Molecular docking and synthesis of caffeic acid analogous and its anti-inflammatory, analgesic and ulcerogenic studies.

A series of caffeic acid (CA) derivatives 7a-j were synthesized via etherification and coupling action and their chemical structures were elucidated spectroscopically. Motivated by the various biological activities displayed by CA derivatives such as anti-inflammatory, antiviral, anticancer and antioxidant and also based on its extensively consumption in the human diet. In the present work, the newly synthesized compounds 7a-j were evaluated for anti-inflammatory and analgesic action and most of them exerted comparable activity to the reference compound celecoxib. Further, ulcer indexes for the most active compounds were calculated and most of them showed less ulcerogenic effect than the reference drug. Among the title series 7a-j, compounds 7f and 7g with electron withdrawing bromo and chloro group respectively, at the para position of the phenoxy ring was showed good activity compared to all other compounds. Interestingly, the COX-I/COX-II activity ratio of potent compounds 7f and7g showed an almost equal inhibitory effect on both isoenzymes. Further, molecular docking studies have been performed for the potent compounds which showed statistically significant result.

Rapid detection of DPP-IV activity in porcine serum: A fluorospectrometric assay.

Dipeptidyl peptidase IV (DPP-IV) is an aminopeptidase that cleaves the N-terminal dipeptide from peptides bearing proline or alanine residues. Currently, DPP-IV activity is quantified by spectrophotometric or fluorometric methods, which employ Gly-Pro-pNA and Gly-Pro-AMC respectively, as substrate. However, these methods require high enzyme and substrate concentrations. In this study, we adapted the DPP-IV fluorospectrometric assay using NanoDrop 3300, which requires only nanogram levels of the enzyme (30 ng crude DPP-IV) and considerably low substrate concentrations (100 µM). Fluorescence measurement required a reaction mixture of only 2 µL, thus eliminating the need for microtiter plates or cuvettes.We employed this assay to demonstrate DPP-IV activity in porcine serum for the first time. The enzymatic activity peaked at pH 8.0 in porcine (84 nM/min), human (87 nM/min) and bovine (89.1 nM/min) sera, with the optimum temperature of 37 °C. The enzyme showed maximum activity upon incubation for 40 min at 37 °C. In contrast, activity in the porcine serum was the highest after incubation for 30 min at the same optimized parameters. The IC50 values of diprotin A against DPP-IV from human, porcine, and bovine sera were 7.83, 8.62, 9.17 µM, respectively. The present assay procedure is a convenient, sensitive, accurate and high-throughput method suitable for primary screening of DPP-IV inhibitors.

Radical scavenging and anti-inflammatory activities of (hetero)arylethenesulfonyl fluorides: Synthesis and structure-activity relationship (SAR) and QSAR studies.

A series of (hetero)arylethenesulfonyl fluorides (1-58) were synthesized and screened for their in vitro antioxidant (DPPH, ABTS and DMPD methods) and anti-inflammatory activities. The results revealed that compounds 4, 15, 16, 24, 25, 26, 38, 39, 40, and 54 exhibited excellent antioxidant activity using all the three performed antioxidant methods, which were superior to the standard antioxidants ascorbic acid and gallic acid. Compounds 6-9, 11, 18, 19, 21, 22, 30, 39, 40, 44, 45, 48-50, 54, 55 and 57 displayed promising anti-inflammatory activity, which were better than the reference drug indomethacin. Preliminary structure-activity relationship (SAR) revealed that compounds containing electron donating (OH and OCH3) groups on the phenyl ring possessed excellent antioxidant properties while compounds containing electron-withdrawing (Cl, NO2, F and Br) groups on the phenyl ring were found to be most potent anti-inflammatory agents. The presence of SO2F group played a crucial role in increases both antioxidant and anti-inflammatory activities.

Discovery of novel arylethenesulfonyl fluorides as potential candidates against methicillin-resistant of Staphylococcus aureus (MRSA) for overcoming multidrug resistance of bacterial infections.

