Antiplasmodial and antimycobacterial activities of crude and lead-like enhanced extracts from Namibian medicinal plants.

ETHNOPHARMACOLOGICAL RELEVANCE: Eight indigenous medicinal plants which are used traditionally for the treatment of tuberculosis (TB), malaria, and associated symptoms, were selected for this study. AIM OF STUDY: The aim of this study was to evaluate the antiplasmodial and antimycobacterial activities of the organic and aqueous crude extracts of different plant parts, by comparing the activities of subfractions (lead-like enhanced [LLE] extracts and methanol fractions) prepared from the bioactive crude extracts. MATERIALS & METHODS: Crude aqueous and organic extracts were prepared for 25 different plant parts obtained from eight plant species. In vitro antiplasmodial activity was evaluated using the parasite lactate dehydrogenase assay against chloroquine-sensitive Plasmodium falciparum NF54 and in vitro antimycobacterial activity determined against the Mycobacterium tuberculosis H37Rv-GFP strain in a standard broth microdilution assay. The bioactive crude extracts were subjected to solid phase extraction with Strata-X 33 µm reversed phase cartridges and eluted with 70:30 MeOH: H2O:1% trifluoroacetic acid to yield the LLE extract, followed by a methanol rinse, herein referred to as the MeOH fraction. Both fractions were evaluated for antiplasmodial and antimycobacterial activity. Proton nuclear magnetic resonance spectroscopy (1H-NMR) and ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) profiling of the crude and active fractions of the phytochemically unexplored Sarcocaulon marlothii Engl. were performed to aid the identification of a potential antiplasmodial lead compound. RESULTS: Ten of the aqueous and organic crude extracts displayed antimycobacterial activity, with minimum inhibitory concentration (MIC90) values ranging from 9.9 to 86.8 µg/mL, and four crude extracts showed antiplasmodial activity with inhibitory concentration (IC50) values between 5.2 and 17.8 µg/mL. Although the stems of S. marlothii are traditionally used to treat TB and related symptoms, the two crude extracts displayed weak antimycobacterial activity (MIC90 > 100 µg/mL) while the crude organic extract displayed moderate antiplasmodial activity with an IC50 value of 8.8 µg/mL. None of the LLE extracts prepared from the ten antimycobacterial-active crudes displayed any significant activity (MIC90 > 125 µg/mL). In contrast, fractionation of three antiplasmodial-active, crude organic extracts yielded MeOH fractions which displayed a 2-fold to 19-fold increase in activity. The 1H-NMR profiles of the active MeOH fraction (IC50 4.3 µg/mL) of S. marlothii (organic, stem) revealed the likely presence of an unidentified trisubstituted cinnamic acid derivative as one of the major compounds and UPLC-MS/MS data provided additional evidence that the compound may be a hydroxycinnamic acid derivative. Unfortunately, owing to the paucity of the material obtained, we were unable to purify and unequivocally determine the structure of this active compound. CONCLUSIONS: This is the first report on the phytochemical profiling of S. marlothii and, based on the antiplasmodial activity recorded, it merits an in-depth phytochemical analysis for the unequivocal characterization of a potential antiplasmodial lead compound. Results from this study lend support to the effectiveness of extract enrichment in combination with NMR fingerprinting for antiplasmodial lead identification.

Recent trends in the chromatographic analysis of volatile flavor and fragrance compounds: Annual review 2020.

The chromatographic analysis of volatile flavor and fragrance compounds is performed routinely in several industries and in many fields of scientific research. Typical applications include food-, environmental-, essential oil- and cosmetics analysis. Even though the analysis of flavors and fragrances have become increasingly standardized during the past decade, there are still a large variety of techniques that can be used for their extraction, chemical analysis, and sensory analysis. Moreover, there are certain less commonly used techniques that are now being used with increased frequency and that are showing the potential of being used as alternatives to the existing standard techniques. In this annual review, the techniques that were most commonly used in 2020 for the investigation of these volatile compounds are discussed. In addition, a number of emerging trends are discussed, notably the use of solvent assisted flavor evaporation (SAFE) for extraction, GC ion mobility spectrometry (IMS) for volatile compound analysis and electronic senses, that is, E-noses and E-tongues, for sensory analysis. Miscellaneous hyphenated techniques, advances in stationary phase chemistry and a number of interesting applications are also highlighted.

