Isolation of lactoperoxidase using different cation exchange resins by batch and column procedures

Lactoperoxidase (LP) was isolated from whey protein by cation-exchange using Carboxymethyl resin (CM-25C) and Sulphopropyl Toyopearl resin (SP-650C). Both batch and column procedures were employed and the adsorption capacities and extraction efficiencies were compared. The resin bed volume to whey volume ratios were 0.96:1.0 for CM-25C and ≤ 0.64:1.0 for SP-650 indicating higher adsorption capacity of SP-650 compared to CM-25C. The effluent LP activity depended on both the enzyme activity in the whey and the amount of whey loaded on the column within the saturation limits of the resin. The percentage recovery was high below the saturation point and fell off rapidly with over-saturation. While effective recovery was achieved with column extraction procedures, the recovery was poor in batch procedures. The whey-resin contact time had little impact on the enzyme adsorption. SDS PAGE and HPLC analyses were also carried out, the purity was examined and the proteins characterised in terms of molecular weights. Reversed phase HPLC provided clear distinction of the LP and lactoferrin (LF) peaks. The enzyme purity was higher in column effluents compared to batch effluents, judged on the basis of the clarity of the gel bands and the resolved peaks in HPLC chromatograms.

A benchmark method for global optimization problems in structure determination from powder diffraction data

Quasi-Newton-Raphson minimization and conjugate gradient minimization have been used to solve the crystal structures of famotidine form B and capsaicin from X-ray powder diffraction data and characterize the chi(2) agreement surfaces. One million quasi-Newton-Raphson minimizations found the famotidine global minimum with a frequency of ca 1 in 5000 and the capsaicin global minimum with a frequency of ca 1 in 10 000. These results, which are corroborated by conjugate gradient minimization, demonstrate the existence of numerous pathways from some of the highest points on these chi(2) agreement surfaces to the respective global minima, which are passable using only downhill moves. This important observation has significant ramifications for the development of improved structure determination algorithms.

Bayesian decision procedures for dose-escalation based on evidence of undesirable events and therapeutic benefit.

In this paper, Bayesian decision procedures are developed for dose-escalation studies based on bivariate observations of undesirable events and signs of therapeutic benefit. The methods generalize earlier approaches taking into account only the undesirable outcomes. Logistic regression models are used to model the two responses, which are both assumed to take a binary form. A prior distribution for the unknown model parameters is suggested and an optional safety constraint can be included. Gain functions to be maximized are formulated in terms of accurate estimation of the limits of a therapeutic window or optimal treatment of the next cohort of subjects, although the approach could be applied to achieve any of a wide variety of objectives. The designs introduced are illustrated through simulation and retrospective implementation to a completed dose-escalation study. Copyright © 2006 John Wiley & Sons, Ltd.

Bayesian decision procedures for binary and continuous bivariate dose-escalation studies

In this paper, Bayesian decision procedures are developed for dose-escalation studies based on binary measures of undesirable events and continuous measures of therapeutic benefit. The methods generalize earlier approaches where undesirable events and therapeutic benefit are both binary. A logistic regression model is used to model the binary responses, while a linear regression model is used to model the continuous responses. Prior distributions for the unknown model parameters are suggested. A gain function is discussed and an optional safety constraint is included. Copyright (C) 2006 John Wiley & Sons, Ltd.

Optimization of protoplast production in white lupin

The influence, was investigated, of abiotic parameters on the isolation of protoplasts from in vitro seedling cotyledons of white lupin. The protoplasts were found to be competent in withstanding a wide range of osmotic potentials of the enzyme medium, however, −2.25 MPa (0.5 M mannitol), resulted in the highest yield of protoplasts. The pH of the isolation medium also had a profound effect on protoplast production. Vacuum infiltration of the enzyme solution into the cotyledon tissue resulted in a progressive drop in the yield of protoplasts. The speed and duration of orbital agitation of the cotyledon tissue played a significant role in the release of protoplasts and a two step (stationary-gyratory) regime was found to be better than the gyratory-only system.

A comparison of two time-domain analysis procedures in the determination of VO2 kinetics by pseudorandom binary sequence exercise testing

The purpose of this study was to apply and compare two time-domain analysis procedures in the determination of oxygen uptake (VO2) kinetics in response to a pseudorandom binary sequence (PRBS) exercise test. PRBS exercise tests have typically been analysed in the frequency domain. However, the complex interpretation of frequency responses may have limited the application of this procedure in both sporting and clinical contexts, where a single time measurement would facilitate subject comparison. The relative potential of both a mean response time (MRT) and a peak cross-correlation time (PCCT) was investigated. This study was divided into two parts: a test-retest reliability study (part A), in which 10 healthy male subjects completed two identical PRBS exercise tests, and a comparison of the VO2 kinetics of 12 elite endurance runners (ER) and 12 elite sprinters (SR; part B). In part A, 95% limits of agreement were calculated for comparison between MRT and PCCT. The results of part A showed no significant difference between test and retest as assessed by MRT [mean (SD) 42.2 (4.2) s and 43.8 (6.9) s] or by PCCT [21.8 (3.7) s and 22.7 (4.5) s]. Measurement error (%) was lower for MRT in comparison with PCCT (16% and 25%, respectively). In part B of the study, the VO2 kinetics of ER were significantly faster than those of SR, as assessed by MRT [33.4 (3.4) s and 39.9 (7.1) s, respectively; P<0.01] and PCCT [20.9 (3.8) s and 24.8 (4.5) s; P < 0.05]. It is possible that either analysis procedure could provide a single test measurement Of VO2 kinetics; however, the greater reliability of the MRT data suggests that this method has more potential for development in the assessment Of VO2 kinetics by PRBS exercise testing.

Alkaline phosphatase activity in pasteurized milk: a quantitative comparison of Fluorophos and colourimetric procedures

Fluorophos and colourimetric procedures for alkaline phosphatase (ALP) testing were compared using milk with raw milk additions, purified bovine ALP additions and heat treatments. Repeatability was between 0.9% and 10.1% for Fluorophos, 3.5% and 46.1% for the Aschaffenburg and Mullen (A&M) procedure and 4.4% and 8.8% for the Scharer rapid test. Linearity (R-2) using raw milk addition was 0.96 between Fluorophos and the Scharer procedure. Between the Fluorophos and the A&M procedures, R-2 values were 0.98, 0.99 and 0.98 for raw milk additions, bovine ALP additions and heat treatments respectively. Fluorophos showed greater sensitivity and was both faster and simpler to perform.