The multidrug-resistant Staphylococcus aureus (MRSA) is one of the most prevalent human pathogens involved in many minor to major disease burdens throughout the world. Inhibition of biofilm formation is an attractive strategy to treat diseases associated with MRSA infection. In the present investigation, a series of functional group diverse (hetero)aryl fluorosulfonyl analogs were designed, synthesized and tested as antibacterial agents against Staphylococcal spp., and as anti-biofilm candidates. Compounds 8, 15, and 67 were found to possess potent in vitro antibacterial activity among this class of sulfonyl fluorides (MIC = 0.818 ±â€¯0.42, 0.840 ±â€¯0.37 and 0.811 ±â€¯0.37 µg/mL respectively). The analogs 8, 15, 36, and 67 exhibited outstanding anti-biofilm properties compared to other available synthetic antibiotics. The efficacy of synthetic analogs displayed membrane-damaging effect and they are also validated by cellular content release assay. The insight physiological changes were explored by studying the intracellular redox activities through changing cyclic voltammetric (CV) method. The compounds 8, 15, 22, 32, 36, 51, and 67 were found to participate in the interfering in the electron transport chain (ETC) of MRSA. The analogs 8, 15, and 67 possess great potentiality for discovery and development of anti-staphylococcal drugs to treat the MRSA infections.

Multi-targeted dihydrazones as potent biotherapeutics.

Hydrazone compounds were considered as a useful moiety in drug design development. Therefore, these studies were aimed at the synthesis of new dihydrazones and were screened for their in vitro H+/K+-ATPase and anti-inflammatory activities. The results revealed that compounds 9 (22 ±â€¯0.62 µg/mL), 10 (26 ±â€¯0.91 µg/mL), 15 (24 ±â€¯0.44 µg/mL), 16 (28 ±â€¯0.63 µg/mL), 17 (12 ±â€¯0.38 µg/mL), 18 (14 ±â€¯0.47 µg/mL), 19 (26 ±â€¯0.54 µg/mL), 20 (16 ±â€¯0.41 µg/mL), 25 (06 ±â€¯0.68 µg/mL) and 26 (08 ±â€¯0.43 µg/mL) showed excellent H+/K+-ATPase activity and their IC50 value were lower than the standard drug Omerazole (48 ±â€¯0.12 µg/mL). Compounds 5 (28 ±â€¯0.65 µg/mL), 6 (24 ±â€¯0.61 µg/mL), 7 (28 ±â€¯0.64 µg/mL), 8 (26 ±â€¯0.45 µg/mL), 11 (30 ±â€¯0.74 µg/mL), 12 (28 ±â€¯0.40 µg/mL), 13 (32 ±â€¯0.24 µg/mL), 14 (30 ±â€¯0.55 µg/mL) and 21 (08 ±â€¯0.47 µg/mL), 22 (12 ±â€¯0.47 µg/mL), 23 (10 ±â€¯0.51 µg/mL) and 24 (14 ±â€¯0.84 µg/mL) showed better anti-inflammatory activity compared to standard indomethacin (44 ±â€¯0.15 µg/mL). The structure activity relationship (SAR) showed that, electron donating groups (OH, OCH3) favored the H+/K+-ATPase and antioxidants activity, whereas, electron withdrawing groups (F, Cl, Br and NO2) favored the anti-inflammatory activity. Furthermore, molecular docking study was performed to investigate the binding interactions of the most active analogs with the active site of H+/K+-ATPase enzyme. Compounds 25 (G-score = -9.063) and 26 (G-score = -8.977) showed the highest docking G-scores for H+/K+-ATPase inhibition activity.

Aryl fluorosulfate analogues as potent antimicrobial agents: SAR, cytotoxicity and docking studies.

A series of aryl fluorosulfate analogues (1-37) were synthesized and tested for in vitro antibacterial and antifungal studies, and validated by docking studies. The compounds 9, 12, 14, 19, 25, 26, 35, 36 and 37 exhibited superior antibacterial potency against tested bacterial strains, while compounds 2, 4, 5, 15, 35, 36 and 37 were found to have better antifungal activity against tested fungal strains, compared to standard antibiotic gentamicin and ketoconazole respectively. Among all the synthesized 37 analogs, compounds 25, 26, 35, 36 and 37 displayed excellent anti-biofilm property against Staphylococcus aureus. The structure-activity relationship (SAR) revealed that the antimicrobial activity depends upon the presence of -OSO2F group and slender effect of different substituent's on the phenyl rings. The electron donating (OCH3) groups in analogs increase the antibacterial activity, and interestingly the electron withdrawing (Cl, NO2, F and Br) groups increase the antifungal activity (except compound 35, 36 and 37). The mechanism of potent compounds showed membrane damage on bacteria confirmed by SEM. Compounds 35, 36 and 37 exhibited highest glide g-scores in molecular docking studies and validated the biocidal property.

