M. I. Choudhary

Chemistry and Mechanism of Urease Inhibition

Studies on enzyme inhibition remain an important area of pharmaceutical research since these studies have led to the discoveries of drugs useful in a variety of physiological conditions. The enzyme inhibitors can interact with enzymes and block their activity towards natural substrates. Urease inhibitors have recently attracted much attention as potential new anti-ulcer drugs. Ironically, urease was the first enzyme crystallized but its mechanism of action is still largely misunderstood. This chapter therefore reviews comprehensive developments in the field of urease inhibitors. Inhibitors of urease can be broadly classified into two categories: (1) active site directed (substrate-like), (2) mechanism-based directed. We present here the examples of selected inhibitors along with their mechanisms of action to characterize their mode of urease inhibition. The observations that urease due to its high substrate (urea) specificity can only bind to a few inhibitors with a similar binding mode as urea is also discussed. Several non-covalent interactions including hydrogen bonds and hydrophobic contacts stabilize the enzyme-inhibitor complex. Regardless of the class of compound, it is reported that only a few functional groups with electronegative atoms such as oxygen, nitrogen and sulfur act either as bidentate (mostly), tridentate (rarely), or as ligand-chelator to form octahedral complexes with two slightly distorted octahedral Ni ions of the enzyme. Bulky groups attached to the pharmacophore were found to decrease the activity of inhibitors, since the lack of a bulky attachment makes it easier for urease inhibitors to enter the substrate-binding pocket as well as avoid unfavorable steric interactions with amino acid residues in its vicinity. This review is intended to provide highlights of the inhibition of urease by hydroxamic acids (HXAs), phosphorodiamidates (PPDs), imidazoles, phosphazene and related compounds. These compounds are compared to previously reported urease inhibitors for the catalytic models proposed for urease activity. The differences in inhibition of urease activities from plants and of bacterial origin by various inhibitors and physiological implications of urease inhibition are discussed.

Quantitative HPLC analysis of withanolides in Withania somnifera

One of the most widely used herbs in Ayurvedic medicine is Ashwaghanda, Withania somnifera, a shrub commonly found on the Indian subcontinent. As this plant is increasingly becoming a popular adaptogenic in the western world, analytical methods for its identification and quality control are in demand. Thus, a HPLC method for the determination of withaferin A and withanolide D was developed. The system was successfully used to investigate the presence of the markers in different W. somnifera plant parts as well as to analyze their content in market products.

Synthesis and in vitro urease inhibitory activity of N,N'-disubstituted thioureas.

Thiourea derivatives (1-38) were synthesized and evaluated for their urease inhibition potential. The synthetic compounds showed a varying degree of in vitro urease inhibition with IC50 values 5.53 ± 0.02-91.50 ± 0.08 μM, most of which are superior to the standard thiourea (IC₅₀ = 21.00 ± 0.11 μM). In order to ensure the mode of inhibition of these compounds, the kinetic study of the most active compounds has been carried out. Most of these inhibitors were found to be mixed-type of inhibitors, except compounds 13 and 30 which were competitive, while compound 19 was identified as non-competitive inhibitor with Ki values between 8.6 and 19.29 μM.

Protein glycation inhibitory activities of Lawsonia inermis and its active principles

The protein glycation inhibitory activity of ethanolic extract of Lawsonia inermis (henna) plant tissues was evaluated in vitro using the model system of bovine serum albumin and glucose. Protein oxidation and glycation are posttranslational modifications that are implicated in the pathological development of many age-related disease processes. This study investigated the effects of Lawsonia inermis ethanolic extract and its components, on protein damage induced by a free radical generator in in vitro assay system. We found that alcoholic extract of Lawsonia inermis can effectively protect against protein damage and showed that its action is mainly due to Lawsone. In addition, the presence of gallic acid also plays an important role in the protective activity against protein oxidation and glycation. Two known compounds, namely, Lawsone and gallic acid previously isolated from this plant were subjected to glycation bioassay for the first time. It was found that the alcoholic extract, lawsone (1) and gallic acid (2) showed significant inhibition of Advanced Glycated End Products (AGEs) formation and exhibit 77.95%, 79.10% and 66.98% inhibition at a concentration of 1500 μg/mL, 1000 μg/mL and 1000 μM respectively. Lawsonia inermis, compounds 1 and 2 were found to be glycation inhibitors with IC50 82.06 ± 0.13 μg/mL, 67.42 ± 1.46 μM and 401.7 ± 6. 23 μM respectively. This is the first report on the glycation activity of these compounds and alcoholic extract of Lawsonia inermis.

