Histochemical methods allow for identification and localization of various components within the tissue. Such information on the spatial heterogeneity is not available with biochemical methods. However, there is limitation of the specificity of such detection in context of complex tissue, which is important to consider, and interpretations of the results should regard suitable control treatments if possible. Such methods are valuable extension to specific optical and spectroscopic analytical methods. Here we present a set of selected simple methods of staining and histochemical tests with comments based on our laboratory experience.
- MeSH
- barvení a značení metody MeSH
- barvicí látky analýza MeSH
- buněčná stěna chemie ultrastruktura MeSH
- celulosa analýza MeSH
- histocytochemie metody MeSH
- lignin analýza MeSH
- lipidy analýza MeSH
- mikroskopie metody MeSH
- pektiny analýza MeSH
- rostliny chemie ultrastruktura MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
OBJECTIVES: The aim of the study was to use methods of pharmaceutical technology, and prepare carriers in the form of pellets suitable as a filling of detection tubes for enzymatic detection of cholinesterase inhibitors. The enzymatic detection was based on enzymatic hydrolysis of acetylthiocholine iodide and the subsequent colour reaction of its hydrolysis product with Ellman's reagent. The suitable carriers should be in the form of white, regular and sufficiently mechanically resistant particles of about 1 mm allowing it to capture the enzyme during the impregnation process and ensuring its high activity for enzymatic detection. METHODS: Carriers consisting of microcrystalline cellulose, lactose, povidone, and sodium carboxymethyl cellulose were prepared using extrusion-spheronization method under three different drying conditions in either a hot air oven or a microwave oven. Subsequently, the carriers were impregnated with acetylcholinesterase and their size, shape, mechanical resistance, bulk, tapped and pycnometric density, Hausner ratio, intraparticular and total tapped porosity, and activity were measured and recorded. RESULTS: In this procedure, carriers with different physical parameters and different acetylcholinesterase activity were evaluated. It was found that higher acetylcholinesterase activity was associated not only with a higher intraparticular porosity but also with more regular particles characterized by high sphericity and low total tapped porosity. CONCLUSION: This unique finding is important for the preparation of detection tubes based on enzymatic detection which is still irreplaceable especially in the field of detection and analysis of super-toxic cholinesterase inhibitors.
- MeSH
- acetylthiocholin analogy a deriváty metabolismus MeSH
- celulosa analýza MeSH
- cholinesterasové inhibitory metabolismus MeSH
- kyselina dithionitrobenzoová MeSH
- laktosa analýza MeSH
- poréznost MeSH
- povidon analýza MeSH
- sodná sůl karboxymethylcelulosy analýza MeSH
- sulfhydrylová reagencia MeSH
- testování materiálů MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
A sensitive and specific high performance liquid chromatography method for the separation and determination of tapentadol enantiomers has been developed and validated. Ten different chiral columns were tested in a normal phase system. Excellent enantioseparation with the resolution more than 2.5 for all enantiomers was achieved on Chiralpak AD-H using mixture of heptane-propan-2-ol-diethylamine (980:20:1, v/v/v). The detection was carried out using fluorescence detector at excitation wavelength of 295 nm and emission wavelength of 273 nm. The influence of mobile phase composition, namely organic modifiers, additives, aliphatic alkanes and water content in mobile phase, on retention and enantioseparation was studied. Validation of the developed method including linearity, limit of detection, limit of quantification, precision, accuracy and selectivity was performed according to the International Conference on Harmonization guidelines. The advantage of the method is a good enantioseparation, short analysis time (less than 20 min) and therefore this method is suitable for routine determination of chiral purity of (R,R)-tapentadol in enantiopure active pharmaceutical ingredient.
V práci jsou mikrokrystalické celulosy porovnány pomocí parametrů trojexponencinální rovnice V/V0 = A1*exp(-P/t1) + A2*exp(-P/t2) + A3*exp(-P/t3) + y0 1). Proces lisování pomocí této rovnice probíhá ve třech fázích, ve fázi redukce interpartikulárních pórů, redukce intrapartikulárních pórů a ve fázi redukce tuhé látky bez pórů. Klasická teorie lisování vychází ze skutečnosti, že jednotlivé procesy probíhají v rozmezí konkrétních lisovacích tlaků. Podle nových rovnic hodnocené procesy probíhají souběžně v celém použitém rozsahu lisovacích tlaků. V práci byly studovány čtyři mikrokrystalické celulosy, Avicely PH 102, PH 103, PH 105 a PH 301. Tyto pomocné látky se liší velikostí a tvarem částic, hustotou a obsahem vlhkosti. Byly hodnoceny velikosti redukcí AR, velikosti energií EA a „půltlaky“ PH. Z výsledků práce vyplynulo, že se zmenšováním částic mikrokrystalických celulos se energie EA snižují, současně se „půltlaky“ zvyšují. Nejlepší lisovatelnost má Avicel PH 102 s hodnotou „půltlaku“ 119,954 MPa a nejhorší lisovatelnost Avicel PH 301 s hodnotou „půltlaku“ 151,449 MPa.
The study compares microcrystalline celluloses by means of the parameters of the three-exponential equation V/V0 = A1*exp(-P/t1) + A2*exp(-P/t2) + A3*exp(-P/t3) + y0 1). The process of compression, according to this equation, takes place in three stages: in the stage of reduction of interparticular pores, reduction of intraparticular pores, and in the stage of reduction of the solid substance without pores. The classic theory of compression is based on the fact that the individual processes take place within the ranges of concrete compression pressures. According to the new equations, the processes under evaluation take place concurrently within the whole range of compression pressures employed. The paper studied four microcrystalline celluloses, Avicels PH 102, PH 103, PH 105, and PH 301. These excipients differ in sizes and shapes of particles, density, and content of humidity. The evaluations included the sizes of reductions AR, sizes of energies EA, and “half-pressures” PH. The results of the study show that with diminishing particles of microcrystalline celluloses energies EA are decreased and “half-pressures” are simultaneously increased. Best compressibility was found in Avicel PH 102 with the value of the “half-pressures” of 119.954 MPa and the worst in Avicel PH 301 with the value of the “half-pressures” of 151.449 MPa.
