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Optical tweezers (OT), or optical traps, are a device for manipulating microscopic objects through a focused laser beam. They are used in various fields of physical and biophysical chemistry to identify the interactions between individual molecules and measure single-molecule forces. In this work, we describe the development of a homemade optical tweezers device based on a cost-effective IR diode laser, the hardware, and, in particular, the software controlling it. It allows us to control the instrument, calibrate it, and record and process the measured data. It includes the user interface design, peripherals control, recording, A/D conversion of the detector signals, evaluation of the calibration constants, and visualization of the results. Particular stress is put on the signal filtration from noise, where several methods were tested. The calibration experiments indicate a good sensitivity of the instrument that is thus ready to be used for various single-molecule measurements.

Terthiophene derivatives attract interest due to their prospective applications in optoelectronic or sensor devices. Due to their nontoxicity they can be considered as suitable candidates in biological applications. Supramolecular organization of the matter is one of the most interesting topics in contemporary materials science. Amphiphilic chromophores based on substituted terthiophenes are capable of self-assembly into supramolecular architectures. In this work, we aim at simulation of the spectral properties of terthiophene with oligo(ethylene oxide) substituents by the methods of quantum chemistry and molecular dynamics (MD). The potential energy surface (PES) of this molecule was determined by the methods of density functional theory (DFT) for the ground state and time-dependent density-functional theory (TD-DFT) for the excited state. MD simulations in water than revealed the most frequented molecular conformations in both these states. Absorption and fluorescence spectra were determined for all these conformations, including the surrounding water molecules, using TD-DFT and averaged over the conformation space to obtain the final absorption and fluorescence spectrum. The calculated spectra were compared with their experimental counterparts and the differences were discussed in context of the supramolecular structure revealed by confocal microscopy. In spite of its simplicity, this approach provides a satisfactory approximation of absorption and fluorescent spectra of these molecules obtained by computational methods.

The work is aimed at the development of the experimental device to study of the motion of different colloidal objects in the combined force field composed of the optical-trapping force originating in the light-intensity gradient and the thermophoretic force caused by the temperature gradient in aqueous solvents. Polystyrene microscopic beads of submicrometer diameters are used as a simple model of the optical tweezers calibration. The methods of generating temperature gradient are under development. A dual optical trap is used as a tool for the investigation of the thermophoretic force acting on the observed particles. Also, the generated optical trap's force constant is calculated by various methods, including Fast Fourier Transform, and the influence of differences in the sample environment and light intensity is compared. Results of these experiments should contribute to the theory of the optical trapping effect, so far not understood in detail, and should open a way for developing novel methods of the investigation of thermophoretic phenomena in a water environment.

High-pressure methods have become an interesting tool of investigation of structural stability of proteins. They are used to study protein unfolding, but dissociation of oligomeric proteins can be addressed this way, too. HIV-1 protease, although an interesting object of biophysical experiments, has not been studied at high pressure yet. In this study HIV-1 protease is investigated by high pressure (up to 600 MPa) fluorescence spectroscopy of either the inherent tryptophan residues or external 8-anilino-1-naphtalenesulfonic acid at 25°C. A fast concentration-dependent structural transition is detected that corresponds to the dimer-monomer equilibrium. This transition is followed by a slow concentration independent transition that can be assigned to the monomer unfolding. In the presence of a tight-binding inhibitor none of these transitions are observed, which confirms the stabilizing effect of inhibitor. High-pressure enzyme kinetics (up to 350 MPa) also reveals the stabilizing effect of substrate. Unfolding of the protease can thus proceed only from the monomeric state after dimer dissociation and is unfavourable at atmospheric pressure. Dimer-destabilizing effect of high pressure is caused by negative volume change of dimer dissociation of -32.5 mL/mol. It helps us to determine the atmospheric pressure dimerization constant of 0.92 μM. High-pressure methods thus enable the investigation of structural phenomena that are difficult or impossible to measure at atmospheric pressure. © 2015 Ingr et al.

