Bridging the maytansine and vinca sites: Cryptophycins target ß-tubulin's T5-loop.

Cryptophycins are microtubule-targeting agents (MTAs) that belong to the most potent antimitotic compounds known to date; however, their exact molecular mechanism of action remains unclear. Here, we present the 2.2 Å resolution X-ray crystal structure of a potent cryptophycin derivative bound to the αß-tubulin heterodimer. The structure addresses conformational issues present in a previous 3.3 Å resolution cryo-electron microscopy structure of cryptophycin-52 bound to the maytansine site of ß-tubulin. It further provides atomic details on interactions of cryptophycins, which had not been described previously, including ones that are in line with structure-activity relationship studies. Interestingly, we discovered a second cryptophycin-binding site that involves the T5-loop of ß-tubulin, a critical secondary structure element involved in the exchange of the guanosine nucleotide and in the formation of longitudinal tubulin contacts in microtubules. Cryptophycins are the first natural ligands found to bind to this new "ßT5-loop site" that bridges the maytansine and vinca sites. Our results offer unique avenues to rationally design novel MTAs with the capacity to modulate T5-loop dynamics and to simultaneously engage multiple ß-tubulin binding sites.

Conditional Cell Penetration of Masked CPPs by an ADEPT-like Approach.

The intracellular delivery of cargos via cell penetrating peptides (CPPs) holds significant promise as a drug delivery vehicle, but a major issue is their lack of cell type specificity, which can lead to detrimental off-target effects. We use an ADEPT-like concept to introduce conditional and selective activation of cellular uptake by using the lysine-rich, cationic, and amphiphilic L17E peptide as a model CPP. By masking the lysine residues of the L17E peptide with enzyme-cleavable acetyl protecting groups, the delivery of the covalently conjugated fluorophore TAMRA to HeLa cells was diminished. Recovery of cellular uptake could be achieved by deacetylation of the masked acetylated L17E peptide using the NAD-dependent sirtuin 2 (SirT2) deacetylase in vitro. Finally, trastuzumab-SirT2 and anti-B7H3-SirT2 antibody-enzyme conjugates were generated for the conditional and selective delivery of a cryptophycin cytotoxin by the L17E peptide. While the masked peptide still demonstrated some cytotoxicity, selective cell killing mediated by the antibody-enzyme conjugates was observed.

Small molecule- and peptide-drug conjugates addressing integrins: A story of targeted cancer treatment.

Targeted cancer treatment should avoid side effects and damage to healthy cells commonly encountered during traditional chemotherapy. By combining small molecule or peptidic ligands as homing devices with cytotoxic drugs connected by a cleavable or non-cleavable linker in peptide-drug conjugates (PDCs) or small molecule-drug conjugates (SMDCs), cancer cells and tumours can be selectively targeted. The development of highly affine, selective peptides and small molecules in recent years has allowed PDCs and SMDCs to increasingly compete with antibody-drug conjugates (ADCs). Integrins represent an excellent target for conjugates because they are overexpressed by most cancer cells and because of the broad knowledge about native binding partners as well as the multitude of small-molecule and peptidic ligands that have been developed over the last 30 years. In particular, integrin αVß3 has been addressed using a variety of different PDCs and SMDCs over the last two decades, following various strategies. This review summarises and describes integrin-addressing PDCs and SMDCs while highlighting points of great interest.

Increasing the Stability of Flavin-Dependent Halogenases by Disulfide Engineering.

Flavin-dependent halogenases allow halogenation of electron-rich aromatic compounds under mild reaction conditions even at electronically unfavored positions with high regioselectivity. In order to expand the application of halogenases, the enzymes need to be improved in terms of stability and efficiency. A previous study with the tryptophan 6-halogenase Thal demonstrated that thermostable Thal variants tend to form dimers in solution while the wild type is present as a monomer. Based on this a dimeric Thal variant was generated that is covalently linked by disulfide bonds. Introducing two cysteine residues at the dimer interface resulted in the variant Thal CC with significantly increased thermostability (▵T50 =15.7 K) and stability over time at elevated temperature compared to the wild type. By introducing the homologous mutations into the tryptophan 5-halogenase PyrH, we were able to show that the stabilization by covalent dimerization can also be transferred to other halogenases. Moreover, it was possible to further increase the thermostability of PyrH by inserting cysteine mutations at alternative sites of the dimer interface.