Synthesis and In Vitro Antiprotozoan Evaluation of 4-/8-Aminoquinoline-based Lactams and Tetrazoles.

A second generation of 4-aminoquinoline- and 8-aminoquinoline-based tetrazoles and lactams were synthesized via the Staudinger and Ugi multicomponent reactions. These compounds were subsequently evaluated in vitro for their potential antiplasmodium activity against a multidrug-resistant K1 strain and for their antitrypanosomal activity against a cultured T. b. rhodesiense STIB900 strain. Several of these compounds (4a-g) displayed good antiplasmodium activities (IC50 = 0.20-0.62 µM) that were comparable to the reference drugs, while their antitrypanosomal activity was moderate (<20 µM). Compound 4e was 2-fold more active than primaquine and was also the most active (IC50 = 7.01 µM) against T. b. rhodesiense and also exhibited excellent aqueous solubility (>200 µM) at pH 7.

Gas Chromatography-Mass Spectrometry (GC-MS) Combined with Retention Index Prediction for the Rapid Identification of Halogenated monoterpenes from a Namibian Plocamium species.

Plocamiun species collected from the Namibian coast display morphological features similar to those of both P. rigidum and P. suhrii which makes identification of these species a difficult task. It has been reported that the major secondary metabolites found in various Plocamium species are unique to each species [1]. In this study GC-MS combined with a retention index (RI) prediction strategy was used for the rapid identification of halogenated monoterpenes characteristic of a particular Namibian Plocatnium species. The RIs of the metabolites were matched with the predicted RIs of halogenated monoterpenes for which similar MS data have been reported for the same species of Plocamium. Based on the identification of the major secondary metabolite, IE,3R,4S,5E,7Z- 1-bromo-3;4,8- trichloro-7-(dichloromethyl)-3-methylocta-1,5,7-triene [2], it was proposed that these Namibian samples are closely related to that of P. suhrii. From. this, it was determined that the proposed P. suhrii specimens collected in Namibia contain four additional metabolites (with molecular formulae C10H16Br2C2, C10H11BrCI4, C10H9BrCl6 and an unknown compound) previously not reported in P. suhrii species. In addition, a compound previously identified in South African P. suhrii was not present in the Namibian Plocamium specimens.

Drug metabolite generation using a laboratory evolved NADPH independent cytochrome P450: application of in vitro and in silico approaches.

Twelve disparate drugs were subjected to metabolite generation by a laboratory evolved bacterial cytochrome P450 to investigate feasibility of the bacterial CYP to generate drug metabolites. Seven drugs were metabolised by the bacterial cytochromes to give diverse metabolites, which were compared to human metabolites reported in literature. Several non human metabolites were also generated by the bacterial CYP in addition to the known human metabolites. From docking studies and in silico sites of metabolism results, it was shown that the binding mode of the drug molecule and its distance from the active site in the binding pocket of the CYP was important for metabolism. This contribution reports, for the first time, previously uncharacterised metabolites of this bacterial cytochrome and demonstrates the potential usefulness of human CYP-based prediction software when used in combination with bacterial CYPs for metabolite generation.

Biotransformation and biocatalysis: roles and applications in the discovery of antimalarials.

Several strategies to discover new antimalarials have been proposed to augment and complement the conventional drug-discovery paradigm. One approach, which has not yet been fully exploited, is the use of drug biotransformation to identify new active molecules. This concept rests on the use of the biotransformation of drugs to their pharmacologically active metabolites. This approach has been used successfully in human chemotherapy, with the discovery and development of several metabolite-based drugs. This review looks at the contribution that biotransformations can play in antimalarial drug discovery.

Comparison of electrospray ionisation, atmospheric pressure chemical ionisation and atmospheric pressure photoionisation for the identification of metabolites from labile artemisinin-based anti-malarial drugs using a QTRAP® mass spectrometer.

