Despite significant differences in size and formal charge, the aldehyde dehydrogenase PaPauC (PA5312) from Pseudomonas aeruginosa PAO1 efficiently catalyzes the NAD+-dependent oxidation of the aminoaldehydes formed in polyamines degradation. We report here that PaPauC also oxidizes 4-guanidinebutyraldehyde, formed in one arginine degradation pathway, trimethylaminobutyraldehyde, of unknown metabolic origin, and indole-3-acetaldehyde, a precursor of the plant growth-promoting hormone indoleacetic acid. PaPauC has been proposed as a potential target for combating P. aeruginosa. However, understanding its structure-function relationships, crucial for developing specific inhibitors, is lacking. Using X-ray crystallography, we identified the structural characteristics that determine PaPauC broad aldehyde specificity: a spacious aldehyde-entrance tunnel and six active-site residues. Docking simulations, site-directed mutagenesis, and kinetic analyses support the interactions of Lys479 with glutamylated aminoaldehydes; Phe169, Trp176, and Phe467 with amino and guanidinium groups through cation-π interactions and with the indole group via NH-π and CH-π interactions; Asp459 with amino and indole groups; and Thr303 with amide and guanidinium groups. Exploiting the distinctive structural features of the PaPauC active site could aid in developing specific inhibitors to combat P. aeruginosa infections in humans and animals, as well as in preventing its colonization of plants, which are abundant P. aeruginosa reservoirs and, therefore, a significant source of human infections.

In the present study the interaction of cannabinoid, PhAR-DBH-Me [(R, Z)-18-((1S,4S)-5-methyl-2,5-diazabicyclo[2.2.1]heptan-2-yl)-18-oxooctadec-9-en-7-ylphenyl-acetate] and tramadol in two neuropathy models, as well as their possible toxic effects, was analyzed. The anti-allodynic effect of PhAR-DBH-Me, tramadol, or their combination, were evaluated in neuropathic rats. Furthermore, the effective dose 35 (as the 35 % of the anti allodynic effect) was calculated from the maximum effect of each drug. Moreover, the isobolographic analysis was performed to determine the type of interaction between the drugs. A plasma acute toxicity study was carried out to assess the hepatic, renal, and heart functions after an individual or combined administration of the drugs, as well as histology using the hematoxylin-eosin or Masson-trichome method. PhAR-DBH-Me, tramadol, and their combination produced an antiallodynic effect on spinal nerve ligation (SNL) and cisplatin-induced neuropathic pain in rats. Moreover, PhAR-DBH-Me and tramadol combination showed a synergistic interaction in neuropathic pain rats induced by SNL but not for cisplatin-induced neuropathy. On the other hand, changes in renal and hepatic functions were not observed. Likewise, analysis of liver, kidney and heart histology showed no alterations compared with controls. Results show that the combination of PhAR-DBH-Me and tramadol attenuates the allodynia in SNL rats; the acute toxicology analysis suggests that this combination could be considered safe in administered doses.

The effect of PHAR-DBH-Me, a cannabinoid receptor agonist, on different cardiovascular responses in adult male rats was analyzed. The blood pressure was measured directly and indirectly. The coronary flow was measured by Langendorff preparation, and vasomotor responses induced by PHAR-DBH-Me in aortic rings precontracted with phenylephrine (PHEN) were analyzed. The intravenous injection of the compound PHAR-DBH-Me (0.018-185 μg/kg) resulted in decreased blood pressure; maximum effect was observed at the dose of 1,850 μg/kg. A concentration-dependent increase in the coronary flow was observed in a Langendorff preparation. In the aortic rings, with and without endothelium, pre-contracted with PHEN (10-6 M), the addition of PHAR-DBH-Me to the superfusion solution (10-12-10-5 M), produced a vasodilator response, which depends on the concentration and presence of the endothelium. L-NAME inhibited these effects. Addition of CB1 receptor antagonist (AM 251) did not modify the response, while CB2 receptor antagonist (AM630) decreased the potency of relaxation elicited by PHAR-DBH-Me. Indomethacin shifted the curve concentration-response to the left and produced an increase in the magnitude of the maximum endothelium dependent response to this compound. The maximum effect of PHAR-DBH-Me was observed with the concentration of 10-5 M. These results show that PHAR-DBH-Me has a concentration-dependent and endothelium-dependent vasodilator effect through CB2 receptor. This vasodilation is probably mediated by the synthesis/release of NO. On the other hand, it is suggested that PHAR-DBH-Me also induces the release of a vasoconstrictor prostanoid.

