Improved synthesis of polyfluorinated l-lysine for 19F NMR-based screening

Polyfluorinated N-α-Fmoc--Boc-l-lysine represents the best-in-class among a set of polyfluorinated amino acids (PFAs) which are useful tools for ¹⁹F NMR-Based Screening. In this communication, optimized reaction conditions that allowed for the multi-gram preparation of this unnatural amino acid are reported.     

Microwave-assisted organocatalytic multicomponent Knoevenagel/hetero Diels-Alder reaction for the synthesis of 2,3-dihydropyran[2,3-c]pyrazoles

A rapid protocol for the multicomponent microwave-assisted organocatalytic domino Knoevenagel/hetero Diels-Alder reaction (DKHDA) has been developed for the synthesis of 2,3-dihydropyran[2,3-c]pyrazoles. The reported procedure could be used for the fast generation of novel substituted 2,3-dihydropyran[2,3-c]pyrazoles with potential anti-tuberculosis activity.     

LC Analysis of Histamine and Other Biogenic Amines in Digesta Cultures

In the intestine, microbial protein degradation to histamine and other biologically active amines is believed to be relevant to the health of species. An LC method with fluorescence detection has been developed for analysis of histamine, phenylethylamine, isoamylamine, putrescine, cadaverine, tyramine, and spermidine. After pre-column derivatisation with 9-fluorenylmethyl chloroformate, amines were separated on a reversed-phase C₁₈ polyamine column with a mobile phase gradient. Validation of the method included calibration experiments, determination of amine recovery, and repeatability tests. The method proved especially useful for detection of histamine, phenylethylamine, putrescine, and cadaverine at relevant concentrations in caecum cultures from horses. The limits of quantification for these four amines ranged between 80 nM (histamine) and 400 nM (phenylethylamine).     

Conformationally Homogeneous Heterocyclic Pseudotetrapeptides as Three‐Dimensional Scaffolds for Rational Drug Design: Receptor‐Selective Somatostatin Analogues

A would‐be amide : A 1,4‐disubstituted 1,2,3‐triazole was used as a surrogate for a trans amide bond to create a library of 16 diastereomeric pseudotetrapeptides as β‐turn mimetics. High‐resolution structural analysis indicated that these scaffolds adopt distinct, rigid, conformationally homogeneous β‐turn‐like structures (see example), some of which bind somatostatin receptor subtypes selectively, and some of which show broad‐spectrum activity.

     

Photoconvertible fluorescent protein EosFP: biophysical properties and cell biology applications

EosFP is a fluorescent protein from the coral Lobophyllia hemprichii that changes its fluorescence emission from green to red upon irradiation with near-UV light. Here we present the spectroscopic properties of wild-type EosFP and a variety of monomeric and dimeric mutants and provide a structural interpretation of its oligomerization and photoconversion, which is based on X-ray structure analysis of the green and red species that we reported recently. Because functional expression of the monomeric EosFP variant is limited to temperatures of 30 degrees C, we have developed a tandem dimer. This construct, in which two EosFP subunits are connected by a flexible 12 amino acid linker, expresses well after fusion with the androgen and endothelin A receptors at 37 degrees C. A variety of applications in cellular imaging, developmental biology and automated high-content screening applications are presented, which demonstrate that EosFP is a powerful tool for in vivo monitoring of cellular processes.     