Enzymatic Peptide and Protein Bromination: The BromoTrp Tag.

Bio-orthogonal reactions for modification of proteins and unprotected peptides are of high value in chemical biology. The combination of enzymatic halogenation with transition metal-catalyzed cross-coupling provides a feasible approach for the modification of proteins and unprotected peptides. By a semirational protein engineering approach, variants of the tryptophan 6-halogenase Thal were identified that enable efficient bromination of peptides with a C-terminal tryptophan residue. The substrate scope was explored using di-, tri-, and tetrapeptide arrays, leading to the identification of an optimized peptide tag we named BromoTrp tag. This tag was introduced into three model proteins. Preparative scale post-translational bromination was possible with only a single cultivation and purification step using the brominating E. coli coexpression system Brocoli. Palladium-catalyzed Suzuki-Miyaura cross-coupling of the bromoarene was achieved with Pd nanoparticle catalysts at 37 °C, highlighting the rich potential of this strategy for bio-orthogonal functionalization and conjugation.

Evaluation of anti-quorum sensing and antibiofilm effects of secondary metabolites from Gambeya lacourtiana (De Wild) Aubr. & Pellegr against selected pathogens.

BACKGROUND: Microbial infections cause serious health problems especially with the rising antibiotic resistance which accounts for about 700,000 human deaths annually. Antibiotics which target bacterial death encounter microbial resistance with time, hence, there is an urgent need for the search of antimicrobial substances which target disruption of virulence factors such as biofilm and quorum sensing (QS) with selective pressure on the pathogens so as to avoid resistance. METHODS: Natural products are suitable leads for antimicrobial drugs that can inhibit bacterial biofilms and QS. Twenty compounds isolated from the medicinal plant Gambeya lacourtiana were evaluated for their antibiofilm and anti-quorum sensing effects against selected pathogenic bacteria. RESULTS: Most of the compounds inhibited violacein production in Chromobacterium violaceum CV12472 and the most active compound, Epicatechin had 100% inhibition at MIC (Minimal Inhibitory Concentration) and was the only compound to inhibit violacein production at MIC/8 with percentage inhibition of 17.2 ± 0.9%. Since the bacteria C. violaceum produces violacein while growing, the inhibition of the production of this pigment reflects the inhibition of signal production. Equally, some compounds inhibited violacein production by C. violaceum CV026 in the midst of an externally supplied acylhomoserine lactone, indicating that they disrupted signal molecule reception. Most of the compounds exhibited biofilm inhibition on Staphyloccocus aureus, Escherichia coli and Candida albicans and it was observed that the Gram-positive bacteria biofilm was most susceptible. The triterpenoids bearing carboxylic acid group, the ceramide and epicatechin were the most active compounds compared to others. CONCLUSION: Since some of the compounds disrupted QS mediated processes in bacteria, it indicates that this plant is a source of antibiotics drugs that can reduce microbial resistance.

Extended Biocatalytic Halogenation Cascades Involving a Single-Polypeptide Regeneration System for Diffusible FADH2.

