Inhibition of Cholinesterase by Dialkylcarbamates
Abstract The pI50 index and separation coefficients of chosen 3-N,N-diethylaminophenyl-N′,N′-dialkylcarbamates were determined. Index pI50 (pI50 = negative logarithm of molar concentration of inhibitor inhibiting the enzyme activity by 50%) describes the effectiveness of the inhibitor. The rate of ability of the inhibitor to pass the blood-brain barrier is usually described by the separation coefficient in a system n-octanol/water (Kow). Obtained results were compared with pI50 and Kow of Exelon®, the commercially used drug against the Alzheimer’s disease.
From the New Method for the Determination of the Half Inhibition Concentration ( IC 50 ) of Cholinesterase Inhibitors
The IC50 is the molar concentration of a given inhibitor (I), which reduces the respective uninhibited reaction rate to one half, and is indicative of the inhibitory potency of a compound. We investigated new inhibitors of the in vitro hydrolysis of acetylcholine (ACh) by cholinesterases: acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). Ellman ́s method (Ellman et al., 1961) is routinely used for the determination of the cholinesterase activity and effi ciency of cholinesterase inhibitors. It is based on the measurement of the absorbance at 412 nm of the yellow complex produced by the reaction between thiocholine (TCh) and DTNB (5,5 ́-dithiobis-2-nitrobenzoic acid, Ellman ́s reagent), as described in detail in Zdražilová et al. (2004). Ellman ́s method has two disadvantages: 1) The composition of the yellow complex and its absorption coeffi cient at 412 nm depend strongly on the composition of the reaction mixture, above all on the pH value. 2) Acetylthiocholine (ATCh) instead of ACh must be used as the substrate. However, the reaction rates of the ACh and ATCh hydrolysis by AChE (or BChE) are not identical, nor are the IC50 values (Zdražilová et al., 2006a). When using ACh, the determination of the rate of the enzymatic reaction, and hence also of IC50, requires the quantitative determination of the concentration of one of the reaction components, i.e. ACh, choline (Ch), acetic acid (HAc), or protons from HAc, in the reaction mixture. The suitable methods, e.g. HPLC or pH-stat (Zdražilová et al., 2006b), are experimentally much more sophisticated and time-consuming. This paper describes a new simple, and fast method of IC50 determination with the possibility of using either ACh or ATCh. The method is based on the measurement of changes in the pH value as a function of time during the enzymatic hydrolysis of ACh or ATCh, it is called the pH(t) method.
Rational Design, Synthesis and Evaluation of γ-CD-Containing Cross-Linked Polyvinyl Alcohol Hydrogel as a Prednisone Delivery Platform
This study describes the in-silico rational design, synthesis and evaluation of cross-linked polyvinyl alcohol hydrogels containing γ-cyclodextrin (γ-CDHSAs) as platforms for the sustained release of prednisone (PDN). Through in-silico studies using semi-empirical quantum mechanical calculations, the effectiveness of 20 dicarboxylic acids to generate a specific cross-linked hydrogel capable of supporting different amounts of γ-cyclodextrin (γ-CD) was evaluated. According to the interaction energies calculated with the in-silico studies, the hydrogel made from PVA cross-linked with succinic acids (SA) was shown to be the best candidate for containing γ-CD. Later, molecular dynamics simulation studies were performed in order to evaluate the intermolecular interactions between PDN and three cross-linked hydrogel formulations with different proportions of γ-CD (2.44%, 4.76% and 9.1%). These three cross-linked hydrogels were synthesized and characterized. The loading and the subsequent release of PDN from the hydrogels were investigated. The in-silico and experimental results showed that the interaction between PDN and γ-CDHSA was mainly produced with the γ-CDs linked to the hydrogels. Thus, the unique structures and properties of γ-CDHSA demonstrated an interesting multiphasic profile that could be utilized as a promising drug carrier for controlled, sustained and localized release of PDN.