High-pressure methods have become attractive tools for investigation of the structural stability of proteins. Besides protein unfolding, dimerization can be studied this way, too. HIV-1 protease is a convenient target of experimental and theoretical high-pressure studies. In this study molecular-dynamics simulations are used to predict the response of HIV-1 protease to the pressure of 0.1 to 600 MPa. The protease conformation of both the monomer and the dimer is highly rigid changing insignificantly with growing pressure. Hydrophobicity of the protease decreases with increasing pressure. Water density inside the active-site cavity grows from 87% to 100% of the bulk water density within the pressure range. The dimer-dissociation volume change is negative for most of the pressure ranges with the minimum of -105 ml mol-1, except for a short interval of positive values at low pressures. The dimer is thus slightly stabilized up to 160 MPa, but strongly destabilized by higher pressures.

Chicken feathers were processed through hydrolysis in two stages and breakdown efficiency was monitored by factor tests. In the first stage, feathers were incubated in an environment of 0.1 % or 0.3 % aqueous solution of KOH (ratio 1:50) for 24 h at 70 degrees C. After adapting pH level to 9, action by proteolytic enzyme polarzyme 12 T (1-5 %, w/w) at temperature 30-50 degrees C broke down the feathers in 4-8 h into keratin hydrolyzate. Hydrolysis of feathers in environment of 0.1 % KOH is quite low (max. 20%). In an environment of 0.3 % KOH the hydrolysis efficiency is more than four-fold. At upper limits of factors under study (8 h hydrolysis at 50 degrees C with 5 % addition of enzyme) 88.7 % feathers were degraded. Test results confirmed that applying proteolytic enzyme polarzyme 12 T achieved efficiency of chicken feather degradation similar to formerly used enzyme Savinase Ultra 16 L.

Poultry feathers make up for as much as 8.5% of chicken weight and represent a considerable amount of almost pure keratin waste which is not being adequately utilized at the present time. The present study dealt with the processing of poultry feathers through a two-stage alkaline-enzymatic hydrolysis. In the first stage, feathers were mixed with a 0.1 or 0.3% KOH water solution in a 1 : 50 ratio and were incubated at 70 degrees C for 24 h. After adjusting pH to 9, the effects examined in the second processing stage on the amount of degraded feathers were those of proteolytic enzyme additions (1-5%), time (4-8 h) and temperature (50-70 degrees C). Processing feathers in 0.3% KOH and hydrolysing for 8 h in the second stage at 70 degrees C with a 5% dose of enzyme (relative to dry feathers weight) produced approx. 91% degradation. Keratin hydrolysate is distinct for its high nitrogen content and reasonable inorganic solids level. Two-stage technology of alkaline-enzymatic hydrolysing of poultry feathers in an environment of 0.3% KOH achieves high efficiency under quite mild reaction conditions (temperature not exceeding 70 degrees C with pH in a mildly alkaline region), and is feasible from an economic viewpoint. Keratin hydrolysate can find particular application in packaging technology (films, foils and encapsulates).

Cílem práce byla příprava biodegradabilních a jedlých filmů ze škrobo-proteinového hydrolysátu amarantové mouky a studium vlivu chemické a fysikální modifikace na rozpustnost filmů. Filmy byly připraveny litím z 25 % (w/w) roztoku hydrolysátu s přídavkem 20 % glycerinu a s různými přídavky dialdehydu škrobu (0, 1a 5 %). Po tepelné exposici filmů při 65 a 95 °C (doba 6 a 48 hodin) byly provedeny zkoušky rozpustnosti filmů ve vodě při 6 a 37 oC. Bylo zjištěno, že chemickou a fysikální modifikací filmů lze výrazně měnit jejich rozpustnost ve vodě. Výsledky zkoušek rozpustnosti filmů mohou posloužit při praktickém využití škrobo-proteinového hydrolysátu amarantové mouky k výrobě biodegradabilních obalových materiálů (filmy, fólie, mikrokapsule) v potravinářství, ve farmacii, v kosmetice či v zemědělství., The objective of the work was to prepare biodegradable and edible films based on amaranth flour starch-protein hydrolysate and evaluate the effect of chemical and physical modifications on solubility properties of films. Films were prepared by casting a 14 % (w/w) water solution of hydrolysate containing 20 % (per hydrolysate dry matter) glycerol and 0, 1 or 5 % (per hydrolysate dry matter) additions of dialdehyde starch. After thermal exposure of films at 65 or 95 oC (for 6 or 48 hours) film solubility tests were performed in water at 6 or 37 oC. It was found that chemical modification (added dialdehyde starch) and physical modification (thermal exposure) of films allows to markedly altering their water solubility. Results of water solubility tests can serve to practically utilising starch-protein hydrolysate of amaranth flour for producing biodegradable packaging materials (foils, coatings) to be employed in the food industry, cosmetics or agriculture.