Safety studies are essential in drug development. This study evaluates the safety of Amphotericin A21 (AmB-A21), a derivative of amphotericin B with antifungal therapeutic potential. We performed a chronic toxicity study, a targeted organ study and a dermal irritation test. To evaluate chronic toxicity, 18 male adult rats were treated orally with AmB-21 (2 mg/kg) for 26 weeks. The effects on body-weight and animal health were measured, and haematological, clinical chemistry and histopathological tests were conducted on various organs. In the target organ toxicity study, male adult rats received a daily oral dose of AmB-21 (2 mg/kg) for 6 and 17 weeks; testicle histology and testosterone levels were then evaluated. For the dermal irritation study, AmB-21 (200 and 1000 mg/kg) was placed on the skin of adult male rabbits; macroscopic and microscopic studies, as well as haematological and clinical chemistry tests were then conducted. The chronic toxicity study revealed that AmB-21 caused testicle damage, and the testicle-targeted study showed structural alterations and changes in testosterone levels at 17 weeks. However, these alterations were no longer observed 8 weeks after discontinuation of treatment, and the testes showed very similar characteristics to those in the control group. The dermal irritation study showed skin thickening and reddening in rabbits treated with 2000 mg of AmB-A21 after 14 days of exposure. This same group also showed changes in liver enzymes, renal parameters and platelet levels. Based on our results, we consider AmB-21 to be a potential candidate for safe, long-term antifungal treatment given its reduced side effects.

The opportunistic human pathogen Pseudomonas aeruginosa exhibits great resistance to antibiotics; so, new therapeutic agents are urgently needed. Since polyamines levels are incremented in infected tissues, we explored whether the formation of a toxic aldehyde in polyamines degradation can be exploited in combating infection. We cloned the gene encoding the only aminoaldehyde dehydrogenase involved in P. aeruginosa polyamines-degradation routes, PaPauC, overexpressed this enzyme, and found that it oxidizes 3-aminopropionaldehyde (APAL) and 3-glutamyl-3-aminopropionaldehyde (GluAPAL) - produced in spermine (Spm), spermidine (Spd), and diaminopropane (Dap) degradation, as well as 4-aminobutyraldehyde (ABAL) and 4-glutamyl-4-aminobutyraldehyde (GluABAL) - formed in putrescine (Put) degradation. As the catalytic efficiency of PaPauC with APAL was 30-times lower than with GluAPAL, and GluAPAL is predominantly formed, APAL will be poorly oxidized 'in vivo'. We found polyamines-induced increases in the PaPauC activity of cell crude-extracts and in the expression of the PapauC gene that were diminished by glucose. Spm, Spd, or Dap, but not Put, were toxic to P. aeruginosa even in the presence of other carbon and nitrogen sources, particularly to a strain with the PapauC gene disrupted. APAL, but not GluAPAL, was highly toxic even to wild-type cells, suggesting that its accumulation, particularly in the absence of, or low, PaPauC activity is responsible for the toxicity of Spm, Spd, and Dap. Our results shed light on the toxicity mechanism of these three polyamines and strongly support the critical role of PaPauC in this toxicity. Thus, PaPauC emerges as a novel potential drug target whose inhibition might help in combating infection by this important pathogen.

The antiallodynic effect of PhAR-DBH-Me was evaluated on two models of neuropathic pain, and the potential roles of CB1, CB2, and TRPV1 receptors as molecular targets of PhAR-DBH-Me were studied. Female Wistar rats were submitted to L5/L6 spinal nerve ligation (SNL) or repeated doses of cisplatin (0.1 mg/kg, i.p.) to induce experimental neuropathy. Then, tactile allodynia was determined, and animals were treated with logarithmic doses of PhAR-DBH-Me (3.2-100 mg/kg, i.p.). To evaluate the mechanism of action of PhAR-DBH-Me, in silico studies using crystallized structures of CB1, CB2, and TRPV1 receptors were performed. To corroborate the computational insights, animals were intraperitoneally administrated with antagonists for CB1 (AM-251, 3 mg/kg), CB2 (AM-630, 1 mg/kg), and TRPV1 receptors (capsazepine, 3 mg/kg), 15 min before to PhAR-DBH-Me (100 mg/kg) administration. Vagal stimulation evoked on striated muscle contraction in esophagus, was used to elicited pharmacological response of PhAR-DBH-ME on nervous tissue. Systemic administration of PhAR-DBH-Me reduced the SNL- and cisplatin-induced allodynia. Docking studies suggested that PhAR-DBH-Me acts as an agonist for CB1, CB2, and TRPV1 receptors, with similar affinity to the endogenous ligand anandamide. Moreover antiallodynic effect of PhAR-DBH-Me was partially prevented by administration of AM-251 and AM-630, and completely prevented by capsazepine. Finally, PhAR-DBH-Me decreased the vagally evoked electrical response in esophagus rat. Taken together, results indicate that PhAR-DBH-Me induces an antiallodynic effect through partial activation of CB1 and CB2 receptors, as well as desensitization of TRPV1 receptors. Data also shed light on the novel vanilloid nature of the synthetic compound PhAR-DBH-Me.

