Immobilization of transaminase from Bacillus licheniformis on copper phosphate nanoflowers and its potential application in the kinetic resolution of RS-α-methyl benzyl amine

This study reports the isolation and partial purification of transaminase from the wild species of Bacillus licheniformis. Semi-purified transaminase was immobilized on copper nanoflowers (NFs) synthesized through sonochemical method and explored it as a nanobiocatalyst. The conditions for the synthesis of transaminase NFs [TA@Cu3(PO4)2NF] were optimized. Synthesized NFs revealed the protein loading and activity yield—60 ± 5% and 70 ± 5%, respectively. The surface morphology of the synthesized hybrid NFs was examined by scanning electron microscopy (SEM) and transmission electron microscopy (TEM), which revealed the average size to be around 1 ± 0.5 μm. Fourier-transform infrared (FTIR) was used to confirm the presence of the enzyme inside the immobilized matrix. In addition, circular dichroism and florescence spectroscopy were also used to confirm the integrity of the secondary and tertiary structures of the protein in the immobilized material. The transaminase hybrid NFs exhibited enhanced kinetic properties and stability over the free enzyme and revealed high reusability. Furthermore, the potential application of the immobilized transaminase hybrid NFs was demonstrated in the resolution of racemic α-methyl benzylamine. Graphical Abstract

Effect of Chlormequat Chloride on Vegetative Growth, Photosynthetic Activities, and its Residual Dissipation in Thompson Seedless Grapevines

Chlormequat chloride is one of the important plant growth regulators which is a highly stable gibberellins biosynthesis inhibitor used to inhibit vegetative growth and cell elongation. Considering the importance of plant growth inhibitors, the present investigation was carried out to understand the effect of Chlormequat chloride with its different concentrations on vegetative growth, photosynthetic activities and its residual dissipation in Grapevines at two different locations one at Pune, Maharashtra India and second at Nashik, Maharashtra, India during the year 2018-2019. Chlormequat chloride was applied as a foliar spray, where the whole vines were sprayed at concentration of 500, 1000, 1500, 2000 and 2500 ppm with untreated control as a water spray; as a treatments with different developmental stages Viz 5th, 7th, and 15th leaf stage after foundation pruning. Morphological observations were recorded at 45 days and 90 days after foundation as well as fruit pruning. Application of Chlormequat chloride recorded reduced shoot length and Internodal distance while increased in cane diameters, leaf thickness after both the pruning’s in this investigation Photosynthetic activity, yield per vines and percent fruitfulness had significantly influenced with the application of Chlormequat chloride in grapevines. The dissipation of Chlormequat chloride was stable and indicated a non- linear pattern of degradation. Thus, implied that simple first-order kinetics might not be adequate to explain the dissipation behaviour of Chlormequat chloride in grapes.

Exposures and behavioral responses to wildfire smoke

The impacts of environmental change on human outcomes often depend on local exposures and behavioral responses that are challenging to observe with traditional administrative or sensor data. We show how data from private pollution sensors, cell phones, social media posts, and internet search activity yield new insights on exposures and behavioral responses during large wildfire smoke events across the US, a rapidly-growing environmental stressor. Health-protective behavior, mobility, and sentiment all respond to increasing ambient wildfire smoke concentrations, but responses differ by income. Indoor pollution monitors provide starkly different estimates of likely personal exposure during smoke events than would be inferred from traditional ambient outdoor sensors, with similar outdoor pollution levels generating >20x differences in average indoor PM2.5 concentrations. Our results suggest that the current policy reliance on self protection to mitigate health risks in the face of rising smoke exposure will result in modest and unequal benefits.

Production of 6-l-[18F]Fluoro-m-tyrosine in an Automated Synthesis Module for 11C-Labeling

6-l-[18F]Fluoro-m-tyrosine (6-l-[18F]FMT) represents a valuable alternative to 6-l-[18F]FDOPA which is conventionally used for the diagnosis and staging of Parkinson’s disease. However, clinical applications of 6-l-[18F]FMT have been limited by the paucity of practical production methods for its automated production. Herein we describe the practical preparation of 6-l-[18F]FMT using alcohol-enhanced Cu-mediated radiofluorination of Bpin-substituted chiral Ni(II) complex in the presence of non-basic Bu4ONTf using a volatile iPrOH/MeCN mixture as reaction solvent. A simple and fast radiolabeling procedure afforded the tracer in 20.0 ± 3.0% activity yield within 70 min. The developed method was directly implemented onto a modified TracerLab FX C Pro platform originally designed for 11C-labeling. This method enables an uncomplicated switch between 11C- and 18F-labeling. The simplicity of the developed procedure enables its easy adaptation to other commercially available remote-controlled synthesis units and paves the way for a widespread application of 6-l-[18F]FMT in the clinic.

