spectroscopy and the following rate law was obtained: −d[flaH]/dt=k_obs[{Cu^II(fla)(idpa)}ClO₄][O₂]. The rate constant, activation enthalpy and entropy at 403 K are k_obs=4.22±0.15×10⁻² M⁻¹ s⁻¹, ΔH^‡=71±6 kJ mol⁻¹, ΔS^‡=−97±15 J mol⁻¹ K⁻¹, respectively.     

Fourier transform infrared spectroscopy used to evidence the prevention of beta-sheet formation of amyloid beta(1-40) peptide by a short amyloid fragment

Reflectance Fourier transform infrared (FT-IR) microspectroscopy was applied to study the prevention of β-sheet formation of amyloid β (Aβ)(1–40) peptide by co-incubation with a hexapeptide containing a KLVFF sequence (Aβ(15–20) fragment). Second-derivative spectral analysis was used to locate the position of the overlapping components of the amide I band of Aβ peptide and assigned them to different secondary components. The result indicates that each intact sample of Aβ(15–20) fragment or Aβ(1–40) peptide previously incubated in distilled water at 37 °C transformed their secondary structure from 1649 (1651) or 1653 cm⁻¹ to 1624 cm⁻¹, suggesting the transformation from -helix and/or random coil structures to β-sheet structure. By co-incubating both samples with different molar ratio in distilled water at 37 °C, the structural transformation was not found for Aβ(1–40) peptide after 24 h-incubation. But the β-sheet formation of Aβ(1–40) peptide after 48 h-incubation was evidenced from the appearance of the IR peak at 1626 cm⁻¹ by adding a little amount of Aβ(15–20) fragment. There was no β-sheet formation of Aβ(1–40) peptide after addition with much amount of Aβ(15–20) fragment, however, suggesting the higher amount of Aβ(15–20) fragment used might inhibit the β-sheet formation of Aβ(1–40) peptide. The more Aβ(15–20) fragment used made the more stable structure of Aβ(1–40) peptide and the less β-sheet formation of Aβ(1–40) peptide. The study indicates that the reflectance FT-IR microspectroscopy can easily evidence the prevention of β-sheet formation of Aβ(1–40) peptide by a short amyloid fragment.     

Incorporation of the antibacterial agent, miconazole nitrate into a cellulosic fabric grafted with β-cyclodextrin

The aim of the study was to investigate the incorporation of the antibacterial agent, miconazole nitrate into cyclodextrin cavities covalently bonded onto cloth fibers. The cellulosic fabric was grafted with β-cyclodextrin molecules through reaction with monochlorotriaziny β-cyclodextrin (MCT-β-CD). The suitable bonded reaction conditions were found to be MCT-β-CD 60–100 g/L, catalyst Na₂CO₃ 50–60 g/L, the reaction temperature 150–160 °C and the reaction time 5–8 min.

The modified and unmodified fabrics were characterized by UV spectrophotometry. The level of miconazole nitrate entrapped in the fabrics were determined by HPLC and was founded to be much higher (0.458% w/w) for the textile functionalized with MCT-β-CD compared to the unmodified fabric (0.056% w/w). The antibacterial abilities measured by shaker flask method showed that the antibacterial property was markedly enhanced by impregnation with miconazole nitrate of the MCT-β-CD grafted textile. The finished fabric kept the antibacterial abilities more than 70% even after washing 10 times, while the antibacterial activity of the unmodified textile was almost lost.     

Luminescent yeast cells entrapped in hydrogels for estrogenic endocrine disrupting chemical biodetection

In the construction of luminescent yeast cell based fibre-optic biosensors, we demonstrate a novel approach for estrogenic endocrine disrupting chemical (EDC) biodetection by entrapping genetically modified Saccharomyces cerevisiae cells, containing the estrogen receptor alpha-mediated expression of the luc reporter gene, in hydrogel matrices based on calcium alginate or PVA. In order to insure a significant signal, an optimal immobilization ratio of 1:2 alginate 3% (w/v): 5 × 10⁶ [cells/ml], respectively, was used with the highest 17-β-estradiol (β-E2) induction factor after 2.5 h of incubation with 10 [nM] β-E2. It was shown that biocompatible alginate beads, 4.27–4.55 × 10⁵ [CFU/bead], which were characterized by a detection limit of 0.08 [μg l⁻¹] and an EC50 of 0.64 [μg l⁻¹] for β-E2, retained their viability for luminescence measurements after 1 month of storage at −80 °C slow freeze condition, and thus repeated cell cultivations were not required. The assay reproducibility for each tested EDC, represented by the coefficients of variation (CV), ranged from 4.35 to 18.47%. An alternative immobilization method, based on a room temperature partial drying of polyvinyl alcohol (PVA) solution (LentiKat^® Liquid) and cell suspension mix, was investigated with only a slightly lower detection limit for β-E2 than that reported with alginate beads. Alginate yeast based hydrogels may also be applicable to the analysis of environmental water samples since the trend of detected estrogenic activities with alginate beads roughly correlated with LC–MS–MS analytical results.     

