Alena Komersová

AMPEROMETRIC DETERMINATION OF GLUCOSE WITH AN MnO2 AND GLUCOSE OXIDASE BULK-MODIFIED SCREEN-PRINTED CARBON INK BIOSENSOR

A simple biosensor, constructed by bulk-modification of carbon ink with manganese dioxide as a mediator and glucose oxidase as a biocomponent was investigated for its ability to serve as amperometric detector for glucose in hydrodynamic as well as in flow injection analysis (FIA) mode. The sensor could be operated at a potential of 0.48 V under physiological conditions (0.1 M phosphate buffer, pH 7.5) and exhibited excellent reproducibility and stability. Factors influencing the amperometric response such as injection volume, flow rate and applied working potential were studied in detail. The screen-printed electrode exhibited a linear amperometric increase with the concentration of D-glucose from 2 to 2500 mg L−1 and gave a 3σ detection limit of 0.085 mg L−1. Due to its remarkable stability such a sensor could be operated continuously for more than four weeks or more than 1000 sample injections. No change of signal height could be observed within an operation period of 12 h. The sensor was exploited for FIA-amperometric determination of glucose in beer and wine samples

Biodiesel fuel from rapeseed oil, methanol, and KOH. Analytical methods in research and production

The enumeration of the analytical methods used in the production of biodiesel from rapeseed oil and methanol catalyzed by KOH and published till 1997 is given. Some of our original methods for individual or simultaneous determination of the main components in the reaction mixture are described. All these methods can be also used to analyse the non-equilibrium complex and heterogencous mixture.

New Findings about Ellman’s Method to Determine Cholinesterase Activity

The original Ellman’s spectrophotometrical method for cholinesterase activity determination uses 5,5′-dithiobis-2-nitrobenzoic acid (DTNB, Ellman’s reagent) as a chromogen and records the level of cholinesterase activity as an increase of absorbance at 412 nm. Although this procedure usually poses no problem, exceptions arise when the concentration of DTNB is far higher than the concentration of acetylthiocholine (ATCH). It was found that the ratio of concentrations of DTNB/ATCH is an important parameter for the ATCH hydrolysis course: high excess of DTNB decreases the hydrolysis rate resulting in a lower measured enzyme activity. Our experiments indicate that this influence of DTNB concentration can be explained by the inhibition of ATCH hydrolysis by DTNB.

Kinetics of Total Enzymatic Hydrolysis of Acetylcholine and Acetylthiocholine

Kinetics and the mechanism of total in vitro hydrolyses (i.e. up to the exhaustion of substrate) of acetylcholine and acetylthiocholine by acetylcholinesterase and butyrylcholinesterase were studied in vitro in a batch reactor at 25 °C, pH 8 and ionic strength of 0.11 ᴍ. Every hydrolysis was monitored by 2 - 3 independent analytical methods. All studied types of enzymatic hydrolyses fulfilled the Michaelis - Menten reaction scheme with the irreversible second step. A table of obtained average values of rate constants and estimations of initial molar enzyme concentrations, and discussion of the results are presented.

Trace iron determination in aminoisophthalic acid using differential-pulse cathodic stripping voltammetry at carbon paste electrodes

Application of differential-pulse cathodic stripping voltammetry using a carbon paste electrode (consisting of carbon powder and liquid paraffin) have been investigated for trace determination of iron in 5-aminoisophthalic acid (AIPA). Samples were dissolved in 1 M HC1, pH was adjusted to 4-5 after addition of EDTA. Voltammetric measurements were performed after filtration. No sample decomposition (mineralization) was necessary. The method showed a good linearity between current and concentration from 3 x 10(-7) to 5 x 10(-5) mol dm-3 of iron, with a detection limit of 3 x 10(-7) mol dm-3 (resp. 1 ppm in solid AIPA). The results agreed well to those obtained by atomic absorption spectrometry (AAS) using electrothermic atomisation. For AAS measurement, however, microwave digestion of samples was necessary.

