Thermal stability of human matrix metalloproteinases.

Matrix metalloproteinases (MMP) are key players in the remodelling of the extracellular matrix under physiological and pathological conditions. Thermodynamic parameters of human recombinant metalloproteinases of the active (rMMP2, 3, 7, 8 and 9) and latent (rPro-MMP2, 3 and 9) forms were obtained by differential scanning calorimetry (DSC). Temperature by itself does not result in autocatalysis of recombinant MMP. The transitions observed by DSC correspond to structural domains of the monomeric protein. In this study, we show the domain organization of these proteins, where the thermal transition (Tm) of the main component is observed at 71.3 °C (ProMMP-2); 74.8 °C (ProMMP-8); 80.0 °C (ProMMP-3); 92.6 °C (ProMMP-9) and 98.3 °C (ProMMP-7). For MMP-3, this main Tm is related to the catalytic domain (CD). The isolated recombinant CD of MMP-3 unfolds as a single transition at Tm 83.4 °C, matching the more stable domain observed in the full-length active form of rMMP-3. The denaturation profile of rProMMP-3 shows the main transition at Tm 80 °C, a less stable domain before the propeptide domain (PD) cleavage. Our results indicate that the structural stability of MMP and particularly their CD are not substantially altered after cleavage of the PD. We propose that the thermodynamic parameters obtained by DSC are relevant for the functional study of MMP, particularly to reveal their contribution in complex biological samples in health and disease.

Reducción de proteínas y glucosa por reacción de Maillard en leche con lactosa hidrolisada / Protein and glucose reduction by Maillard reaction in milk with hydrolyzed lactose

Se evaluó el efecto de la temperatura sobre la desnaturalización de proteínas y la reacción de Maillard en leche entera y descremada con lactosa hidrolizada. Las leches hidrolizadas se trataron térmicamente a 100, 110, 120 y 130 °C durante un período de 1 hora y se midió la concentración de glucosa, el grado de pardeamiento y la desnaturalización de proteínas. El grado de dorado en la leche entera varió de 14.4 (100 °C) a 42.6 (130 °C). Para la leche descremada fue de 20.2 (100 °C) a 38.0 (130 °C). La concentración de glucosa en leche entera (47% p/v) y en leche descremada (41% p/v) después del tratamiento térmico (130 °C) mostró una reducción significativa en relación con el control (25 °C). El efecto de la temperatura en la desnaturalización de proteínas en leche entera y descremada en relación con el control (25 °C) fue del 100%. La leche tratada térmicamente con lactosa hidrolizada promovió la desnaturalización de proteínas con un aumento del pardeamiento característico de la reacción de Maillard, lo que afectó la calidad nutricional.

The effect of temperature in protein denaturation and Maillard reaction in whole and skim milk with hydrolyzed lactose was evaluated. Hydrolyzed milk was thermally treated at 100, 110, 120 and 130 °C over a period of 1 hour and glucose concentration, browning degree and protein denaturation were measured. The browning degree in whole milk varied from 14.42 (100 °C) to 42.63 (130 °C) and 20.21 (100 °C) to 38.03 (130 °C) in skim milk. Glucose concentration in whole milk (47% - w/v) and skim milk (41% - w/v) after heat treatment (130 °C) showed a significant reduction in relation to the control (25 °C). The temperature effect in protein denaturation in whole and skim milk in relation to the control (25 °C) was 100%. Thermally treated milk with hydrolyzed lactose promoted protein denaturation with increasing browning characteristic of the Maillard reaction, thus affecting the nutritional quality.

Quality parameters of chicken breast meat affected by carcass scalding conditions.

OBJECTIVE: The influence of broiler carcass scalding conditions on chicken breast meat quality parameters was investigated. METHODS: Two hundred and seventy Cobb broiler chickens from 42 to 48 days old were slaughtered according to the standard industry practice and scalded in five temperature/time combinations-T1, 54°C/210 s; T2, 55°C/180 s; T3, 56°C/150 s; T4, 57°C/120 s; T5, 58°C/90 s. RESULTS: Scalding temperature increase resulted in higher values of external and ventral lightness and in protein functionality reduction-determined by emulsification capacity and protein denaturation-in chicken breast fillets 24 h post-mortem. Protein secondary structures had conformational changes, with a decrease of the α-helix and an increase of the ß-sheet and ß-turn proportions, mainly in T1 and T5 samples, determined by Fourier-transform infrared spectroscopy in an attenuated reflectance mode analysis. The chemical composition, pH, water holding capacity and Warner-Bratzler shear force did not differ among the treatments. In the fatty acid profile, the 18:1n-9 was lower in T5, which suggested that the high scalding-temperature could have caused the lipid oxidation. The values of the polyunsaturated fatty acids (PUFA), such as 22:2, 20:4n-6, and 22:6n-3, were highest in the T5, thus being related to the phospholipid cellular membrane collapse in this experimental condition and subsequent release of these PUFA. CONCLUSION: Intermediate scalding-parameters avoided the negative changes in the chicken meat quality.

