Screening,Molecular Cloning,and Biochemical Characterization of an Alcohol Dehydrogenase from Pichia pastoris Useful for the Kinetic Resolution of a Racemic β‐Hydroxy‐β‐trifluoromethyl Ketone

The stereoselective synthesis of chiral 1,3‐diols with the aid of biocatalysts is an attractive tool in organic chemistry. Besides the reduction of diketones, an alternative approach consists of the stereoselective reduction of β‐hydroxy ketones (aldols). Thus, we screened for an alcohol dehydrogenase (ADH) that would selectively reduce a β‐hydroxy‐β‐trifluoromethyl ketone. One potential starting material for this process is readily available by aldol addition of acetone to 2,2,2‐trifluoroacetophenone. Over 200 strains were screened, and only a few yeast strains showed stereoselective reduction activities. The enzyme responsible for the reduction of the β‐hydroxy‐β‐trifluoromethyl ketone was identified after purification and subsequent MALDI‐TOF mass spectrometric analysis. As a result, a new NADP⁺‐dependent ADH from Pichia pastoris (PPADH) was identified and confirmed to be capable of stereospecific and diastereoselective reduction of the β‐hydroxy‐β‐trifluoromethyl ketone to its corresponding 1,3‐diol. The gene encoding PPADH was cloned and heterologously expressed in Escherichia coli BL21(DE3). To determine the influence of an N‐ or C‐terminal His‐tag fusion, three different recombinant plasmids were constructed. Interestingly, the variant with the N‐terminal His‐tag showed the highest activity; consequently, this variant was purified and characterized. Kinetic parameters and the dependency of activity on pH and temperature were determined. PPADH shows a substrate preference for the reduction of linear and branched aliphatic aldehydes. Surprisingly, the enzyme shows no comparable activity towards ketones other than the β‐hydroxy‐β‐trifluoromethyl ketone.     

Waste disposal and waste avoidance

Waste disposal and waste avoidance are examined from two different perspectives, from the point of view of the waste 'producing' company on the one hand, and from the point of view of the waste disposing company on the other hand. The idea of the basic model lies in the 'turning upside down' of the lot-sizing approach to inventory control theory. We derive a corresponding formula for disposal that shows that the number of waste disposals depends positively on the waste accumulation rate and negatively on the fixed costs for a pick-up. Moreover, we formally work out at which waste avoidance costs a company will enlist the services of a disposal company. Specifically, disposal tends to be preferred to avoidance when waste-avoidance is relatively expensive and three cost rates to be specified are relatively low. Finally, we take the perspective of a monopolistic disposal company that has to decide on the optimal pricing policy. In our model, we determine that the price for a pick-up should be chosen just to cover costs. The disposal company can earn a profit by the suitable choice of the price per unit waste to be disposed of.     

Experimental and numerical analysis of seat belt bunching phenomenon

In current cars, loops are commonly used to redirect the webbing which reels out from the retractor to the passenger's shoulder. Some types of pillar loops, also called D-rings, lead to a non-systematic instability. The webbing, which should scroll without hindrance through the D-ring, laterally shifts, bunches and produces the overturning of the ring.     

Impact of selective oxidation during inline annealing prior to hot-dip galvanizing on Zn wetting and hydrogen-induced delayed cracking of austenitic FeMnC steel

The present study discusses the impact of selective oxidation during in-line annealing of Fe–23%Mn–0.6%C–0.3%Si steel on surface and sub-surface properties and is focused on hot-dip galvanizability and susceptibility to hydrogen-induced delayed cracking. Annealing temperature (700–1100 °C) and dewpoint DP (? 15/?30/?50 °C) of the 5%H₂–N₂ annealing atmosphere were varied in order to investigate Zn wetting in dependence on selective oxidation of Mn and Si. Sub-surface microplasticity (hardness, pop-in frequency, pop-in activation load) was examined by electrochemical nanoindentation in-situ to hydrogen charging (ECNI) to assess hydrogen/material interactions. Zn wetting fails if external Mn and Si oxidation is not avoided by performing high reductive bright annealing (1100 °C/DP ? 50 °C). Zn wetting will however turn to increase if a roughly globular MnO layer appears and Si is internally oxidized (700–900 °C/DP ? 15 °C). Selective oxidation further affects hydrogen/material interactions by influencing the local distribution of solid-soluted Mn: ECNI results indicate hydrogen-induced dislocation demobilization (HEDE mechanism) or dislocation mobilization (HELP mechanism) in dependence on the local amount of solid-soluted Mn within the sub-surface. Macroscopic delayed cracking seems to occur earlier if HELP is predominating. The gained results benefit understanding the impact of selective oxidation on galvanizability and susceptibility to hydrogen-induced failure of austenitic FeMnC steel and advance further developments in processing high Mn alloyed steels.