RATIONALE: Artemisinin-based drugs and their metabolites are prone to in-source fragmentation under atmospheric pressure ionisation mass spectrometry (API-MS) conditions. To facilitate correct and efficient identification of all possible drug metabolites using full scan MS analyzer methods, stable [M + NH(4) ](+) ions should be produced in the MS source. METHODS: Using a high-performance liquid chromatography (HPLC) hybrid triple quadrupole linear ion trap MS system, electrospray ionisation (ESI), atmospheric pressure chemical ionisation (APCI) and atmospheric pressure photoionisation (APPI) methods were developed for the detection of [M + NH(4) ](+) ions of the test compounds dihydroartemisinin, artemisinin, artemether and artesunic acid. The optimised methods employed ammonium formate buffered HPLC mobile phase in combination with moderate source temperatures (100-200 °C) and showed satisfactorily reduced in-source fragmentation. RESULTS: With a full scan MS analyser method for the detection of the in vitro metabolites of the test compounds, the respective performance of the ESI and APCI methods was found to be comparable. ESI generally resulted in less in-source fragmentation. Incorrect assignment of metabolites resulted from strong in-source fragmentation of artemether using the APPI method. The most number of metabolites could be detected using ESI in combination with a selective MS analyser method. CONCLUSIONS: ESI and APCI full scan methods proved to be capable of detecting any drug metabolites present in reasonable concentrations, and are useful when employed in addition to selective scan methods that target low level expected metabolites. APPI can be a valuable alternative for detecting expected metabolites due to good signal-to-noise (S/N) ratio.

New ketomethylene inhibitor analogues: synthesis and assessment of structural determinants for N-domain selective inhibition of angiotensin-converting enzyme.

Angiotensin-converting enzyme (ACE) is a zinc metallopeptidase containing two homologous domains. While the C-domain plays a major role in blood pressure regulation, the N-domain hydrolyzes the antifibrotic agent N-acetyl-Ser-Asp-Lys-Pro. Thus, N-domain selective (N-selective) inhibitors could be useful in the treatment of conditions relating to excessive tissue fibrosis. New keto-ACE analogues were designed that contained functionalities considered important for N-selective inhibitor RXP407 binding, namely, a P(2) Asp, N-acetyl group, and C-terminal amide. Such functionalities were incorporated to assess the structural determinants for N-selective binding in a novel inhibitor template. Inhibitors containing a C-terminal amide and modified P(2)' group were poor inhibitors of the N-domain, with several of these displaying improved inhibition of the C-domain. Molecules with both a C-terminal amide and P(2) Asp were also poor inhibitors and not N-selective. Compounds containing a free C-terminus, a P(2) Asp and protecting group displayed a change of more than 1000-fold N-selectivity compared with the parent molecule. Molecular docking models revealed interaction of these P(2) groups with S(2) residues Tyr369 and Arg381. This study emphasizes the importance of P(2) functionalities in allowing for improved N-selective binding and provides further rationale for the design of N-selective inhibitors, which could be useful in treating tissue fibrosis.

Lizard epidermal gland secretions. II. Chemical characterization of the generation gland secretion of the sungazer, Cordylus giganteus.

In lizards, the epidermal glands of the femoral and precloacal regions are involved in the production of semiochemicals. In addition to its femoral glands, the giant girdled lizard, or sungazer, Cordylus giganteus, which is endemic to South Africa, has generation glands as an additional potential source of semiochemicals. These epidermal glands are described as glandular scales that overlay the femoral glands and are included in the normal epidermal profile located in the femoral (thigh) and anterior antebrachial (fore-leg) regions of the male sungazer. GC-MS analysis of the generation gland secretions and the trimethylsilyl derivatives of some of the steroidal constituents was employed to identify 59 constituents, including alkenes, carboxylic acids, alcohols, ketones, aldehydes, esters, amides, nitriles, and steroids. The quantitative differences of the volatile constituents of the fore- and hind-leg generation glands were compared between individuals. This is the first report on the chemical composition of generation glandular material of lizards.

Synthesis and in vitro antitubercular activity of ferrocene-based hydrazones.

We report here the synthesis and in vitro antitubercular activity of a new series of ferrocenyl derivatives. The quinoline-ferrocene hybrid 5 exhibited significant activity (MIC=2.5-5 µg/ml) against Mycobacterium tuberculosis. Results indicate that such hybrid compounds provide an efficient approach for future pharmacological developments to fight against tuberculosis. Moreover, the antimalarial drug candidate ferroquine (FQ, SSR97193) was also evaluated mainly because of its structural similarity. FQ was found to display moderate inhibitory activity (MIC=10-15 µg/ml) against M. tuberculosis. This new drug may offer an interesting alternative in endemic area where malaria and tuberculosis coexist.

Catching a gently thrown ball.