Cryopreservation of cultivated and wild potato varieties by droplet vitrification: effect of subculture of mother-plants and of preculture of shoot tips

In this paper, we studied the effect of subculture of mother-plants and of preculture of shoot tips of two potato varieties (Dejima, cultivated and STN13, wild) cryopreserved using the droplet-vitrification technique. The subculture conditions (light intensity, aeration and planting density) significantly affected survival of both non-cryopreserved and cryopreserved shoot-tips in both varieties. The subculture duration and the position of the shoot tips on the axis of the in vitro plantlets had a significant (P<0.0001) effect on survival of cryopreserved shoot tips. The optimal subculture duration was 7 and 5 weeks and the optimal size of shoot tips was 1.5-2.0 and 1.0-1.5 mm for var. Dejima and STN13, respectively. Survival of cryopreserved shoot tips was influenced by the sucrose concentration in the preculture medium and the preculture duration. The highest survival of cryopreserved shoot tips was observed after preculture with 0.3 M sucrose for 8 h followed by 0.7 M sucrose for 18 h. These results indicate that the parameters of the subculture of mother-plants and of preculture of shoot tips should be carefully optimized, especially in the case of wild species.     

New determinants for tolerance of coffee (Coffea arabica L.) seeds to liquid nitrogen exposure

The present work establishes for the first time that tolerance of coffee seeds to liquid nitrogen (LN) exposure depends on the initial quality of the seedlot and on the rewarming regime employed. Seedlot quality was estimated by the parameters of a quantal response model of desiccation sensitivity developed previously. The percentage of seedlings recovered from cryopreserved seeds was very well correlated with the relative humidity (RH) at which 90 percent of the initial viability was retained, RH90, as estimated by the model. Whatever the cooling regime employed, rewarming the seeds slowly by exposing them to ambient air was highly detrimental. Slow rewarming-induced viability loss was not due to imbibitional damage since seeds pre-heated at 37 degree C after slow rewarming to 0 degree C exhibited a survival percentage lower than seeds thawed rapidly to 0 degree C before sowing. The optimal hydration status for coffee seed cryopreservation was also re-examined. Drying seeds in 81 percent RH provided survival percentages considerably higher than those obtained using the drying RH always employed until now, i.e. 78 percent. A new procedure for slowly precooling the seeds prior to immersion in LN was also established. It consisted of placing the vials containing the seeds in a dry ice-bath for 25 min. Using this procedure in combination with seed drying in 81 percent RH and rapid rewarming in a 37 degree C water-bath for 30 min ensured the highest survival percentages ever obtained with coffee seeds, i.e. 89 percent, a value which was not significantly different from the initial viability percentage.     

Cryopreservation of garlic bulbil primordia by the droplet-vitrification procedure

The droplet-vitrification protocol, a combination of droplet-freezing and solution-based vitrification was applied for cryopreserving garlic bulbil primordia. The highest survival and regeneration percentages of cryopreserved primordia (90.1 to 95.0 percent and 82.7 to 85.0 percent, respectively) were achieved after preculture for 2-4 days at 10 degree C on solid medium with 0.1 - 0.3 M sucrose, loading for 50 minutes in liquid medium with 2 M glycerol + 0.5 M sucrose, dehydration with PVS3 vitrification solution for 90-150 min, cooling primordia in 5 microl droplets of PVS3 vitrification solution placed on aluminum foil strips by dipping these strips in liquid nitrogen, warming them by plunging the foil strips into pre-heated (40 degree C) 0.8 M sucrose solution for 30 s and further incubation in the same solution for 30 minutes. The optimized droplet-vitrification protocol was successfully applied to bulbil primordia of five garlic varieties originating from various countries and to immature bulbils of two vegetatively propagated Allium species, with regeneration percentages ranging between 77.4 - 95.4 percent.     

Relative angular precision in electron backscatter diffraction: A comparison between cross correlation and Hough transform based analysis

In the present study the relative angular resolution of an electron backscatter diffraction system based on Hough transform analysis has been determined with a silicon single crystal wafer. The resolution is found to be better than 0.1° and can be easily improved by repetition of measurements. A test measurement on a BaFe₂As₂ thin film, where disorientations of 0.1° and less are present, was performed using the cross correlation electron backscatter diffraction technique. The same measurement is evaluated with the Hough transform technique. Comparing both techniques give evidence of a relative resolution of better than 0.1°. However, in specimen areas with strain inhomogeneities a deviation along one rotation axis can be observed.