Salmonella Typhimurium and Pseudomonas aeruginosa Respond Differently to the Fe Chelator Deferiprone and to Some Novel Deferiprone Derivatives

The ability to obtain Fe is critical for pathogens to multiply in their host. For this reason, there is significant interest in the identification of compounds that might interfere with Fe management in bacteria. Here we have tested the response of two Gram-negative pathogens, Salmonella enterica serovar Typhimurium (STM) and Pseudomonas aeruginosa (PAO1), to deferiprone (DFP), a chelating agent already in use for the treatment of thalassemia, and to some DFP derivatives designed to increase its lipophilicity. Our results indicate that DFP effectively inhibits the growth of PAO1, but not STM. Similarly, Fe-dependent genes of the two microorganisms respond differently to this agent. DFP is, however, capable of inhibiting an STM strain unable to synthesize enterochelin, while its effect on PAO1 is not related to the capability to produce siderophores. Using a fluorescent derivative of DFP we have shown that this chelator can penetrate very quickly into PAO1, but not into STM, suggesting that a selective receptor exists in Pseudomonas. Some of the tested derivatives have shown a greater ability to interfere with Fe homeostasis in STM compared to DFP, whereas most, although not all, were less active than DFP against PAO1, possibly due to interference of the added chemical tails with the receptor-mediated recognition process. The results reported in this work indicate that DFP can have different effects on distinct microorganisms, but that it is possible to obtain derivatives with a broader antimicrobial action.

Unravelling the intricate cooperativity of subunit gating in P2X2 ion channels

Ionotropic purinergic (P2X) receptors are trimeric channels that are activated by the binding of ATP. They are involved in multiple physiological functions, including synaptic transmission, pain and inflammation. The mechanism of activation is still elusive. Here we kinetically unraveled and quantified subunit activation in P2X2 receptors by an extensive global fit approach with four complex and intimately coupled kinetic schemes to currents obtained from wild type and mutated receptors using ATP and its fluorescent derivative 2-[DY-547P1]-AET-ATP (fATP). We show that the steep concentration-activation relationship in wild type channels is caused by a subunit flip reaction with strong positive cooperativity, overbalancing a pronounced negative cooperativity for the three ATP binding steps, that the net probability fluxes in the model generate a marked hysteresis in the activation-deactivation cycle, and that the predicted fATP binding matches the binding measured by fluorescence. Our results shed light into the intricate activation process of P2X channels.

Fluorimetric Method for the Determination of Histidine in Random Human Urine Based on Zone Fluidics

In the present study, the determination of histidine (HIS) by an on-line flow method based on the concept of zone fluidics is reported. HIS reacts fast with o-phthalaldehyde at a mildly basic medium (pH 7.5) and in the absence of additional nucleophilic compounds to yield a highly fluorescent derivative (λex/λem = 360/440 nm). The flow procedure was optimized and validated, paying special attention to its selectivity and sensitivity. The LOD was 31 nmol·L−1, while the within-day and day-to-day precisions were better than 1.0% and 5.0%, respectively (n = 6). Random urine samples from adult volunteers (n = 7) were successfully analyzed without matrix effect (<1%). Endogenous HIS content ranged between 116 and 1527 μmol·L−1 with percentage recoveries in the range of 87.6%–95.4%.

Ultra-fast and universal detection of Gram-negative bacteria in complex samples based on colistin derivatives

A rapid and universal assay for detection of Gram-negative bacteria was developed using a fluorescent derivative of colistin. Labeling is achieved within 10 min in various bacteria relevant to hospital-acquired infections in complex samples.

Plasma membrane imaging with a fluorescent benzothiadiazole derivative

This work describes a novel fluorescent 2,1,3-benzothiadiazole derivative designed to act as a water-soluble and selective bioprobe for plasma membrane imaging. The new compound was efficiently synthesized in a two-step procedure with good yields. The photophysical properties were evaluated and the dye proved to have an excellent photostability in several solvents. DFT calculations were found in agreement with the experimental data and helped to understand the stabilizing intramolecular charge-transfer process from the first excited state. The new fluorescent derivative could be applied as selective bioprobe in several cell lines and displayed plasma-membrane affinity during the imaging experiments for all tested models.

Archangelolide: A sesquiterpene lactone with immunobiological potential from Laserpitium archangelica

Sesquiterpene lactones are secondary plant metabolites with sundry biological effects. In plants, they are synthesized, among others, for pesticidal and antimicrobial effects. Two such compounds, archangelolide and trilobolide of the guaianolide type, are structurally similar to the well-known and clinically tested lactone thapsigargin. While trilobolide has already been studied by us and others, there are only scarce reports on the biological activity of archangelolide. Here we present the preparation of its fluorescent derivative based on a dansyl moiety using azide–alkyne Huisgen cycloaddition having obtained the two sesquiterpene lactones from the seeds of Laserpitium archangelica Wulfen using supercritical CO2 extraction. We show that dansyl-archangelolide localizes in the endoplasmic reticulum of living cells similarly to trilobolide; localization in mitochondria was also detected. This led us to a more detailed study of the anticancer potential of archangelolide. Interestingly, we found that neither archangelolide nor its dansyl conjugate did exhibit cytotoxic effects in contrast to the structurally closely related counterparts trilobolide and thapsigargin. We explain this observation by a molecular dynamics simulation, in which, in contrast to trilobolide, archangelolide did not bind into the sarco/endoplasmic reticular calcium ATPase cavity utilized by thapsigargin. Last, but not least, archangelolide exhibited anti-inflammatory activity, which makes it promising compound for medicinal purposes.

Archangelolide: Sesquiterpene lactone with immunobiological potential from Laserpitium archangelica

Sesquiterpene lactones are secondary plant metabolites with sundry biological effects. In plants, they are synthesized mainly for their pesticidal, antimicrobial and other effects. Two such compounds, archangelolide and trilobolide of the guaianolide type, are structurally similar to the well-known and clinically tested lactone thapsigargin. Here we present the development of a facile method for isolation of these two sesquiterpene lactones from the seeds of Laserpitium archangelica Wulfen using supercritical CO2 extraction. Furthermore, not much has been reported on biological activity of archangelolide. We prepared its fluorescent derivative based on a dansyl moiety using azide-alkyne Huisgen cycloaddition. We showed that dansyl-archangelolide localized in endoplasmic reticulum of living cells similarly to trilobolide; nevertheless, localization in mitochondria was also detected. This led us to study the anticancer potential of archangelolide. Interestingly, we found that neither archangelolide nor its dansyl conjugate did exhibit cytotoxic effect in contrast to its structurally closely related counterparts trilobolide and thapsigargin. We explain this observation by a molecular dynamics simulation, in which, in contrast to trilobolide, archangelolide did not bind into the sarco/endoplasmic reticular calcium ATPase cavity utilized by thapsigargin. Last, but not least, archangelolide exhibited anti-inflammatory activity, which makes it promising compound for medicinal purposes.