Nε-Acetyl L-α Lysine Improves Activity and Stability of α-Amylase at Acidic Conditions: A Comparative Study with other Osmolytes

Background: Nε-acetyl L-α lysine is an unusual acetylated di-amino acid synthesized and accumulated by certain halophiles under osmotic stress. Osmolytes are generally known to protect proteins and other cellular components under various stress conditions. Objective: The structural and functional stability imparted by Nε-acetyl L-lysine on proteins were unknown and hence was studied and compared to other commonly known bacterial osmolytes - ectoine, proline, glycine betaine, trehalose and sucrose. Methods: Effects of osmolytes on the temperature and pH profiles, pH stability and thermodynamic stability of the model enzyme, α-amylase were analyzed. Results: At physiological pH, all the osmolytes under study increased the optimal temperature for enzyme activity and improved the thermodynamic stability of the enzyme. At acidic conditions (pH 3.0), Nε-acetyl L-α lysine and ectoine improved both the catalytic and thermodynamic stability of the enzyme; it was reflected in the increase in residual enzyme activity after incubation of the enzyme at pH 3.0 for 15 min by 60% and 63.5% and the midpoint temperature of unfolding transition by 11°C and 10°C respectively. Conclusion: Such significant protective effects on both activity and stability of α-amylase imparted by addition of Nε-acetyl L-α lysine and ectoine at acidic conditions make these osmolytes interesting candidates for biotechnological applications.

Evaluation of the Diagnostic Accuracy of the CareStart™ Glucose-6-Phosphate Dehydrogenase Deficiency Rapid Diagnostic Test among Chinese Newborns

Previous studies have shown that the CareStart™ G6PD Deficiency rapid diagnostic test has high diagnostic accuracy on G6PD deficiency in Africa and Thailand, but not in China. As there are regional differences of G6PD genotype distribution, we are attending to verify the effectiveness of the kit in Chinese population. The study cohort included 247 newborns admitted to our hospital for jaundice. The quantitative detection of G6PD enzyme activity and G6PD gene mutations analysis was used to classify the status of G6PD deficiency. The performance of CareStart™ assays was verified by calculating the sensitivity, specificity and area under the receiver operating characteristic curve (AUC) based on the corrected G6PD deficiency status. In male newborns, the sensitivity of the CareStart™ assay was 98.9%, the specificity was 94.2% and the AUC was 0.97. In female newborns, the sensitivity was 58.5% when the cutoff value of residual enzyme activity was 100%; however, the sensitivity was 100% when the cutoff value was 60%. Therefore, the CareStart™ test can effectively screen G6PD deficiency in male newborns and female infants with less than 60% residual enzyme activity, female infants with residual enzyme activities of 60–100% are more likely to be missed diagnosed among Chinese newborns.

Chemical, enzymatic and physical characteristic of cloudy apple juices

In this study cloudy juices from six apple cultivars: ‘Alwa’, ‘Fiesta’, ‘Gloster’, ‘Golden Delicious’, ‘Mutsu’ and ‘Pinova’ were characterized in respect of polyphenols content by UPLC, PME (pectin methylesterase) activity, color, viscosity, and stability of turbidity. Apple cultivar affected significantly the chemical, enzymatic and physical properties of juices. Total quantitated polyphenols ranged from 686.63 mg l-1 (‘Gloster’) to 988.63 mg l-1 (‘Alwa’), and polymeric proanthocyanidins were a dominant group of these compounds. All of products contained also high content of phenolic acids, mainly chlorogenic. The thermal treatment of juices did not cause a complete inactivation of pectin methylesterase. Taking into account the % of residual enzyme activity, the pasteurization was more efficient in the case of ‘Fiesta’ and ‘Pinova’ juices (13% and 14% of the initial activity, respectively). Examined juices were characterized by low values of a stable turbidity (18.07–37.75%), despite relatively high viscosity (2.40–9.60 mPas).

Factors Affecting the Softening of Pickled Pasteurised Cucumbers

During the last three seasons the specific softening of pickled cucumbers was observed. The defective samples were analysed, but no microbial contamination was confirmed and no residual enzyme activity as well. The hypothesis of residual activity of microbial pectinases and cellulases as the most probable softening cause was proposed. The cellulolytic and pectolytic activities of nineteen strains of moulds and yeasts isolated from the samples of soils, cucumbers and cucumber plants rests were compared. The inactivation parameters (D and z values) of pectolytic enzymes of the most active strains were determined. The inhibitory effect of Ca²⁺ addition was evaluated within the model experiments. The residual enzyme activities were confirmed as the main cause of the defect, together with other factors such as the characteristic composition of microbial contamination, the stress or other damage of the cucumbers during the postharvest manipulation (chilling injury, humidity stress, etc.), microbial contamination of cucumbers before processing, conditions of washing, heat treatment parameters, etc. The practical recommendations for the prevention of the defect were formulated.