Práce se zabývá vlivem síťování výšemolekulárními glykol diepoxidy (a glycerol triglycidyleterem) na biologický anaerobní rozklad proteinového hydrolyzátu (značky Hykol) vyrobeného z chromo-činěných postružin. Vzorky Hykolu síťované vybranými výšemolekulárními glykol diepoxidy byly anaerobně degradovány kalem čistírny městských odpadních vod při 35o C. Biodegradací testovaných vzorků vznikalo sledované množství bioplynu (CH4, CO2). Byla popsána příprava, stanovení rozpustnosti, limitní viskozitní čísla a IR spektra připravených vzorků. Ve srovnání s Hykolem stupeň degradace se zmenšil cca na 70-50%., This work focuses on the influence of cross-linking with higher-molecular glycol diepoxides (and glycerol triglycidyleter) on biologic anaerobic degradation of protein hydrolysate (trade name Hykol) produced from chrome-tanned wastes (shavings). Samples of Hykol cross-linked with selected higher-molecular diepoxides were degraded with anaerobic sludge from a municipal plant at temperature 35o C. Biodegradation of tested modified hydrolysate samples produced recorded quantities of biogas (CH4, CO2). Attention was also given to preparation, the determination of solubility, intrinsic viscosity and IR spectra of prepared samples. In comparision with Hykol the degree of degradation decreased approx. from 70-50%.

Koželužský průmysl zabývající se zpracováním surových kůží a jejich přeměnou na usně produkuje značné množství tuhých a kapalných odpadů. Rizikové jsou především tuhé činěné odpady obsahující chróm a všechny druhy odpadních lázní obsahujících zbytkový chróm. Aby se zabránilo ztrátám bílkovinného odpadu a únikům sloučenin chrómu, byly vyvinuty recyklační technologie pro kapalné i tuhé odpady. Recyklace tuhých činěných odpadů je úspěšně realisována enzymovou hydrolýsou. Problémem zůstává tuhý zbytek (chromitý koláč) obsahující MgO, který byl použit jako promotor při hydrolyse chromočiněného tuhého odpadu. Vysoký obsah MgO v chromitém kalu komplikuje jeho další využití. Předložená práce se zabývá možnostmi isolace hořčíku, který je obsažen ve formě jeho hydroxidu z chromitého koláče s využitím odpadního činícího koželužského roztoku. Výsledky ukázaly že je možné dosáhnout až 70 % účinnosti odstranění Mg., The tanning industry processes raw hides and converts them into leather, producing considerable quantities of solid and liquid by-products. The solid tanned by-products contain chromium and by-product liquors contain residual chromium. In order to prevent loss of protein material and chrome compounds, technologies were developed for recycling liquid and solid by-products. Recycling solid by-products is being successfully realized through enzymatic hydrolysis. The chrome cake remains an issue as it contains MgO which was used as a promoter in hydrolysis of chrome-tanned solid by-products. High content of MgO in the chrome cake complicates its further use. This presents work addresses the possibility of isolating magnesium, present as hydroxide, from the chrome cake using by-product (spent) tanning liquor. Results showed that it is possible to achieve magnesium removal efficiency up to 70 %.

Bylo studováno sítování hydrolyzátu kolagenu diepoxibutanem nebo (oktanem) při 25oC. Optimální pH pro sítování bylo 9. Rozpustnost vzorku klesala úměrně s množstvím použitého sítovadla. Pro stanovení limitního viskozitního čísla je optimální měřit viskozity vzorku v 1M KCl roztoku. Diepoxyoktan byl účinnější sítovadlo než diepoxybutan. V IR spektrech se sítování projevilo slabě a jen při použití diepoxyoktanu., Colagen hydrolyzate was crosslinked with diepoxibutane or (octane) at 25oC. Suitable conditions for crosslinking were found at pH = 9. Solubility of samples decreased proportionately with increasing concentration of crosslinking agent. Viscosity measurements yielded best results for estimation of intrinsic viscosity in 1M KCl solution. Diepoxyoctane was better crosslinking agent than diepoxybutane. In IR spectra crosslinking demonstrated only weakly with diepoxyoctane.