Flavin-dependent halogenases have attracted increasing interest for aryl halogenation at unactivated C-H positions because they are characterised by high regioselectivity, while requiring only FADH2 , halide salts, and O2 . Their use in combined crosslinked enzyme aggregates (combiCLEAs) together with an NADH-dependent flavin reductase and an NADH-regeneration system for the preparative halogenation of tryptophan and indole derivatives has been previously described. However, multiple cultivations and protein purification steps are necessary for their production. We present a bifunctional regeneration enzyme for two-step catalytic flavin regeneration using phosphite as an inexpensive sacrificial substrate. This fusion protein proved amenable to co-expression with various flavin-dependent Trp-halogenases and enables carrier-free immobilisation as combiCLEAs from a single cultivation for protein production and the preparative synthesis of halotryptophan. The scalability of this system was demonstrated by fed-batch fermentation in bench-top bioreactors on a 2.5 L scale. Furthermore, the inclusion of a 6-halotryptophan-specific dioxygenase into the co-expression strain further converts the halogenation product to the kynurenine derivative. This reaction cascade enables the one-pot synthesis of l-4-Cl-kynurenine and its brominated analogue on a preparative scale.

Two new 30-norfriedelane triterpenes from Caloncoba glauca (P. Beauv.) Gilg (Achariaceae).

The chemical investigation of the methanolic root extract of Caloncoba glauca (P. Beauv.) Gilg exhibited two new 30-norfriedelane triterpenes, glaucalactones A and B (1-2), together with eight known compounds, caloncobalactone (3), friedelin (4), friedelanol (5), 3-oxo-friedelan-28-oic acid (6), stigmasterol (7), ß-sitosterol (8), ß-sitosterol-3-O-ß-D-glucopyranoside (9) and pentacosanoic acid (10). The structures of the isolates were elucidated by extensive spectroscopic and spectrometric analyses (1D and 2D NMR, ESI-MS) and by comparison with previously reported data. All the compounds were tested for their antioxidant, antifungal and antibacterial activities. Compound 1 displayed weak antibacterial effect with MIC value of 125 µg/mL against Staphylococcus aureus and Escherishia coli. Compound 6 exhibited moderated antifungal activity against Candida krusei with MIC value of 62.5 µg/mL. All the isolates were found to be inactive as antioxidants in the DPPH, ABTS and FRAP assays.

Polyoxygenated Stigmastane-Type Steroids from Vernonia kotschyana Sch. Bip. ex Walp. and Their Chemophenetic Significance.

Four polyoxygenated stigmastanes (1-4) alongside known analogues (7-8) and flavonoids (5-6) were isolated from a dichloromethane/methanol (1:1, v/v) extract of the whole plant of Vernonia kotschyana Sch. Bip. ex Walp. (Asteraceae). Their structures were determined by means of spectroscopic and spectrometric analysis. The relative stereochemistry of the new compounds was established and confirmed via biosynthesis evidence and cyclization of 1 under acidic conditions. A plausible biosynthetic pathway to the new compounds and the chemophenetic significance of the isolated constituents were also discussed. The crude extract, fractions, and compounds (1-3) were assessed for their antibacterial activity against five highly prevalent bacterial strains. The fractions and compounds showed low to moderate activity with minimal inhibitory concentrations (MICs) > 125 µg/mL.

In vitro antiplasmodial activity and toxicological profile of extracts, fractions and chemical constituents of leaves and stem bark from Dacryodes edulis (Burseraceae).