The differential regulation of aldosterone output in hamster adrenal by angiotensinII and adrenocorticotropin.

Aldosterone was isolated from hamster adrenal cells and was identified by high performance liquid chromatography and thermospray mass spectroscopy analysis. Basal outputs from adrenal cell suspensions were of the same order of magnitude, 8.4 ± 1.9 ng and 8.0 ± 0.7 ng/2 h/50,000 cells, for aldosterone and corticosteroid, respectively. The outputs of aldosterone and corticosteroid increased with K⁺ concentrations to reach maxima of 3.3- and 1.6-fold at 10 meq/l of K⁺. Angiotensin_II (A_II) produced dose-dependent increases in aldosterone and corticosteroid outputs with maxima of 3- and 4-fold, respectively. In contrast, ACTH induced relatively no changes in aldosterone output, whereas dose-dependent increases in corticosteroid output were found. In time study experiments, with 10⁻⁸ M A_II, aldosterone and corticosteroid outputs were maximally increased after 1 h (6-fold) and 3 h (1.8-fold), respectively. At 10⁻⁸ M, ACTH had a small stimulatory effect on aldosterone output after 6 h, whereas it provoked a gradual increase in corticosteroid output (up to 7-fold after 8 h of incubation). The effects of A_II and ACTH on adrenal cytochrome P-450_11β involved in the last steps of aldosterone formation were evaluated by c combined in vivo andin vitro experiments. The P-450_11β mRNA level was increased by a low sodium intake but not by a 24 h ACTH stimulus. These results taken together indicate that ACTH and A_II differentially regulate P-450_11β. It is postulated that these two regulatory peptides regulate the hamster adrenal steroidogenesis by different P-450 genes.     

Influence of lipid peroxidation on [H]ketanserin binding to 5-HT2 prefrontal cortex receptors

The influence of lipid peroxidation on 5-HT₂ receptor binding was examined in prefrontal cortex membranes from sheep brain. Lipid peroxidation was induced with ascorbic acid and ferrous sulphate and measured by the thiobarbituric acid method. In lipid-peroxidized membranes, [³H]ketanserin specific binding was inhibited. The B_max values decreased by 80%, from 50.1±3.5 fmol/mg protein in control membranes to 10.1±2.0 fmol/mg protein in peroxidized membranes, indicating a decrease in the number of 5-HT₂ binding sites. However, the K_D values for the [³H]ketanserin specific binding did not significantly change. In order to further characterize [³H]ketanserin binding, the inhibition potency (IC₅₀ values) of antagonists or agonists of serotonin and dopamine receptors for [³H]ketanserin specific binding was determined. In control membranes, the order of the inhibition potency of the drugs tested was the following: ketanserin (−log [IC₅₀] = 8.56±0.70) ritanserin (−log [IC₅₀] = 8.13±0.30) methysergide (−log [IC₅₀] = 7.42±0.50) spiperone (−log [IC₅₀] = 7.23±0.18) serotonin (−log [IC₅₀] = 6.99±0.65) haloperidol (−log [IC₅₀] = 6.95±0.65) dopamine (−log [IC₅₀] = 5.82±0.76). After membrane lipid peroxidation, the IC₅₀ value for ritanserin was significantly increased, suggesting a decreased capacity for displacing [³H]ketanserin specific binding. Other antagonists of 5-HT₂ receptors showed apparent increases in IC₅₀ values upon peroxidation, whereas spiperone was shown to be the most potent drug (−log [IC₅₀] = 7.19±1.06) in inhibiting [³H]ketanserin specific binding. A decrease in polyunsaturated fatty acids, namely docosahexaenoic acid (22:6) was also observed in peroxidized membranes. These results indicate a modulating role of the surrounding lipids and of the physical properties of the membranes on the binding activity of 5-HT₂ receptors upon the lipid peroxidation process, which can be involved in the tissue impairment that occurs during the aging process and in post-ischemic situations.     

Influence of phosphorus concentration on the growth kinetics and stoichiometry of the microalga Scenedesmus obliquus

The growth of the freshwater microalga Scenedesmus obliquus was studied at 30°C in a mineral culture medium with phosphorus concentrations of between 0 and 372 μ . The values for the specific growth rates, between and , fitted a semistructured substrate-limitation model with μ_m1 = 0·0466 h⁻¹, μ_m2 = 0·0256 h⁻¹ and . The specific uptake rate of phosphorus reached a maximum value of q_Sm1 = 658·01 × 10⁻⁴ μmol P mg⁻¹ biomass h⁻¹.     

