Statistical Evaluation of HTS Assays for Enzymatic Hydrolysis of ß-Keto Esters.

ß-keto esters are used as precursors for the synthesis of ß-amino acids, which are building blocks for some classes of pharmaceuticals. Here we describe the comparison of screening procedures for hydrolases to be used for the hydrolysis of ß-keto esters, the first step in the preparation of ß-amino acids. Two of the tested high throughput screening (HTS) assays depend on coupled enzymatic reactions which detect the alcohol released during ester hydrolysis by luminescence or absorption. The third assay detects the pH shift due to acid formation using an indicator dye. To choose the most efficient approach for screening, we assessed these assays with different statistical methods-namely, the classical Z'-factor, standardized mean difference (SSMD), the Kolmogorov-Smirnov-test, and t-statistics. This revealed that all three assays are suitable for HTS, the pH assay performing best. Based on our data we discuss the explanatory power of different statistical measures. Finally, we successfully employed the pH assay to identify a very fast hydrolase in an enzyme-substrate screening.

Injuries in competitive boxing. A prospective study.

Boxing remains a subject of controversy and is often classified as dangerous. But the discussion is based mostly on retrospective studies. This survey was conducted as a prospective study. From October 2012 to September 2013, 44 competitive boxers were asked to report their injuries once a month. The questionnaire collected general information (training, competition) and recorded the number of bouts fought, injuries and resulting lost days. A total of 192 injuries were recorded, 133 of which resulted in interruption of training or competition. Each boxer sustained 3 injuries per year on average. The injury rate was 12.8 injuries per 1 000 h of training. Boxers fighting more than 3 bouts per year sustain more injuries (p=0.0075). The injury rate does is not a function of age (age≤19 vs. > 19a, p=0.53). Injuries to the head and the upper limbs occur most frequently. The most common injuries are soft tissue lacerations and contusions. Head injuries with neurological symptoms rarely occur (4.2%). Boxing has a high injury rate that is comparable with other contact sports, but most injuries are minor. Injury frequency is not a function of whether the boxer competes in the junior or adult category. Athletes fighting many bouts per year have a greater risk of injury.

Stereoselective hydrolysis of aryl-substituted dihydropyrimidines by hydantoinases.

In this study, we investigated the possibility of using a modified hydantoinase process for the production of optically pure ß-amino acids. Two aryl-substituted dihydropyrimidines D,L-6-phenyl-5,6-dihydrouracil (PheDU) and para-chloro-D,L-6-phenyl-5,6-dihydrouracil (pClPheDU) were synthesized. Hydrolysis of these novel substrates to the corresponding N-carbamoyl-ß-amino acids by three recombinant D-hydantoinases and several bacterial strains was tested. All applied recombinant D-hydantoinases and eight bacterial isolates catalyzed the conversion of PheDU to N-carbamoyl-ß-phenylalanine (NCßPhe). Some of these biocatalysts showed an enantioselectivity for either the D- or the L-PheDU enantiomer. The second dihydropyrimidinase substrate pClPheDU was hydrolyzed by all three recombinant D-hydantoinases and six of the wild-type strains. To our knowledge, this is the first dihydropyrimidinase activity reported with this aryl-substituted dihydropyrimidine. For selected biocatalysts, hydantoinase activity towards aryl-substituted hydantoins was demonstrated as well. However, none of the bacterial strains tested so far exhibited any carbamoylase activity towards NCßPhe.

Injuries and overuse syndromes in powerlifting.

Powerlifting is a discipline of competitive weightlifting. To date, no investigations have focused on pain encountered during routine training. The aim of the study was to identify such pain, assign it to particular exercises and assess the data regarding injuries as well as the influence of intrinsic and extrinsic factors. Data of 245 competitive and elite powerlifters was collected by questionnaire. Information regarding current workout routines and retrospective injury data was collected. Study subjects were selected from 97 incorporated powerlifting clubs. A percentage of 43.3% of powerlifters complained of problems during routine workouts. Injury rate was calculated as 0.3 injuries per lifter per year (1 000 h of training=1 injury). There was no evidence that intrinsic or extrinsic factors affected this rate. Most commonly injured body regions were the shoulder, lower back and the knee. The use of weight belts increased the injury rate of the lumbar spine. Rate of injury to the upper extremities was significantly increased based on age >40 years (shoulder/p=0.003, elbow/p=0.003, hand+wrist/p=0.024) and female gender (hand+wrist/p=0.045). The daily workout of a large proportion of powerlifters is affected by disorders which do not require an interruption of training. The injury rate is low compared to other sports.