BACKGROUND: Dacryodes edulis is a plant that belongs to the Burseraceae family. It is widely used traditionally alone or in association with other plants in Cameroonian folk medicine to cure wounds, fever, headaches, and malaria. The aim of this work was to investigate the leaves and stem bark of D. edulis with an emphasis on the antiplasmodial and cytotoxic effects of extracts, fractions, and isolated compounds. METHODS: Extracts, fractions, and some isolated compounds were subjected to antiplasmodial activity screening in vitro against chloroquine-sensitive 3D7 and multidrug resistant Dd2 strains of Plasmodium falciparum using a SyBr Green fluorescence-based assay. The cytotoxicity of active extracts, fractions, and compounds was tested against mammalian Raw cell lines using an in vitro resazurin-based viability assay. The structures of the compounds were determined based on their NMR and MS data. The in vivo toxicity using female BALB/c mice was performed on the most active extract according to the protocol of OECD (2002), guideline 423. RESULTS: The hydroethanolic extract from the leaves of D. edulis displayed good antiplasmodial activity with IC50 values of 3.10 and 3.56 µg/mL respectively on sensitive (3D7) and multiresistant (Dd2) strains of P. falciparum. Of the sixteen compounds isolated, 3,3',4-tri-O-methylellagic acid (4) exhibited the highest antiplasmodial activity against PfDd2 strains with an IC50 value of 0.63 µg/mL. All extracts, fractions, and isolated compounds demonstrated no cytotoxicity against Raw cell lines with CC50 > 250 µg/mL. In addition, the most active extract on both strains of P. falciparum was nontoxic in vivo, with a LD50 greater than 2000 and 5000 mg/kg. A phytochemical investigation of the stem bark and leaves of D. edulis afforded sixteen compounds, including two xanthones (1-2), three ellagic acid derivatives (3-5), one phenolic compound (6), one depside (7), one triglyceride (8), one auranthiamide acetate (9), one gallic acid derivative (10), four triterpenoids (11-14), and two steroids (15-16). Compounds 1, 2, 5, 7, 8, and 9 were herein reported for the first time from the Burseraceae family. CONCLUSION: This work highlights the good in vitro antiplasmodial potency of the hydroethanolic extract of the leaves of this plant and that of two isolated constituents (3,3',4-tri-O-methylellagic acid and ethylgallate) from the plant. These biological results support the use of D. edulis in traditional medicine against malaria.

Co-Immobilization of a Multi-Enzyme Cascade: (S)-Selective Amine Transaminases, l-Amino Acid Oxidase and Catalase.

An enzyme cascade was established previously consisting of a recycling system with an l-amino acid oxidase (hcLAAO4) and a catalase (hCAT) for different α-keto acid co-substrates of (S)-selective amine transaminases (ATAs) in kinetic resolutions of racemic amines. Only 1 mol % of the co-substrate was required and l-amino acids instead of α-keto acids could be applied. However, soluble enzymes cannot be reused easily. Immobilization of hcLAAO4, hCAT and the (S)-selective ATA from Vibrio fluvialis (ATA-Vfl) was addressed here. Immobilization of the enzymes together rather than on separate beads showed higher reaction rates most likely due to fast co-substrate channeling between ATA-Vfl and hcLAAO4 due to their close proximity. Co-immobilization allowed further reduction of the co-substrate amount to 0.1 mol % most likely due to a more efficient H2 O2 -removal caused by the stabilized hCAT and its proximity to hcLAAO4. Finally, the co-immobilized enzyme cascade was reused in 3 cycles of preparative kinetic resolutions to produce (R)-1-PEA with high enantiomeric purity (97.3 %ee). Further recycling was inefficient due to the instability of ATA-Vfl, while hcLAAO4 and hCAT revealed high stability. An engineered ATA-Vfl-8M was used in the co-immobilized enzyme cascade to produce (R)-1-(3-ethoxy-4-methoxyphenyl)-2-(methylsulfonyl)ethanamine, an apremilast-intermediate, with a 1,000 fold lower input of the co-substrate.

Foetidumins A-D, and other chemical constituents from Helichrysum foetidum (L.) Moench (Asteraceae) with antiparasite activity.

The phytochemical investigation of the MeOH and CH2Cl2-MeOH (1:1) extracts from the flowers and twigs of Helichrysumfoetidum (L.) Moench (Asteraceae), which showed antileishmanial and antiplasmodial activities during the preliminary screening, led to the isolation of four undescribed compounds, including two ent-beyer-15-ene-type diterpenoids, foetidumins A (1) and B (2), one flavonoid, foetidumin C (3) and one chalcopyrone, foetidumin D (4). Additionally, fourteen known compounds comprising, two ent-beyer-15-ene-type diterpenoids (5-6), six flavonoids (7-12), two steroids (13-14), three triterpenoids (15-17), and one glyceryl monostearate (18) were also isolated. The chemical structures of foetidumins A-D were fully elucidated by analyses of their spectroscopic data. The structure and the stereochemistry of foetidumin A (1) were confirmed by SC-XRD analyses. Among the tested compounds, foetidumin C (3), erythroxylol A (6), and kaempferol (7) displayed the highest antileishmanial potency with IC50 values of 13.0, 11.8, and 11.1 µM, respectively. Foetidumin C (3) had no cytotoxicity toward Vero cells with the selectivity index > 3.59. Meanwhile, extracts of flowers and twigs had higher activity against Plasmodium falciparum chloroquine-sensitive (Pf3D7) strain with IC50 values of 3.66 and 10.52 µg/mL, respectively.