Promising bactericidal approach of dihydrazone analogues against bio-film forming Gram-negative bacteria and molecular mechanistic studies.

Gram-negative members of the ESCAPE family are more difficult to treat, due to the presence of an additional barrier in the form of a lipopolysaccharide layer and the efficiency of efflux pumps to pump out the drugs from the cytoplasm. The development of alternative therapeutic strategies to tackle ESCAPE Gram-negative members is of extreme necessity to provide a solution to the cause of life-threatening infections. The present investigations demonstrated that compounds 17, 20, 25 and 26 possessing the presence of electron donating (OH and OCH3) groups on the phenyl rings are highly potent; whereas compounds 9, 10, 15, 16, 18, 33 and 36 showed moderate activity against Gram-negative bacteria. An excellent dose-dependent antibacterial activity was established compared to that of the standard antibiotic ampicillin. Significant anti-biofilm properties were measured quantitatively, showing optical density (O.D) values of 0.51 ± 015, 0.63 ± 0.20, 0.38 ± 0.07 and 0.62 ± 0.11 at 492 nm and the leakage of cellular components by the compounds, such as 17, 20, 25 and 26, increased the O.D. of respective treated samples compared to the control. In addition, the implication of experimental results is discussed in the light of the lack of survivability of planktonic bacteria and biofilm destruction in vitro. These results revealed the great significance of the development of a new generation of synthetic materials with greater efficacy in anti-biofilm properties by targeting to lock the bio-film associated protein Bap in Gram-negative bacteria.

Synthesis, SAR and molecular docking studies of benzo[d]thiazole-hydrazones as potential antibacterial and antifungal agents.

A series of new benzo[d]thiazole-hydrazones analogues were synthesized and screened for their in vitro antibacterial and antifungal activities. The results revealed that compounds 13, 14, 15, 19, 20, 28 and 30 exhibited superior antibacterial potency compared to the reference drug chloramphenicol and rifampicin. Compounds 5, 9, 10, 11, 12, 28 and 30 were found to be good antifungal activity compared to the standard drug ketoconazole. A preliminary study of the structure-activity relationship (SAR) revealed that the antimicrobial activity depended on the effect of different substituents on the phenyl ring. The electron donating (OH and OCH3) groups presented in the analogues, increase the antibacterial activity (except compound 12), interestingly, while the electron withdrawing (Cl, NO2, F and Br) groups increase the antifungal activity (except compound 19 and 20). In addition, analogues containing thiophene (28) and indole (30) showed good antimicrobial activities. Whereas, aliphatic analogues (24-26) shown no activities in both bacterial and fungal stains even in high concentrations (100µg/mL). Molecular docking studies were performed for all the synthesized compounds of which compounds 11, 19 and 20 showed the highest glide G-score.

Caralluma umbellata Peroxidase: Biochemical Characterization and Its Detoxification Potentials in Comparison with Horseradish Peroxidase.

Caralluma umbellata peroxidase (CUP) is an acidic heme-containing protein having a molecular weight of ~42 kDa and is specific to guaiacol. It is not a glycoprotein. It was purified to 12.5-fold purity with 6.16 % yield. Its activity is dependent on hydrogen peroxide and has an optimum pH and temperature of 6.2 and 45 °C respectively. It can decolorize dyes, viz., Aniline Blue, Reactive Black 5, and Reactive Blue 19 but not Congo Red, while HRP can decolorize Congo Red also. It has lignin-degrading potentiality as it can decompose veratryl alcohol. Detoxification of phenol was more by CUP compared to HRP while with p-nitrophenol HRP has a greater detoxification rate. Based on our results, CUP was identified to be capable of oxidizing a variety of hazardous substances and also a lignin-degrading plant biocatalyst.

Synthesis of benzo[d]thiazole-hydrazone analogues: molecular docking and SAR studies of potential H+/K+ ATPase inhibitors and anti-inflammatory agents.