Formulation and dissolution kinetics study of hydrophilic matrix tablets with tramadol hydrochloride and different co-processed dry binders

The aim of this study is to present the possibility of using of co-processed dry binders for formulation of matrix tablets with drug controlled release. Hydrophilic matrix tablets with tramadol hydrochloride, hypromellose and different co-processed dry binders were prepared by direct compression method. Hypromelloses Methocel™ K4M Premium CR or Methocel™ K100M Premium CR were used as controlled release agents and Prosolv® SMCC 90 or Disintequik™ MCC 25 were used as co-processed dry binders. Homogeneity of the tablets was evaluated using scanning electron microscopy and energy dispersive X-ray microanalysis. The release of tramadol hydrochloride from prepared formulations was studied by dissolution test method. The dissolution profiles obtained were evaluated by non-linear regression analysis, release rate constants and other kinetic parameters were determined. It was found that matrix tablets based on Prosolv® SMCC 90 and Methocel™ Premium CR cannot control the tramadol release effectively for >12h and tablets containing Disintequik™ MCC 25 and Methocel™ Premium CR >8h.

Kinetics of 13 new cholinesterase inhibitors

Kinetics of hydrolysis of acetylcholine and acetylthiocholine by two types of acetylcholinesterase and butyrylcholinesterase inhibited by 13 new inhibitors (5 carbamates and 8 carbazates - hydrazinium derivatives) was measured in vitro in a batch reactor at 25 °C, pH 8, ionic strength 0.11 ᴍ and enzyme activity 3.5 U by four nondependent analytical methods. Sevin®, rivastigmin (Exelon®) and galantamin (Reminyl®) served as comparative inhibiting standards. Kinetics of hydrolyses inhibited by all studied carbamates, sevin, carbazates (with exceptions) and rivastigmin (with exceptions) can be simulated by the competitive inhibition model with irreversible reaction between enzyme and inhibitor. Galantamin does not fulfil this model. In positive simulations, the value of inhibition (carbamoylation) rate constant k3 was calculated, describing the reaction velocity between the given enzyme and inhibitor. Physiologically important hydrolyses of acetylcholine catalyzed by acetylcholinesterase from electric eel or bovine erythrocytes and butyrylcholinesterase from horse plasma can be most quickly inhibited by carbamoylation of the mentioned enzymes by the 3-N,N-diethylaminophenyl- N′-(1-alkyl) carbamates 4 and 5. Probably this is due to a long enough hydrocarbon aliphatic substituent (hexyl and octyl) on the amidic nitrogen atom. The tested carbazates failed as inhibitors of cholinesterases. The regeneration ability of the inhibited enzymes was not measured.

Two new methods monitoring kinetics of hydrolysis of acetylcholine and acetylthiocholine.

Hydroxylamine and HPLC methods, measuring in vitro kinetics of enzymatic hydrolysis of acetylcholine or acetylthiocholine by cholinesterases, are described. The hydroxylamine method determines the dependence of substrate concentration vs. time, the HPLC method is able to measure simultaneously the time dependences of substrate and both primary products, choline or thiocholine, and acetic acid. Practical determinations are shown, comparison with known (above all Ellman’s and pH-stat) methods, advantages and disadvantages are discussed.

Compressibility of tableting materials and properties of tablets with glyceryl behenate

Abstract The paper studies the compressibility of directly compressible tableting materials with dry binders, spray-dried lactose and microcrystalline cellulose, and glyceryl dibehenate at various concentrations. Compressibility was evaluated by means of the energy profile of compression and tensile strength of tablets. Release rate of the active ingredient, salicylic acid, from the tablets was also examined. In the case of microcrystalline cellulose, a higher concentration of glyceryl dibehenate increased the strength of tablets, while this did not occur in the case of spray-dried lactose. Increasing concentration of glyceryl dibehenate prolonged the release of salicylic acid; however, no statistically significant difference was found compared to the type of the dry binder used

A Study of Tablets with a Co-Processed Dry Binder Containing Hypromellose and Α-Lactose Monohydrate

The paper examines the co-processed dry binder RetaLac from the aspect of its compressibility and dissolution of the active ingredient from tablets. RetaLac contains α-lactose monohydrate and hypromellose in the identical proportion. The same parameters are tested in the corresponding physical mixtures of Flowlac100 and various types of hypromellose (Metolose 100SR, Metolose 4000SR, Metolose 100000SR) and compared with the substance RetaLac. Compressibility is evaluated by means of the energy profile of compression and tensile strength of tablets. The values of the total energy of compression and plasticity were higher in the substance RetaLac than in the physical mixtures of lactose and various types of hypromellose; tablet strength, on the contrary, was lower. Dissolution profile of the active ingredient from tablets with RetaLac corresponded to the dissolution profile of tablets from a physical mixture of Flowlac 100 and Metolose 4000SR.