Several studies have shown that people can catch a ball even if it is visible only during part of its flight. Here, we examine how well they can do so. We measured the movements of a ball and of the hands of both the thrower and the catcher during one-handed underarm throwing and catching. The catcher's sight was occluded for 250 ms at random moments. Participants could catch most balls without fumbling. They only really had difficulties if vision was occluded before the ball was released and was restored less than 200 ms before the catch. In such cases, it was impossible to accurately predict the ball's trajectory from motion of the ball and of the thrower's hand before the occlusion, and there was not enough time to adjust the catching movement after vision was restored. Even at these limits, people caught most balls quite adequately.

High-efficiency hydrophilic interaction chromatography by coupling 25 cm x 4.6mm ID x 5 microm silica columns and operation at 80 degrees C.

Recently, hydrophilic interaction chromatography (HILIC) has emerged as a valuable orthogonal tool to reversed-phase liquid chromatography (RP-LC) as it allows for resolution of highly polar ionisable compounds. The relationships between separation efficiency, column length and speed of analysis for 4.6 mm ID x 5 microm silica particle columns in HILIC are demonstrated using kinetic plots. The kinetic plots constructed for conventional pressure systems operating at 350 bar and at 30 degrees C and 80 degrees C are confirmed using experimental data for different column lengths. Efficiencies of more than 130,000 theoretical plates could be achieved by connecting up to six columns of 25 cm. As expected, a significant gain in analysis speed without loss of efficiency could be obtained by operating at 80 degrees C compared to 30 degrees C. The advantages of using long columns in HILIC in combination with elevated column temperature for the pharmaceutical industry are illustrated using test mixtures comprised of commercially available ionisable compounds (including some containing functional groups with potential genotoxic typical structural alerts) as well as real polar ionisable pharmaceuticals.

Slant cues are processed with different latencies for the online control of movement.

For the online control of movement, it is important to respond fast. The extent to which cues are effective in guiding our actions might therefore depend on how quickly they provide new information. We compared the latency to alter a movement when monocular and binocular cues indicated that the surface slant had changed. We found that subjects adjusted their movement in response to three types of information: information about the new slant from the monocular image, information about the new slant from binocular disparity, and information about the change in slant from the change in the monocular image. Responses to changes in the monocular image were approximately 40 ms faster than responses to a new slant estimate from binocular disparity and about 90 ms faster than responses to a new slant estimate from the monocular image. Considering these delays, adjustments of ongoing movements to changes in slant will usually be initiated by changes in the monocular image. The response will later be refined on the basis of combined binocular and monocular estimates of slant.

Serial coupling of reversed-phase and hydrophilic interaction liquid chromatography to broaden the elution window for the analysis of pharmaceutical compounds.

It is presently a common practice in drug discovery to analyse samples by reversed-phase liquid chromatography (RPLC) and hydrophilic interaction chromatography (HILIC). To increase throughput, HILIC was connected in series to RPLC by means of a T-piece with make-up flow. The first column is a 2mm I.D. column having an optimal flow between 0.1 and 0.2mL/min. Via the T-piece, the flow for the second dimension column with an I.D. of 4.6mm is adjusted to 1.5-2.0mL/min with a high acetonitrile content (i.e. >/=80%) mobile phase. Therefore, even in gradient RPLC analysis starting with a mobile phase with high water content, the HILIC column is always operated at high acetonitrile concentration which is required to obtain retention on the HILIC column. The performance of the hyphenated RPLC/HILIC set-up is illustrated with the analysis of two model samples of pharmaceutical interest. Optimization of the conditions in the HILIC dimension is discussed.

Lizard epidermal gland secretions I: Chemical characterization of the femoral gland secretion of the sungazer, Cordylus giganteus.

The giant girdled lizard or sungazer, Cordylus giganteus, is endemic to South Africa. It has been suggested that in this species, as in other lizard species, epidermal glands in the femoral, pre-cloacal regions, and cloacal glands are the main sources of semiochemicals and that these secretions could play an important role at different levels of the social biology of the animals. To gain a better understanding of the nature of the femoral gland secretions of the sungazer, characterization of the constituents of the secretions was carried out. By using GC-MS analysis, in conjunction with auxiliary techniques, such as solventless sample introduction and trimethylsilyl derivatization, 53 relatively involatile compounds, including carboxylic acids, alcohols, ketones, esters, and steroids, were identified in the secretions of both sexes. The study showed that the secretions of male and female sungazers contain only semi-volatile chemicals.

