Cladodes from prickly pear as a functional ingredient: effect on fat retention, oxidative stability, nutritional and sensory properties of cookies.

The stems of Opuntia ficus-indica known as cladodes are rich source of bioactive and functional substances, which make them important candidate for the production of health-promoting food. Cladodes powder was incorporated at different levels of substitution (2.5%, 5% and 7.5%) in cookies (butter/wheat flour: 55/100 m/m). Substitution of wheat flour by cladodes powder improved dietary fiber, ash, potassium, magnesium and calcium contents of enriched cookies. The results also revealed that cladodes supplementation increased hardness; however, it decreased a* and b* values and reduced exudate loss of cookies during storage. Moreover, rising levels of cladodes powder contribute to the increase of antioxidant activity of cookies and decreased their oxidative degradation. Sensory evaluation showed that cladodes supplementation at 5% level remained acceptable at 5-point hedonic scale. The present study suggested that cladodes supplementation in high-fat cookies not only added nutritional value to food, but also improved its functional characteristics.

A cysteine protease isolated from the latex of Ficus microcarpa: purification and biochemical characterization.

A plant protease named microcarpain was purified from the latex of Ficus microcarpa by acetonic (20-40 % saturation) precipitation, Sephadex G-75 filtration, and Mono Q-Sefinose FF chromatography. The protease was purified with a yield of 9.25 % and a purification factor of 8. The molecular weight of the microcarpain was estimated to be 20 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The purified enzyme showed maximum activity at pH 8.0 and at a temperature of 70 °C. Proteolytic activity was strongly inhibited by dithio-bis-nitrobenzoic acid (DTNB), Hg(2+), and Cu(2+). The N-terminal amino acid sequence of the purified microcarpain "VPETVDWRSKGAV" showed high homology with a protease from Arabidopsis thaliana. Inhibition studies and N-terminal sequence classified the enzyme as a member of the cysteine peptidases family.

Optimization of acid protease production by Aspergillus niger I1 on shrimp peptone using statistical experimental design.

Medium composition and culture conditions for the acid protease production by Aspergillus niger I1 were optimized by response surface methodology (RSM). A significant influence of temperature, KH(2)PO(4), and initial pH on the protease production was evaluated by Plackett-Burman design (PBD). These factors were further optimized using Box-Behnken design and RSM. Under the proposed optimized conditions, the experimental protease production (183.13 U mL(-1)) closely matched the yield predicted by the statistical model (172.57 U mL(-1)) with R(2) = 0.914. Compared with the initial M1 medium on which protease production was 43.13 U mL(-1), a successful and significant improvement by 4.25 folds was achieved in the optimized medium containing (g/L): hulled grain of wheat (HGW) 5.0; KH(2)PO(4) 1.0; NaCl 0.3; MgSO(4)(7H(2)O) 0.5; CaCl(2) (7H(2)O) 0.4; ZnSO(4) 0.1; Na(2)HPO(4) 1.6; shrimp peptone (SP) 1.0. The pH was adjusted at 5 and the temperature at 30°C. More interestingly, the optimization was accomplished using two cheap and local fermentation substrates, HGW and SP, which may result in a significant reduction in the cost of medium constituents.

Molecular and biochemical characterization of an extracellular serine-protease from Vibrio metschnikovii J1.

A protease-producing bacterium was isolated from an alkaline wastewater of the soap industry and identified as Vibrio metschnikovii J1 on the basis of the 16S rRNA gene sequencing and biochemical properties. The strain was found to over-produce proteases when it was grown at 30 degrees C in media containing casein as carbon source (14,000 U ml(-1)). J1 enzyme, the major protease produced by V. metschnikovii J1, was purified by a three-step procedure, with a 2.1-fold increase in specific activity and 33.3% recovery. The molecular weight of the purified protease was estimated to be 30 kDa by SDS-PAGE and gel filtration. The N-terminal amino acid sequence of the first 20 amino acids of the purified J1 protease was AQQTPYGIRMVQADQLSDVY. The enzyme was highly active over a wide range of pH from 9.0 to 12.0, with an optimum at pH 11.0. The optimum temperature for the purified enzyme was 60 degrees C. The activity of the enzyme was totally lost in the presence of PMSF, suggesting that the purified enzyme is a serine protease. The kinetic constants K (m) and K (cat) of the purified enzyme using N-succinyl-L-Ala-L-Ala-L-Pro-L-Phe-p-nitroanilide were 0.158 mM and 1.14 x 10(5) min(-1), respectively. The catalytic efficiency (K (cat) /K (m)) was 7.23 x 10(8) min(-1) M(-1). The enzyme showed extreme stability toward non-ionic surfactants and oxidizing agents. In addition, it showed high stability and compatibility with some commercial liquid and solid detergents. The aprJ1 gene, which encodes the alkaline protease from V. metschnikovii J1, was isolated, and its DNA sequence was determined. The deduced amino acid sequence of the preproenzyme differs from that of V. metschnikovii RH530 detergent-stable protease by 12 amino acids, 7 located in the propeptide and 5 in the mature enzyme.