A series of new benzo[d]thiazole-hydrazones were synthesized and characterized by analytical and spectroscopic techniques. All the compounds were screened for their in vitro inhibition of H+/K+ ATPase and anti-inflammatory effects. The results revealed that compounds 6-8, 13-15, 18-20, 22, 23 and 27-30 displayed excellent inhibitory activity against H+/K+ ATPase, and their IC50 values were lower than those of the standard compound omeprazole. Compounds 2-5, 9-12, 28 and 30 exhibited better anti-inflammatory activity in comparison to the standard compound indomethacin. Studies of the structure-activity relationship (SAR) showed that electron-donating groups (OH and OCH3) favored inhibitory activity against H+/K+ ATPase, whereas electron-withdrawing groups (F, Cl, Br and NO2) favored anti-inflammatory activity, and derivatives with both electron-donating (OH and OCH3) and electron-withdrawing (Br) groups (16-18) displayed reasonable activity, whereas aliphatic analogues (24-26) exhibited less activity and heterocyclic analogues (27-30) displayed moderate activity in both biological studies. Molecular docking studies were performed for all the synthesized compounds, among which compounds 19 and 20 exhibited the highest docking scores for inhibitory activity against H+/K+ ATPase, whereas compounds 10 and 12 displayed the highest docking scores for anti-inflammatory activity.

Synthesis and molecular docking studies of xanthone attached amino acids as potential antimicrobial and anti-inflammatory agents.

A series of novel xanthone conjugated amino acids were synthesised and characterised by analytical and spectroscopic methods. All the synthesized analogues (2-23) were screened for their in vitro antimicrobial and anti-inflammatory activities. Compounds 7, 8, 9, 12, 18, 19, 20, 21 and 23 showed excellent antimicrobial activities compared to antibacterial and antifungal reference drugs gentamicin and bavistin, respectively. Compounds 7-12 and 18-23 showed good anti-inflammatory activity compared to a standard drug, indomethacin. The preliminary structure-activity relationship revealed that tryptophan, tyrosine, phenylalanine, proline and cysteine conjugated compounds showed excellent antimicrobial and anti-inflammatory activities. This may be explained by the contribution of aromaticity and hydrophobicity of amino acids. Molecular docking studies were performed for all the synthesised compounds, among which compounds 20, 21 and 23 showed the highest docking scores for antimicrobial activity while compounds 9, 20 and 22 showed the highest docking scores for anti-inflammatory activity. Different amino acids conjugated xanthone derivatives were synthesized and evaluated for their in vitro biological activities. The conjugation was found to play a major role in improving the biological activities of those compounds.

Purification and characterization of novel fibrin(ogen)olytic protease from Curcuma aromatica Salisb.: Role in hemostasis.

BACKGROUND: The proteases from turmeric species have procoagulant and fibrinogenolytic activity. This provides a scientific basis for traditional use of turmeric to stop bleeding and promote wound healing processes. PURPOSE: Our previous studies revealed that fibrinogenolytic action of crude enzyme fraction of Curcuma aromatica Salisb., was found to be more influential than those of Curcuma longa L., Curcuma caesia Roxb., Curcuma amada Roxb. and Curcuma zedoria (Christm.) Roscoe. Hence, the purpose of this study is to purify and characterize protease from C. aromatica and to explore its role in wound healing process. METHODS: The protease was purified by Sephadex G-50 gel permeation chromatography. Peak with potent proteolytic activity was subjected to rechromatography and then checked for homogeneity by SDS-PAGE and native PAGE. Furthermore purity of the peak was assessed by RP-HPLC and MALDI-TOF. The biochemical properties, type of protease, kinetic studies, fibrinogenolytic, coagulant and fibrinolytic activities were carried out. RESULTS: The two proteolytic peaks were fractionated in gel permeation chromatography. Among these, the peak-II showed potent proteolytic activity with specific activity of 10units/mg/min and named as C. aromatica protease-II (CAP-II). This protein resolved into a single sharp band both in SDS-PAGE (reducing and non-reducing) as well as in native (acidic) PAGE. It is a monomeric protein, showing sharp peak in RP-HPLC and its relative molecular mass was found to be 12.378kDa. The caseinolytic and fibrinolytic activity of CAP-II was completely inhibited by phenylmethylsulfonylfluoride (PMSF). The CAP-II exhibited optimum temperature of 45°C and optimum pH of 7.5. The Km and Vmax of CAP-II was found to be 1.616µg and 1.62units/mg/min respectively. The CAP-II showed hydrolysis of all three subunits of fibrinogen in the order Aα>Bß>γ. The CAP-II exhibited strong procoagulant activity by reducing the human plasma clotting time. It also showed fibrinolytic activity by complete hydrolysis of α-polymer and γ-γ dimer present in fibrin. CONCLUSION: The CAP-II is a novel serine protease from C. aromatica, which has been demonstrated to stop bleeding and initiate wound healing through its procoagulant and fibrin(ogen)olytic activities. Our study demonstrates the possible role of CAP-II, as therapeutic enzyme to stop bleeding at the time of wounding.