Elemane-Type Sesquiterpene, Acetonide Derived Polyacetylene and Other Constituents from the Whole Plant of Gymnanthemum theophrastifolium (Schweinf. ex Oliv. & Hiern) H.Rob. and Their Chemophenetic Significance.

The chemical investigation of the methanol extract of the whole plant of Gymnanthemum theophrastifolium (Schweinf. ex Oliv. & Hiern) H.Rob. (Asteraceae) led to the isolation of a new elemane-type sesquiterpene (1), a new acetonide derived polyacetylene (2) and a naturally occurring compound (3) from the plant kingdom along with sixteen known compounds (4-19). Their structures were elucidated by extensive NMR and MS analysis. This is the first report on the chemical constituents of G. theophrastifolium. Furthermore, compounds 12, 13, and 14 are reported for the first time from the family Asteraceae, while compound 9 is reported for the first time from the genus Gymnanthemum. Thus, the present results provide valuable insights to the chemophenetic knowledge of G. theophrastifolium, which is also discussed in this work.

Constituents of the Stem Bark of Symphonia globulifera Linn. f. with Antileishmanial and Antibacterial Activities.

The chemical investigation of the n-hexane fraction from the methanol extract of the stem bark of Symphonia globulifera Linn f., which displayed good in vitro activity against Leishmania donovani NR-48822 promastigotes (IC50 43.11 µg/mL), led to the isolation of three previously unreported polyprenylated benzophenones, guttiferone U (1), V (2)/W (3), and a new tocotrienol derivative named globuliferanol (4), along with 11 known compounds (5-15). Their structures were elucidated based on their NMR and MS data. Some isolated compounds were assessed for both their antileishmanial and cytotoxic activities against L. donovani and Vero cells, respectively. Guttiferone K (5) exhibited the best potency (IC50 3.30 µg/mL), but with low selectivity to Vero cells. The n-hexane fraction and some compounds were also assessed in vitro for their antibacterial activity against seven bacterial strains. All the samples exhibited moderate to potent antibacterial activity (MICs ≤ 15.6 µg/mL) against at least one of the tested strains.

Constituents of the Stem Bark of Trichilia monadelpha (Thonn.) J. J. De Wilde (Meliaceae) and Their Antibacterial and Antiplasmodial Activities.

The chemical investigation of the EtOH extract from the stem bark of Trichilia monadelpha (Thonn.) J. J. De Wilde afforded two new limonoids (1 and 2): 24-acetoxy-21,25-dihydroxy-21,23-epoxytirucall-7-en-3-one (1) and (6R)-1-O-deacetylkhayanolide E (2), together with eleven known compounds (3-13), including additional limonoids, flavonoids, triterpenoids, steroids, and fatty acid. Their structures were determined using 1D- and 2D-NMR experiments, ESI mass spectrometry, and single crystal X-ray diffraction analysis. The antibacterial and antiplasmodial activities of the extracts, sub-extracts, fractions, and some of the isolated compounds were evaluated in known pathogenic strains, including Staphylococcus aureus and Plasmodium falciparum. Fraction E (n-Hex/EtOAc 30:70, v/v) showed significant activity against S. aureus ATCC 25923 with a MIC value of 3.90 µg/mL, while one of its constituents (epicatechin (9)) exhibited significant activity with MIC values of 7.80 µg/mL. Interestingly, grandifotane A (6) (IC50 = 1.37 µM) and khayanolide D (5) (IC50 = 1.68 µM) were highly active against the chloroquine-sensitive/sulfadoxine-resistant plasmodium falciparum 3D7 strain, unlike their corresponding plant extract and fractions.