Evaluation of in vitro and in vivo therapeutic efficacy of Ribes alpestre Decne in Rheumatoid arthritis

Rheumatoid arthritis is an autoimmune inflammatory disorder, despite the discovery of numerous drugs there is still need to introduce newer, safer and more effective sources of drugs such as medicinal herbs. Present research work was an attempt to appraise the antiarthritic potential of Ribes alpestre Decne in rheumatoid arthritis. In vitro inhibition of protein (bovine serum albumin and egg albumin) denaturation, Human red blood cell membrane stabilization assays along with formaldehyde induced arthritis in rats were commenced in this study. Findings of present investigation demonstrated significant and dose dependent antiarthritic effect of Ribes alpestre. Aqueous ethanolic extract, butanol and aqueous fraction illustrated 95%, 69.233% and 92.840% protection at 6400 ug/mL against bovine serum albumin denaturation respectively. Similarly, plant extract together with butanol and aqueous fractions showed 3653.47%, 1484.03% and 3563.19% inhibition of pathological alteration of egg albumin in that order. Moreover, hydroethanolic extract with butanol and aqueous fraction exhibited 91.29%, 65.73% and 89.62% stabilization against erythrocyte hemolysis at 6400 ug/mL correspondingly. Furthermore, hydroethanolic extract, butanol and aqueous fraction notably 73.49%, 66.42% and 68.87% decreased paw edema at highest dose (200 mg/kg). Similarly aqueous ethanolic extract, butanol and aqueous fraction illustrated 72.38%, 54.90% and 66.33% decrease in paw thickness at 200 mg/kg. Hence results suggested that Ribes alpestre possess antiarthritic potential thus supporting its use as natural remedy in rheumatic conditions.

Kinetics and Thermodynamics of Thermal Inactivation of β-Galactosidase from Aspergillus oryzae

ABSTRACT For optimization of biochemical processes in food and pharmaceutical industries, the evaluation of enzyme inactivation kinetic models is necessary to allow their adequate use. Kinetic studies of thermal inactivation of β-galactosidase from Aspergillus oryzae were conducted in order to critically evaluate mathematical equations presented in the literature. Statistical analysis showed that Weibull model presented the best adequacy to residual enzymatic activity data through the processing time and its kinetic parameters as a function of the temperature, in the range of 58-66 ºC. The investigation suggests the existence of a non-sensitive heat fraction on the enzyme structure, which is relatively stable up to temperatures close to 59 ºC. Thermodynamic parameters were evaluated and showed that such β-galactosidase presents activation energy of 277 kJ mol-1 and that the enzyme inactivation is due to molecular structural changes. Results shown that the enzyme is quite stable for biotechnological applications.

Further studies on membrane stabilizing, anti-inflammatory and FCA induced arthritic activity of various fractions of bark of Machilus macrantha in rats

Machilus macrantha Nees, Lauraceae, bark is traditionally used in the treatment of asthma, tuberculosis and rheumatoid arthritis. In order to validate, mechanism based anti-inflammatory activity of fractions M. macrantha bark are investigated for first time. Test materials viz. petroleum ether (PE), alkaloidal fraction (CH), acetone extracts (TAN) and mucilage (MM) (250 and 500 mg/kg, p.o.) obtained from M. macrantha bark were tested for membrane stabilizing, anti-nociceptive; anti inflammatory and Freund's complete adjuvant (FCA) induced arthritis activity. Diclofenac sodium and morphine were used as the reference standards in pharmacological assay. Test materials have significantly (p<0.01) inhibited paw edema after Carrageenan and histamine induction at higher doses. Administration of test materials of M. macrantha (250 and 500 mg/kg b.w.) significantly reduced abdominal writhing, formalin nociception, cotton pellet granuloma and vascular permeability in experimental animal. In addition to this, bark of M. macrantha showed chronic anti-rheumatic effect by suppressing the swelling volume, arthritis index, hematological and biochemical parameters (ESR, RA factor, CRP, liver transferase enzyme) in FCA-induced arthritis. It also significantly inhibited protein denaturation, heat-induced haemolysis of RBC and reduction in total leukocyte migration. Bioassay guided fractionation of the pet. ether extract of bark of M. macrantha led to isolation and characterization of β-sitosterol and stigma sterol confirmed by its HPLC, NMR and GC-MS study. In conclusion, extracts of M. macrantha bark can be explored as a therapeutic agent for the treatment of acute and chronic arthritis.

Caracterización de la formulación de -Macroagregados de albúmina / Characterization of - macroaggregates formulation

RESUMEN En este trabajo se realizó la caracterización del proceso de formación y marcaje de los macroagregados de albúmina (MAA) con . Con este fin se efectuaron estudios comparativos para establecer el papel de la desnaturalización en presencia del durante la formulación de -Sn-MAA, para ello se utilizaron tres formulaciones. Se realizó un análisis estructural cualitativo de la albúmina sérica humana para describir el sistema a través de un modelo gráfico. Los resultados mostraron diferencias importantes en el comportamiento radioquímico de las tres formulaciones. La formulación de -Sn-MAA (A) mostró un comportamiento superior, con purezas radioquímicas cercanas a 95% en los tiempos estudiados y una disociación inferior al 20% en 24 h.El modelo gráfico permitió explicar, a través de sencillas representaciones, los procesos involucrados en el sistema estudiado, permitiendo una mejor comprensión de este.

ABSTRACT The aim of this paper was to study the formation and radiolabelling of 99m Tcalbumin macroaggregates. Comparatives studies of three formulations to establish the role of denaturalization and in the formulation of -Sn-MAA were carried out. A qualitative structural analysis of human serum albumin was performed in order to give a description of the studied system using a graph model. The results showed important differences in the radiochemical behavior of the three formulations. -Sn-MAA (A) formulation had the best behavior, with radiochemical purity close to 95% and dissociation below 20% after 24 h of being radiolabeled. The graph model explained with simple representations the process involved in the studied system;thus allowing a better knowledge of it.