Introduction: Cardiovascular effects of endocannabinoids (eCBs) have generated considerable interest since it has been suggested that the eCB system could become the new pharmacological target, either by blocking its activity or by promoting its effects on several cardiovascular diseases such as hypovolemic and septic shock or hypertension. The purpose of this study was to examine the effects of oleamide on several vasomotor responses in adult rats. Materials and Methods: Blood pressure (BP) was measured both directly and indirectly. Coronary flow was quantified with Langendorf preparation, and the vasomotor responses induced by oleamide were analyzed in the aortic rings. Results: Oleamide induced a decrease in BP, by both direct and indirect methods, which were dose dependent. An increase in coronary flow was observed with Langendorf preparation depending on the dose. Oleamide produced a vasodilator response in aortic rings pre-contracted with phenylephrine (10-5 M), which was concentration and endothelium dependent. This relaxing effect was of minor magnitude than that induced with the same dose on BP. L-NAME did not modify these effects. However, indomethacin induced a shift to the left of the concentration-response curve to oleamide and an increase in the magnitude of maximum vasodilation in rings with endothelium. Oleamide produced the maximal relaxant response at 10-5 M concentration. Conclusions: Oleamide has both in vivo and in vitro vasodilator effects. Vasodilator effects could be mediated by compounds synthesized/released by the endothelium (hyperpolarizing factor) or acting directly on vascular smooth muscle in aortic rings. The TRPV1 and CB1R receptors could mediate these effects. Finally, the results suggest that oleamide probably induces the synthesis/release of a vasoconstrictor prostanoid.

(−)-Sparteine sulfate is a stable salt of the quinolizidine chiral diamine alkaloid (−)-sparteine. This compound, in addition to presenting various biological activities, has successfully been used as a ligand and organocatalyst in enantioselective lithiation reactions in asymmetric synthesis. (−)-Sparteine free base deteriorates gradually, even in refrigeration, but its stable salt, (−)-sparteine sulfate, offers advantages in handling and storage. To the best of our knowledge, no process exists in the literature for obtaining (−)-sparteine sulfate from the species of the plant genus Lupinus. In this work, we present an efficient and sustainable process to obtain (−)-sparteine sulfate and free base (−)-sparteine from leaves and stems of Lupinus montanus.

The ALDH27 family of the aldehyde dehydrogenase superfamily comprises bacterial enzymes poorly studied so far in spite of seemingly being widely distributed, and of their putative important metabolic roles. With the double aim of exploring their occurrence within bacteria and investigating their physiological functions, we performed comprehensive phylogenetic and genomic context analyses. We also biochemically and structurally characterized the three ALDH27 enzymes present in Pseudomonas aeruginosa PAO1 and tested the growth of PAO1 mutant strains on compounds that produce the putative substrate aldehydes. We retrieved 245 non‐redundant ALDH27 sequences from 149 reference genomes, most of them from αβγ‐proteobacteria, some from actinobacteria and a few from firmicutes. These sequences formed two subfamilies: ALDH27A and ALDH27B, which in turn consists of three clades. The neighboring gene of the aldh27A locus is a class I aldolase not characterized so far. Therefore, we could not infer the physiological role of these enzymes from their genomic context. The genomic contexts of the aldh27B1 and aldh27B3 are consistent with their participation in polyamines catabolism. In the neighborhood of the aldh27B2 locus there are genes encoding for either histamine dehydrogenase or histamine‐pyruvate aminotransferase, suggesting their involvement in histamine catabolism. The in vitro and in vivo specificity of PA5312, which belongs to the ALDH27B3 clade, confirmed that it is essential for the catabolism of agmatine, putrescine, cadaverine, spermine, spermidine, and diaminopropane. Similar studies showed that PA0219, which belongs to the ALDH27B2 clade, is essential for the catabolism of histamine. PA4189, which belongs to the ALDH27A subfamily, oxidizes most of the aminoaldehydes substrates of PA5312 and PA0219 enzymes, but they are not its physiological substrates as the PAO1 mutant strain lacking PA4189 activity grew in polyamines or histamine. To find out the structural basis of aldehyde specificity, we obtained the three‐dimensional structures of PA4189 and PA5312 by X‐ray crystallography and performed docking simulations with aldehydes that in vitro are substrates of these enzymes. Then, we examined the conservation within each subfamily and clade of the active site residues in critical positions, and found highly conserved acidic residues, aspartate or glutamate, which may be important for binding the aldehyde substrate. Together, our results provide insights into the diversification of the ALDH27 family in bacteria and of the physiological functions of the members of this family.Support or Funding InformationFinancially supported by DGAPA‐UNAM (PAPIIT IN220317) and CONACYT 283524 grants to RAMC, and DGAPA‐UNAM (PAPIIT IN218819) grant to HRR. ALJV is a recipient of a postdoctoral scholarship from DGAPA‐UNAM and YVCC of a doctoral scholarship from CONACYT.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