Anticonvulsant activities of ethanolic extract and aqueous fraction isolated from Delphinium denudatum

Dried roots of Delphinium denudatum Wall. are a popular folk remedy for the treatment of epilepsy in the traditional Unani system of medicine in the sub-continent. We carried out anticonvulsant screening of the ethanolic extract (EE) and aqueous fraction (AF) of this plant utilising the maximal electroshock (MEST) and subcutaneous pentylenetetrazole (scPTZ), bicuculline (scBIC), picrotoxin (scPTX) and strychnine (scSTN) tests for anticonvulsant activity. EE had weak dose-dependent anticonvulsant effects on seizures induced by PTZ and BIC. AF exhibited dose-dependent activity against hind limb tonic extension phase (HLTE) of MEST and comparatively stronger anticonvulsant activity against seizures induced by PTZ and BIC. The results suggest the presence of potent anticonvulsant compounds in AF of D. denudatum and deserve further investigation for isolation of active compounds and elucidation of the mechanism of anticonvulsant action.

Minor secondary metabolic products from the stem bark of Plumeria rubra Linn. displaying antimicrobial activities.

Four new iridoids viz., plumeridoids A, B, and C and epiplumeridoid C were isolated from the stem bark of Plumeria rubra Linn. together with twenty-four known compounds viz., 1-( P-hydroxyphenyl)propan-1-one, isoplumericin, plumericin, dihydroplumericin, allamcin, fulvoplumerin, allamandin, plumieride, P- E-coumaric acid, 2,6-dimethoxy- P-benzoquinone, scopoletin, cycloart-25-en-3 beta,24-diol, 2,4,6-trimethoxyaniline, ajunolic acid, ursolic acid, oleanolic acid, beta-amyrin acetate, betulinic acid, lupeol and its acetate, 2,3-dihydroxypropyl octacosanoate, glucoside of beta-sitosterol, and a mixture of common sterols (stigmasterol and beta-sitosterol). Their structures were determined by means of spectroscopic data including HREIMS, 1H NMR, 13C NMR, 2D NMR (HMQC, HMBC, NOESY) and by comparison with published data. All but one of thirteen tested compounds exhibited antifungal, antialgal, and/or antibacterial activities.

Synthesis of some N4-substituted isatin-3-thiosemicarbazones

In this article, we describe a simple and new method for the synthesis of some N4-substituted isatin-3-thiosemicarbazones based on the reactions of the common intermediate, methyl 2-(2-oxo-1,2-dihydro-3H-indol-3-ylidene)-1-hydrazinecarbodithioate, prepared by condensing isatin with methyl 1-hydrazinecarbodithioate, and the readily available amines in essentially a one-step reaction. The synthesized thiosemicarbazones were fully characterized by their physical, analytical, and spectral (IR, 1H-NMR, EIMS) data.

New diterpenoid alkaloids from Aconitum heterophyllum Wall: Selective butyrylcholinestrase inhibitors.

Two new diterpenoid alkaloids, heterophyllinine-A (1) and heterophyllinine-B (2), along with two known alkaloids dihydroatisine (3) and lycoctonine (4) were isolated from the roots of Aconitum heterophyllum Wall. The structure of (1) and (2), were deduced on the basis of spectral data. Compounds 1–2 inhibited acetylcholinesterase (AChE, EC 3.1.1.7) and butyrylcholinesterase (BChE, EC 3.1.1.8) enzymes in a concentration-dependent manner with percent inhibition ranging between 4.24% and 6.94 % and 79.1 % and 82.75 % for AChE and BChE, respectively indicating that compounds 1 and 2 are about thirteen times more specific to BChE than AChE.

Synthesis and inhibitory potential towards acetylcholinesterase, butyrylcholinesterase and lipoxygenase of some variably substituted chalcones

A series of variably substituted chalcones were synthesized by condensation of substituted acetophenones with mono-, di- or trisubstituded benzaldehydes. It was observed that some of these compounds have the potential to inhibit acetylcholinesterase, whereas others show activity against butyrylcholinesterase, depending on the substitution pattern at the two aromatic rings of these chalcones. Similarly, lipoxygenase was inhibited by two of these compounds. It has been observed that inhibition of the three enzymes was concentration dependent with the IC50 values ranging from 28.2–134.5 μM against acetylcholinesterase, 16.0–23.1 μM against butyrylcholinesterase and 57.6–71.7 μM against lipoxygenase, respectively.

In vitro inhibition of pentylenetetrazole and bicuculline-induced epileptiform activity in rat hippocampal pyramidal neurons by aqueous fraction isolated from Delphinium denudatum.

Roots of Delphinium denudatum W. are used for the treatment of epilepsy by traditional healers in subcontinent. Aqueous fraction (AF) isolated from D. denudatum has previously shown significant anticonvulsant activity in in vivo and in vitro models of seizures. We investigated anticonvulsant effects of AF on pentylenetetrazole (PTZ) and bicuculline (BIC)-induced epileptiform activity in primary hippocampal neuronal cultures. Electrophysiological studies on single pyramidal neurons were carried out by using whole-cell current clamp technique. Introduction of AF (0.6 mg/ml) in perfusate blocked PTZ (10 mM) and BIC (100 micro M)-induced epileptiform activity comprising of paroxysmal depolarization shifts (PDS). The PDS were elicited again when AF was removed from perfusate. We conclude that AF contains anticonvulsant compounds that possibly interact with GABA(A) receptor to produce blockade of epileptiform activity. Further studies on isolation of compounds from AF may lead to discovery of new class of anticonvulsants.