The effects of molar activity on [18F]FDOPA uptake in patients with neuroendocrine tumors

Background 6-[18F]fluoro-l-3,4-dihydroxyphenyl alanine ([18F]FDOPA) is a commonly used PET tracer for the detection and staging of neuroendocrine tumors. In neuroendocrine tumors, [18F]FDOPA is decarboxylated to [18F]dopamine via the enzyme amino acid decarboxylase (AADC), leading to increased uptake when there is increased AADC activity. Recently, in our hospital, a new GMP compliant multi-dose production of [18F]FDOPA has been developed, [18F]FDOPA-H, resulting in a higher activity yield, improved molar activity and a lower administered mass than the conventional method ([18F]FDOPA-L). Aims This study aimed to investigate whether the difference in molar activity affects the [18F]FDOPA uptake at physiological sites and in tumor lesions, in patients with NET. It was anticipated that the specific uptake of [18F]FDOPA-H would be equal to or higher than [18F]FDOPA-L. Methods We retrospectively analyzed 49 patients with pathologically confirmed NETs and stable disease who underwent PET scanning using both [18F]FDOPA-H and [18F]FDOPA-L within a time span of 5 years. A total of 98 [18F]FDOPA scans (49 [18F]FDOPA-L and 49 [18F]FDOPA-H with average molar activities of 8 and 107 GBq/mmol) were analyzed. The SUVmean was calculated for physiological organ uptake and SUVmax for tumor lesions in both groups for comparison, and separately in subjects with low tumor load (1–2 lesions) and higher tumor load (3–10 lesions). Results Comparable or slightly higher uptake was demonstrated in various physiological uptake sites in subjects scanned with [18F]FDOPA-H compared to [18F]FDOPA-L, with large overlap being present in the interquartile ranges. Tumor uptake was slightly higher in the [18F]FDOPA-H group with 3–10 lesion (SUVmax 6.83 vs. 5.19, p < 0.001). In the other groups, no significant differences were seen between H and L. Conclusion [18F]FDOPA-H provides a higher activity yield, offering the possibility to scan more patients with one single production. Minor differences were observed in SUV’s, with slight increases in uptake of [18F]FDOPA-H in comparison to [18F]FDOPA-L. This finding is not a concern for clinical practice, but could be of importance when quantifying follow-up scans while introducing new production methods with a higher molar activity of [18F]FDOPA.

The Effects of Molar Activity on [18f]FDOPA Uptake in Patients With Neuroendocrine Tumors.

Background6-[18F]fluoro-L-3,4-dihydroxyphenyl alanine ([18F]FDOPA) is a commonly used PET tracer for the detection and staging of neuroendocrine tumors. In neuroendocrine tumors. [18F]FDOPA is decarboxylated to [18F]dopamine via the enzyme amino acid decarboxylase (AADC), leading to increased uptake when there is increased AADC activity. Recently In our hospital, a new GMP compliant multi-dose production of [18F]FDOPA has been developed, [18F]FDOPA-H, resulting in a higher activity yield, improved molar activity and a lower administered mass than the conventional method ([18F]FDOPA-L). AimThis study aimed to investigate whether the difference in molar activity affects the [18F]FDOPA uptake at physiological sites and in tumor lesions, in patients with NET. It was anticipated that the specific uptake of [18F]FDOPA-H would be equal to or higher than [18F]FDOPA-L.Methods We retrospectively analyzed 50 patients with pathologically confirmed NETs and stable disease who underwent PET scanning using both [18F]FDOPA-H and [18F]FDOPA-L within a time span of 5 years. A total of 100 [18F]FDOPA scans (50 [18F]FDOPA-L and 50 [18F]FDOPA-H with average molar activities of 8 and 107 GBq/mmol) were analyzed. The SUVmean was calculated for physiological organ uptake and SUVmax for tumor lesions in both groups for comparison, and separately in subjects with low tumor load (1-2 lesions) and higher tumor load (3-10 lesions). ResultsComparable or slightly higher uptake was demonstrated in various physiological uptake sites in subjects scanned with [18F]FDOPA-H compared to [18F]FDOPA-L, with large overlap being present in the interquartile ranges. Tumor uptake was slightly higher in the [18F]FDOPA-H group with 3-10 lesion (SUVmax 6.83 vs. 5.19, p<0.001). In the other groups, no significant differences were seen between H and L.Conclusion[18F]FDOPA-H provides a higher activity yield, offering the possibility to scan more patients with one single production. Minor differences were observed in SUV’s, with slight increases in uptake of [18F]FDOPA-H in comparison to [18F]FDOPA-L. This finding is not an concern for clinical practice, but could be of importance when quantifying follow-up scans while introducing new production methods with a higher molar activity of [18F]FDOPA

Immobilization of Phenylalanine Ammonia-lyase via EDTA Based Metal Chelate Complexes – Optimization and Prospects