Comparative analysis of procoagulant and fibrinogenolytic activity of crude protease fractions of turmeric species.

ETHNAOPHARMACOLOGIAL RELEVANCE: Turmeric rhizome is a traditional herbal medicine, which has been widely used as a remedy to stop bleeding on fresh cuts and for wound healing by the rural and tribal population of India. AIM OF THE STUDY: To validate scientific and therapeutic application of turmeric rhizomes to stop bleeding on fresh cuts and its role in wound healing process. MATERIALS AND METHODS: The water extracts of thoroughly scrubbed and washed turmeric rhizomes viz., Curcuma aromatica Salisb., Curcuma longa L., Curcuma caesia Roxb., Curcuma amada Roxb. and Curcuma zedoria (Christm.) Roscoe. were subjected to salting out and dialysis. The dialyzed crude enzyme fractions (CEFs) were assessed for proteolytic activity using casein as substrate and were also confirmed by caseinolytic zymography. Its coagulant activity and fibrinogenolytic activity were assessed using human citrated plasma and fibrinogen, respectively. The type of protease(s) in CEFs was confirmed by inhibition studies using specific protease inhibitors. RESULTS: The CEFs of C. aromatica, C. longa and C. caesia showed 1.89, 1.21 and 1.07 folds higher proteolytic activity, respectively, compared to papain. In contrast to these, C. amada and C. zedoria exhibited moderate proteolytic activity. CEFs showed low proteolytic activities compared to trypsin. The proteolytic activities of CEFs were confirmed by caseinolytic zymography. The CEFs of C. aromatica, C. longa and C. caesia showed complete hydrolysis of Aα, Bß and γ subunits of human fibrinogen, while C. amada and C. zedoria showed partial hydrolysis. The CEFs viz., C. aromatica, C. longa, C. caesia, C. amada and C. zedoria exhibited strong procoagulant activity by reducing the human plasma clotting time from 172s (Control) to 66s, 84s 88s, 78s and 90s, respectively. The proteolytic activity of C. aromatica, C. longa, C. caesia and C. amada was inhibited (>82%) by PMSF, suggesting the possible presence of a serine protease(s). However, C. zedoria showed significant inhibition (60%) against IAA and moderate inhibition (30%) against PMSF, indicating the presence of cysteine and serine protease(s). CONCLUSION: The CEFs of turmeric species exhibited strong procoagulant activity associated with fibrinogenolytic activity. This study provides the scientific credence to turmeric in its propensity to stop bleeding and wound healing process practiced by traditional Indian medicine.

Exploring a new serine protease from Cucumis sativus L.

Coagulation is an important physiological process in hemostasis which is activated by sequential action of proteases. This study aims to understand the involvement of aqueous fruit extract of Cucumis sativus L. (AqFEC) European burp less variety in blood coagulation cascade. AqFEC hydrolyzed casein in a dose-dependent manner. The presence of protease activity was further confirmed by casein zymography which revealed the possible presence of two high molecular weight protease(s). The proteolytic activity was inhibited only by phenyl methyl sulphonyl fluoride suggesting the presence of serine protease(s). In a dose-dependent manner, AqFEC also hydrolysed Aα and Bß subunits of fibrinogen, whereas it failed to degrade the γ subunit of fibrinogen even at a concentration as high as 100 µg and incubation time up to 4 h. AqFEC reduced the clotting time of citrated plasma by 87.65%. The protease and fibrinogenolytic activity of AqFEC suggests its possible role in stopping the bleeding and ensuing wound healing process.

1-(3-Bromo-2-thien-yl)ethanone.

In the title compound, C(6)H(5)BrOS, the non-H and aromatic H atoms lie on a crystallographic mirror plane. In the crystal, mol-ecules are linked into chains propagating along the c axis by inter-molecular C-H⋯O hydrogen bonds.