The present study seeks to describe the design and synthesis of six new Michael adducts of (1S,4S)-2,5-diazabicyclo[2.2.1]heptane-dithiocarbamate with nitrostyrenes and their in vitro antiproliferative activity against human cervical cancer cell lines [HeLa (HPV 18 positive), CaSki (HPV 16 positive) and ViBo (HPV negative) cervical cancer cell lines]. Virtual screening of the physicochemical properties of all compounds have also been presented. All the compounds exploited significant antiproliferative activity on the three cervical cancer cell lines. Compound 8a was found to be most potent, displaying in vitro antiproliferative activity against HeLa, CaSki and ViBo cervical cancer cell lines superior to Cisplatin and Paclitaxel with IC50 values 0.99 ± 0.007, 2.36 ± 0.016 and 0.73 ± 0.002 μM respectively. In addition, compound 8a did not trigger the necrosis cell death to the test cancer cell lines. Further mechanistic study revealed that compound 8a could inhibit the cancer cell proliferation by inducing apoptosis through caspase-3 activation. Moreover, cell cycle analysis indicated that compound 8a could arrest the cell cycle at the G1 phase for HeLa and CaSki cancer cells. At the predetermined IC50 values on cancer cells, compound 8a did not induce any necrotic (cytotoxic) death to the normal human lymphocytes. In the present design, (1S,4S)-2,5-diazabicyclo[2.2.1]heptane system was found to be superior than the piperazine counterpart 11.

Identification of a new class of antitumor agent capable to induce apoptosis without triggering necrotic cell death event is challenging. The present communication describes the multicomponent synthesis of seven new (1S,4S)-2,5-diazabicyclo[2.2.1]heptane-dithiocarbamates and their in vitro antiproliferative activity on cervical cancer cell line (CaSki), breast cancer cell line (MDA-MB231), lung cancer cell line (SK-Lu-1) and human lymphocytes. Among the synthesized dithiocarbamates, compound 9e displayed significant antiproliferative activity without inducing any necrotic cell death (both on tumour cells and lymphocytes) and induced apoptosis in tumor cells by the caspase dependent apoptotic pathway. The compound 9e also exhibited greater tumor selectivity than human lymphocytes. In silico ADME predictions revealed that compound 9e has the potential to be developed as a drug candidate. Rapid chemical modifications of this lead are thus highly necessary for further investigation as a drug like safer antitumor candidate and also to achieve compounds with better activity profile.

We report the synthesis and characterization of N-alkyl modified peptides by efficient coupling of N-methyl amino acids in solution phase. As a model peptide, the segment 37-42 (GGVVIA) of the Aβ-42 amyloid peptide derived from the amyloid precursor protein (Aβ-PP) was chosen. This peptide and its derivatives with N-methyl groups on Val40 and Ile41 residues were synthesized and character-ized. Because the synthesis was performed in solution-phase, the procedure can be easily scaled up for the production of larger amounts of the peptides described in this work or any linear N-methyl peptide with potential therapeutic application.

A21 (2) is a new polyene macrolide Amphotericin B amide antibiotic derived from amphotericin B AmB (1), which has been tested extensively on preclinical trials showing the same antimycotic effectiveness and increased margin of safety over AmB (1). We present the multigram scale synthesis, isolation, purity assessment by HPLC, and key aspects of its characterization by NMR studies of A21 (2).