Immobilized metal ion affinity chromatography principles were applied for selective immobilization of recombinant polyhistidine tag fused phenylalanine ammonia-lyase from parsley (PcPAL) on porous polymeric support with aminoalkyl moieties modified with an EDTA dianhydride (EDTADa)-derived chelator and charged with transition metal ions. Out of the five investigated metal ions - Fe3+, Co2+, Ni2+, Cu2+, Zn2+ - the best biocatalytic activity of PcPAL was achieved when the enzyme was immobilized on the Co2+ ion-charged support (31.8 ± 1.2 U/g). To explore the features this PcPAL obtained by selective immobilization, the thermostability and reusability of this PAL biocatalyst were investigated. To maximize the activity of the immobilized PcPAL the surface functionalization of the aminoalkylated polymeric carrier was fine-tuned with using glycidol as a thinning group beside EDTADa. The maximal activity yield (YA=103 %) was earned when the EDTADa and glycidol were used in 1 to 24 ratio. The reversibility of the immobilization method allowed the development of a support regeneration protocol which enables easy reuse of the functionalized support in case of enzyme inactivation.

Scalable 18F Processing Conditions for Copper-Mediated Radiofluorination Chemistry Facilitate "Design of Experiments" (DoE) Optimization Studies and Afford an Improved Synthesis of [18F]Olaparib.

A convenient, scalable, and azeotropic drying free method for processing [18F]fluoride as base free<br>[18F]TBAF is reported and applied to copper-mediated radiofluorination (CMRF) radiosyntheses. A central<br>feature of this method is that a single production of [18F]TBAF can be divided into small aliquots that can be<br>used to perform multiple small-scale reactions in DoE optimization studies. The results of these studies can<br>then be reliably translated to full batch tracer productions using automated synthesizers. This processing<br>technique was successfully applied to the manual DoE optimization, DoE study validation, and subsequent<br>full-batch automation of the PARP-1 tracer [18F]olaparib. After DoE optimization, we were able to produce<br>[18F]olaparib in high radiochemical yields via both manual (%RCY (CMRF step only) = 78 ± 6 %, n = 4) and<br>automated (up to 80% radiochemical yield (%RCY); 41% activity yield (%AY)) radiosynthesis procedures.<br>This work further demonstrates the power of the DoE approach for improving the radiochemical yields and<br>radiosynthesis performance of clinically relevant tracer productions

Scalable 18F Processing Conditions for Copper-Mediated Radiofluorination Chemistry Facilitate "Design of Experiments" (DoE) Optimization Studies and Afford an Improved Synthesis of [18F]Olaparib.

A convenient, scalable, and azeotropic drying free method for processing [18F]fluoride as base free<br>[18F]TBAF is reported and applied to copper-mediated radiofluorination (CMRF) radiosyntheses. A central<br>feature of this method is that a single production of [18F]TBAF can be divided into small aliquots that can be<br>used to perform multiple small-scale reactions in DoE optimization studies. The results of these studies can<br>then be reliably translated to full batch tracer productions using automated synthesizers. This processing<br>technique was successfully applied to the manual DoE optimization, DoE study validation, and subsequent<br>full-batch automation of the PARP-1 tracer [18F]olaparib. After DoE optimization, we were able to produce<br>[18F]olaparib in high radiochemical yields via both manual (%RCY (CMRF step only) = 78 ± 6 %, n = 4) and<br>automated (up to 80% radiochemical yield (%RCY); 41% activity yield (%AY)) radiosynthesis procedures.<br>This work further demonstrates the power of the DoE approach for improving the radiochemical yields and<br>radiosynthesis performance of clinically relevant tracer productions

Fully Automated Synthesis of Novel TSPO PET Imaging Ligand [18F]Fluoroethyltemazepam

Introduction: Benzodiazepines, including temazepam are described as TSPO antagonists. In fact, TSPO was initially described as a peripheral benzodiazepine receptor (PBR) with a secondary binding site for diazepam. TSPO is a potential imaging target of neuroinflammation because there is an amplification of the expression of this receptor. Objectives: Herein, we developed a novel fluorinated benzodiazepine ligand, [18F]Fluoroethyltemazepam ([18F]F-FETEM), for positron emission tomography (PET) imaging of translocator protein (18 kDa). Methods: [18F]F-FETEM was radiolabelled with an automated synthesizer via a one-pot procedure. We conducted a [18F]F-aliphatic nucleophilic substitution of a tosylated precursor followed by purification on C18 and Alumina N SPE cartridges. Quality control tests was also carried out. Results: We obtained 2.0–3.0% decay-uncorrected radiochemical activity yield (3.7% decay-corrected) within the whole synthesis time about 33 min. The radiochemical purity of [18F]F-FETEM was over 90% by TLC analysis. Conclusion: This automated procedure may be used as basis for future production of [18F]F-FETEM for preclinical PET imaging studies.