Bioactive constituents from Flacourtia vogelii Hook.f. (Flacourtiaceae).

The chemical investigation of the methanolic root extract of Flacourtia vogelii led to the isolation of a new arylbenzoate derivative, vogelinal (1), together with thirteen known compounds (2-14). The structures of the isolates were elucidated by extensive spectroscopic and spectrometric analyses (1D and 2D NMR, ESI-MS) and by comparison with previously reported data. All the compounds were tested for their antioxidant, antifungal and antibacterial activities. Compound 7 exhibited the highest antioxidant potential, with RSa50of 11.80 ± 2.13 µg/mL, RSa50of 42.60 ± 6.32 µg/mL and RC50 of 51.60 ± 7.71 µg/mL for the DPPH, ABTS and FRAP assay, respectively. Compound 13 displayed weak antifungal effect with MIC value of 125 µg/mL against Candida parapsilosis. Compound 8 showed weak antibacterial effect with MIC value of 125 µg/mL, against Shigella dysenteria. The present study, conclude that this species could be a promising source of antioxidant and antibacterial constituents.

Coumarinolignoid and Indole Alkaloids from the Roots of the Hybrid Plant Citrus × paradisi Macfad (Rutaceae).

A phytochemical investigation of the roots of Citrus × paradisi Macfad. (Rutaceae) led to the isolation of two new compounds, namely 1-formyl-5-hydroxy-N-methylindolin-1-ium (1) and decyloxycleomiscosin D (2), along with ten known compounds: 1,1-dimethylpyrrolidin-1-ium-2-carboxylate (3), furan-2,3-diol (4), 5-methoxyseselin (5), umbelliferone (6), scopoletin (7), citracridone I (8), citracridone II (9), citracridone III (10), limonin (11) and lupeol (12). The structures were determined through the comprehensive spectroscopic analysis of 1D and 2D NMR and EI- and ESI-MS, as well as a comparison with the published data. Notably, compounds 3 and 4 from the genus Citrus are reported here for the first time. In addition, the MeOH extract of the roots and compounds 1-7 were screened against the human adenocarcinoma alveolar basal epithelial cell line A549 and the Caucasian prostate adenocarcinoma cell line PC3 using the MTT assay. While the extract showed significant activity, with IC50 values of 35.2 and 38.1 µg/mL, respectively, compounds 1-7 showed weak activity, with IC50 values of 99.2 to 250.2 µM and 99.5 to 192.7 µM, respectively.

Cytotoxicity of the methanol extracts and compounds of Brucea antidysenterica (Simaroubaceae) towards multifactorial drug-resistant human cancer cell lines.