Amphotericin B is the most potent antimycotic known to date. However due to its large collateral toxicity, its use, although long standing, had been limited. Many attempts have been made to produce derivatives with reduced collateral damage. The molecular mechanism of polyene has also been closely studied for this purpose and understanding it would contribute to the development of safe derivatives. Our study examined polyene action, including chemical synthesis, electrophysiology, pharmacology, toxicology and molecular dynamics. The results were used to support a novel Amphotericin B derivative with increased selectivity: L-histidine methyl ester of Amphotericin B. We found that this derivative has the same form of action as Amphotericin B, i.e. pore formation in the cell membrane. Its reduced dimerization in solution, when compared to Amphotericin B, is at least partially responsible for its increased selectivity. Here we also present the results of preclinical tests, which show that the derivative is just as potent as Amphotericin B and has increased safety.

A problem of uncontrolled internal corrosion of over 20 mpy was root cause analyzed within a system of pipelines connecting a set of oil wells onshore central Gulf of Mexico coastline. The root cause analysis was focused on a detailed characterization of the gas, mostly methane, and liquid phases, mostly genic waters as well as CO2, H2S, chlorides and MIC bacteria. The metallurgy of the corrosion damage over the steel pipeline involving both zone of uniform corrosion as well as special points of localized corrosion. In the past these pipelines received unsuccessful chemical treatments. Both the inhibitor and the dosage were not available. A trial of a new palm oil based corrosion inhibitor was performed. The corrosion inhibitor passed successful testing in the laboratory employing electrochemical testing by linear polarization resistance as well as harmonic analysis. The corrosion inhibitor formulation was furthermore adapted for massive fabrication involving a set of parallel variables from pipeline engineering such as foam formation elimination, not gel formation and other. The program was conducted producing improvements from the onset. Corrosion inhibitor injection was immediately and a program of pipeline cleaning was launched. In the first four months of the new corrosion inhibitor solution the results produced the desired target of overall compliance to the internal pipeline corrosion control specification below 2 mpy.

Fatty-amide derivatives were evaluated to study the effect of the double bonds into the hydrocarbon chain (C18) on the corrosion behavior of carbon steel. Electrochemical impedance spectroscopy (EIS) and real-time corrosion measurements were used to evaluate the inhibition mechanism of the fatty-amides on carbon steel in CO 2 -saturated (3% NaCl + 10% diesel) emulsion at 50°C. EIS results demonstrated that the unsaturation present into the hydrocarbon chain contributes to the efficiency of fatty-amides, because they can be adsorbed on the metal surface by a flat-adsorption process reducing the presence of active sites and blocking the corrosion process and preventing the diffusion of corrosive species, such as H 2 O, H + , Cl − , andHCO3-. Real-time corrosion measurements also indicated that the effectiveness of the inhibitors is dependent on the unsaturation into the hydrocarbon chain, being also a good technique to determine the stability of the adsorption process of the inhibitors.

AbstractThis article describes a study on the Michael addition reaction of cyclohexanone to nitroolefins catalyzed by the chiral secondary amine (1S,4S)-2-tosyl- 2,5-diazabicyclo[2.2.1]heptane. Reactions were carried out under solvent-free conditions to make them more environmentally friendly. Initially, the observed diastereoand enantioselectivities were moderate to good, but were significantly improved by lowering the reaction temperature. Furthermore, a variety of chiral acids were also tested as co-catalysts in both of their enantiomeric forms, which revealed that (R)-mandelic acid affords excellent results in terms of yield and stereoselectivity. Monitoring the reaction by MS-TOF allowed for the detection of key reaction intermediates, and a reasonable reaction mechanism in which both catalysts are involved is proposed.

Capsaicin is a potent inducer of apoptosis in tumourreceptor potential vanilloid 1 (TRPV1). The present study determined the IC50 and cytotoxic and apoptotic activities of the Capsaicin analogues Rinvanil and Phenylacetylrinvanil (PhAR) on three cervical cancer cell lines: HeLa, CaSKi and ViBo. These analogues possess an increased affinity for TRPV1 receptors. The IC50 obtained proved to be cytotoxic for all three cell lines; however, in the cells treated with Capsaicin both active caspase-3 and nuclear fragmentation were present. Capsaicin and its analogues also inhibited the normal proliferation of lymphocytes, suggesting that they are non-selective antitumour compounds. Finally, we discuss the possible loss of the relation between apoptosis and affinity to TRPV1, and the need for other strategies to synthesise Capsaicin analogues that can be useful in cancer treatments.