Judging surface slant for placing objects: a role for motion parallax.

People have a variety of sources of information (cues) about surface slant at their disposal. We used a simple placing task to evaluate the relative importance of three such cues (motion parallax, binocular disparity and texture) within the space in which people normally manipulate objects. To do so, we projected a stimulus onto a rotatable screen. This allowed us to manipulate texture cues independently of binocular disparity and motion parallax. We asked people to stand in front of the screen and place a cylinder on the screen. We analysed the cylinder's orientation just before contact. Participants mainly relied on binocular cues (weight between 50 and 90%), in accordance with binocular cues being known to be reliable when the stimulus surface is nearby and almost frontal. Texture cues contributed between 2 and 18% to the estimated slant. Motion parallax was given a weight between 1 and 9%, despite the fact that it only provided information when the head began to move, which was just before the arm did. Thus motion parallax is used to judge surface slant, even when one is under the impression of standing still.

The contribution of covariation to skill improvement is an ambiguous measure: comment on Müller and Sternad (2004).

It has been proposed that it is possible to decompose changes in variability of human motor behavior into 3 independent components: covariation, task tolerance, and stochastic noise (H. Müller & D. Sternad). The authors simulate learning to throw accurately and show that for this task the proposed analysis does not give an unambiguous answer to the question of what the 3 components contribute to the simulated skill improvement. It is argued that this is caused by the fact that the component covariation depends on the choice of control variables. The authors conclude that it is not possible to distinguish between the 3 components of noise reduction without knowing the controlled variables.

Haptic detection of sine-wave gratings.

We studied human haptic perception of sine-wave gratings. In the first experiment we measured the dependence of amplitude detection thresholds on the number of cycles and on the wavelength of the gratings. In haptic perception of sine-wave gratings, the results are in agreement with neural summation. The rate at which detection thresholds decrease with increasing number of cycles is much higher than can be accounted for by probability summation alone. Further, neural summation mechanisms describe the detection thresholds accurately over the whole spatial range probed in the experiment, that is wavelengths from 14 mm up to 225 mm. Earlier, we found a power-law dependence of thresholds on the spatial width of Gaussian profiles (Louw et al, 2000 Experimental Brain Research 132 369-374). The current results extend these findings; the power-law dependence holds not only for Gaussian profiles, but also for a broad range of sine-wave gratings with the number of cycles varying between 1 and 8. Haptic perception involves tactual scanning combined with an active, dynamic exploration of the environment. We measured characteristics of the velocity and force with which stimuli were scanned while performing a psychophysical task. One particularly surprising finding was that, without being instructed, participants maintained an almost constant scanning velocity during each 45-min session. A constant velocity in successive trials of the experiment might facilitate or even be necessary for discrimination. Further, a large systematic dependence of velocity on scanning length was found. An eightfold increase in scanning length resulted in about a fourfold increase in scanning velocity. A second experiment was conducted to study the influence of scanning velocity on psychophysical detection thresholds. This was done by systematically imposing specific scanning velocities to the participants while the thresholds were measured. The main result of the second experiment was that psychophysical detection thresholds are constant over a relatively broad range of scanning velocities.

Haptic discrimination of stimuli varying in amplitude and width.

We studied active haptic discrimination of the geometrical features of an object. The geometrical parameters under investigation were the amplitude and width of a gaussian-shaped surface. Haptic discrimination thresholds were measured with regard to three values of these geometrical parameters. We found that humans discriminate up to about 300 shapes when both amplitude and width are extrapolated to the range between 1 micro m and 1 m. Over this range, which covers the span of the arms, the number of discriminations is small compared to the number across the full range of chromaticities in vision. Roughly speaking, humans are far better at discriminating sharp (extensive amplitude and little width) gaussian surfaces from smooth (small amplitude and extensive width) ones than they are at discriminating small (small amplitude and width) surfaces from large (extensive amplitude and width) ones. Our main conclusion is that discrimination in the geometrical domain is poorest when the proportion between amplitude and width is roughly the same for both shapes. Our results are in close agreement with results of earlier experiments on detection thresholds. This indicates that similar, or even the same, neural mechanisms were used for detection and discrimination of the geometrical parameters under investigation.