BACKGROUND: Cancer remains a global health concern and constitutes an important barrier to increasing life expectancy. Malignant cells rapidly develop drug resistance leading to many clinical therapeutic failures. The importance of medicinal plants as an alternative to classical drug discovery to fight cancer is well known. Brucea antidysenterica is an African medicinal plant traditionally used to treat cancer, dysentery, malaria, diarrhea, stomach aches, helminthic infections, fever, and asthma. The present work was designed to identify the cytotoxic constituents of Brucea antidysenterica on a broad range of cancer cell lines and to demonstrate the mode of induction of apoptosis of the most active samples. METHODS: Seven phytochemicals were isolated from the leaves (BAL) and stem (BAS) extract of Brucea antidysenterica by column chromatography and structurally elucidated using spectroscopic techniques. The antiproliferative effects of the crude extracts and compounds against 9 human cancer cell lines were evaluated by the resazurin reduction assay (RRA). The activity in cell lines was assessed by the Caspase-Glo assay. The cell cycle distribution, apoptosis via propidium iodide (PI) staining, mitochondrial membrane potential (MMP) through 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolylcarbocyanine iodide (JC-1) staining, and the reactive oxygen species (ROS) via 2´,7´-dichlorodihydrofluoresceine diacetate (H2DCFH-DA) staining, were investigated by flow cytometry. RESULTS: Phytochemical studies of the botanicals (BAL and BAS) led to the isolation of seven compounds. BAL and its constituents 3, (3-(3-Methyl-1-oxo-2-butenyl))1H indole (1) and hydnocarpin (2), as well as the reference compound, doxorubicin, had antiproliferative activity against 9 cancer cell lines. The IC50 values varied from 17.42 µg/mL (against CCRF-CEM leukemia cells) to 38.70 µg/mL (against HCT116 p53-/- colon adenocarcinoma cells) for BAL, from 19.11 µM (against CCRF-CEM cells) to 47.50 µM (against MDA-MB-231-BCRP adenocarcinoma cells) for compound 1, and from 4.07 µM (against MDA-MB-231-pcDNA cells) to 11.44 µM (against HCT116 p53+/+ cells) for compound 2. Interestingly, hypersensitivity of resistant cancer cells to compound 2 was also observed. BAL and hydnocarpin induced apoptosis in CCRF-CEM cells mediated by caspase activation, the alteration of MMP, and increased ROS levels. CONCLUSION: BAL and its constituents, mostly compound 2, are potential antiproliferative products from Brucea antidysenterica. Other studies will be necessary in the perspective of the discovery of new antiproliferative agents to fight against resistance to anticancer drugs.

Curative anti-typhoid effect of Detarium microcarpum Guill. & Perr. (Leguminosae) hydroethanolic extract root bark based-on in vivo and molecular docking analyses.

ETHNOPHARMACOLOGICAL RELEVANCE: Detarium microcarpum is used to treat typhoid fever, a major public health problem, by indigenous population in Africa. Though its preventive activities have been documented, the curative effect is still to be confirmed. AIM OF THE STUDY: This study aimed at evaluating the curative effects of the hydroethanolic extract of Detarium microcarpum root bark on Salmonella typhimurium-induced typhoid in rat and exploring the in-silico inhibition of some bacterial key enzymes. STUDY DESIGN: In vitro antioxydant, in vivo antisalmonella of the extract and in silico molecular docking assay on the isolated compounds were carried out to explore the anti-salmonella effects of Detarium microcarpum. MATERIAL AND METHODS: The in vitro antioxidant properties of the extract were evaluated using DPPH, ABTS and FRAP tests. The anti-salmonella activity of the extract was assessed through feacal sample from Salmonella typhimurium-infected rat cultured in Salmonella-Shigella agar (SS agar) medium. The affinity of isolated compounds (Rhinocerotinoic acid and Microcarposide) from the extract were performed on four key enzymes (Adenylosuccinate lyase, Acetyl coenzyme A synthetase, Thymidine phosphorylase and LuxS-Quorum sensor) using molecular docking simulation to elucidate the molecular level inhibition mechanism. RESULTS: Crude extract of D. microcarpum root bark showed variable activities on DPPH (RSa50: 6.09 ± 1.04 µg/mL), ABTS (RSa50: 24.46 ± 0.27), and FRAP (RSa50: 23.30 ± 0.23). The extract at all the doses exhibited significant healing effect of infected rats, with the complete clearance. The extract restored hematological, biochemical and histological parameters closed to the normal control. The molecular docking results indicates that rhinocerotinoic acid and microcarposide present more affinity to the LuxS-Quorum sensor and Acetyl coenzyme A synthetase protein as compared to the others. CONCLUSION: These results demonstrate potent anti-typhoid activities of the hydroethanolic of Detarium microcarpum root bark extract through antioxidant properties and high inhibitory affinity of its compounds on some bacterial key enzymes that justify its use as traditional medicine to typhoid fever.