Chaperone activities of bovine and camel beta-caseins: Importance of their surface hydrophobicity in protection against alcohol dehydrogenase aggregation

β-Casein (β-CN) showing properties of intrinsically unstructured proteins (IUP) displays many similarities with molecular chaperones and shows anti-aggregation activity in vitro. Chaperone activities of bovine and camel β-CN were studied using alcohol dehydrogenase (ADH) as a substrate. To obtain an adequate relevant information about the chaperone capacities of studied caseins, three different physical parameters including chaperone constant (k_c, μM⁻¹), thermal aggregation constant (k_T, °C⁻¹) and aggregation rate constant (k_t, min⁻¹) were measured. Bovine β-CN displays greater chaperone activity than camel β-CN. Fluorescence studies of 8-anilino-1-naphthalenesulfonic acid (ANS) binding demonstrated that bovine β-CN is doted with larger effective hydrophobic surfaces at all studied temperatures than camel β-CN. Greater relative hydrophobicity of bovine β-CN than camel β-CN may be a factor responsible for stronger interactions of bovine β-CN with the aggregation-prone pre denatured molecular species of the substrate ADH, which resulted in greater chaperone activity of bovine β-CN.     

Immobilization of horseradish peroxidase on chitosan/silica sol-gel hybrid membranes for the preparation of hydrogen peroxide biosensor

A simple and effective strategy for fabrication of hydrogen peroxide (H₂O₂) biosensor has been developed by entrapping horseradish peroxidase (HRP) in chitosan/silica sol–gel hybrid membranes (CSHMs) doped with potassium ferricyanide (K₃Fe(CN)₆) and gold nanoparticles (GNPs) on platinum electrode surface. The hybrid membranes are prepared by cross-linking chitosan (CS) with 3-aminopropyltriethoxysilane (APTES), while the presence of GNPs improved the conductivity of CSHMs, and the Fe(CN)₆^3−/4− was used as a mediator to transfer electrons between the electrode and HRP due to its excellent electrochemistry activity. UV–Vis absorption spectroscopy was employed to characterize the different components in the CSHMs and their interaction. The parameters influencing the performance of the resulting biosensor were optimized and the characteristic of the resulting biosensor was characterized by cyclic voltammetry and chronoamperometry. Linear calibration for hydrogen peroxide was obtained in the range of 3.5 × 10^− 6 to 1.4 × 10^− 3 M under the optimized conditions with the detection limit (S/N = 3) of 8.0 × 10^− 7 M. The apparent Michaelis–Menten constant of the enzyme electrode was 0.93 mM. The enzyme electrode retained about 78% of its response sensitivity after 30 days. The system was applied for the determination of the samples, and the results obtained were satisfactory.     

Thermodynamics of the disproportionation of adenosine 5'-diphosphate to adenosine 5'-triphosphate and adenosine 5'-monophosphate, II. Experimental data

High-pressure liquid-chromatography and microcalorimetry have been used to determine equilibrium constants and enthalpies of reaction for the disproportionation reaction of adenosine 5′-diphosphate (ADP) to adenosine 5′-triphosphate (ATP) andadenosine 5′-monophosphate (AMP). Adenylate kinase was used to catalyze this reaction. The measurements were carried out over the temperature range 286 to 311 K, at ionic strengths varying from 0.06 to 0.33 mol kg⁻¹, over the pH range 6.04 to 8.87, and over the pMg range 2.22 to 7.16, where pMg = -log a(Mg²⁺). The equilibrium model developed by Goldberg and Tewari (see the previous paper in this issue) was used for the analysis of the measurements. Thus, for the reference reaction: 2 ADp³⁻ (ao) AMp²⁻ (ao)+ ATp⁻ (ao), K° = 0.225 ± 0.010, ΔG° = 3.70 +- 0.11 kJ mol ⁻¹, ΔH° = −1.5 ± 1. 5 kJ mol ⁻¹, °S ° = −17 ± 5 J mol⁻¹ K⁻¹, and ACP_p°≈ = −46 J mo1l⁻¹ K⁻¹ at 298.15 K and 0.1 MPa. These results and the thermodynamic parameters for the auxiliary equilibria in solution have been used to model the thermodynamics of the disproportionation reaction over a wide range of temperature, pH, ionic strength, and magnesium ion morality. Under approximately physiological conditions (311.15 K, pH 6.94, [Mg²⁺] = 1.35 × 10⁻³ mol kg⁻¹, and I = 0.23 mol kg⁻¹) the apparent equilibrium constant (K_A′ = m(ΣAMP)m(ΣATP)/[ m(ΣADP)]²) for the overall disproportionation reaction is equal to 0.93 ± 0.02. Thermodynamic data on the disproportionation reaction and literature values for this apparent equilibrium constant in human red blood cells are used to calculate a morality of 1.94 × 10⁻⁴ mol kg⁻¹ for free magnesium ion in human red blood cells. The results are also discussed in relation to thermochemical cycles and compared with data on the hydrolysis of the guanosine phosphates.