Deprotenizace chromitého koláče získaného po enzymatické hydrolýze chromočiněných postružin byla provedena alkalickou hydrolýzou v tlakové nádobě roztokem KOH. Kinetika podle provedených výpočtů odpovídá s dobrou shodou reakci II. řádku; efektivní hodnota rychlostní konstanty k= 8,9.10 na -3 l /(mol.s) a aktivační energie Ea = 130kJ/mol při teplotě hdrolýzy 116°C., After enzymatic hydrolysis chrome-tanned shavings and subsequent filtration, cake is formed, composed of hydrated Cr2O3 with protein products of hydrolysis as well as with non-hydrolysable fraction. After hydrolysis with KOH was evaluated hydrolysis kinetics. Formal kinetics of this alkaline hydrolysis, appears to agree with a second-order reaction, with effective values of rate constant c=3.9 .10 na-3 l/(mol.s) and activation energy Ea = 130 kJ/mol at a temperature of 116°C.

Biologické testy byly prováděny ve vodním prostředí za anaerobních podmínek. Po síťování s 1-13 hm. % diepoxybutanu (oktann) biologická rozložitelnost klesla ze 72% na 23% úměrně s hustotou vytvořené sítě., Biodegradability testing was performed in aqueos environment under anaerobic conditions. After crosslinking with 1-13 weight % diepoxybutane (octane) the degradability dropped from 72% to 23% apparently owing tu built-up a network forming.

Práce byla zaměřena na vliv síťování bílkovinného hydrolyzátu a polyethylenglykol diepoxidy (a glycerol triglycidyl etheru) na jeho anaerobní rozklad. Biologické testy byly zaměřeny na bilanci množství vznikajícího bioplynu (CO2 + CH4). Koncentrace inokula byla 4-5%, teplota 35 st.Celsia. Vzorky Hykolu síťované vybranými diepoxidy byly nasazovány v množství 200mg/l. Rychlost anaerobního rozkladu byla nejméně snížena při síťování pomocí polyethylen glykol (600) diglycidyl etheru a nejvíce při aplikaci glycerol triglycidyl etheru., The work focuses on influence exerted by cross-linking with polyethylene glycol diepoxides (and glycerol triglycidyl ether ) on biologic anaerobic degradation of protein hydrolyzate (trade name Hykol). Biodegradation of tested hydrolysate samples produced recorded quantities of bio-gas (CH4, CO2). Anaerobic degradation rate of samples was least reduced when polyethylene glycol (600) diglycidyl ether was employed as cross-linking agent, and most reduced when glycerol triglycidyl ether was applied.

Byl zpracován matematický model acidobazické rovnováhy CO2 v systému plyn - kapalina umožňující stanovit množství produkovaného CO2 chromatografickou analýzou plynné fáze. Za vytipovaných podmínek byla sledována biologická rozložitelnost modelových vzorků, průběh biorozkladu byl sledován podle produkce CO2 stanovované GC analýzou plynné fáze a standardním "titračním" postupem. Jedná se o "zastupitelné" postupy sledování průběhu aerobního biologického rozkladu látek ve vodním prostředí., A mathematical model of acid-basic CO2 equilibrium in a gas - liquid system was worked out, enabling to determine quantity of produced CO2 through chromatographic analysis of gaseous phase. Under appropriate test conditions, biodegradability of model samples was explored. Course of biodegradation was studied through produced CO2 determined by analyzing gaseous phase through GC on the one hand, and through customary "titration" procedure on the other. The procedures in question are "substituting" procedures for observing course of aerobic biodegradation of substances in an aqueous environment.

Cílem práce bylo posouzení možnosti využití plynově chromatografické analýzy plynné fáze pro hodnocení aerobní biologické rozložitelnosti látek ve dvoufázovém systému plyn-kapalina. Příspěvek je rozdělen do dvou částí: v první části je diskutována acidobazická rovnováha CO2 v reálných podmínkách biologických testů; druhá část příspěvku pak zahrnuje experimentální ověření teoretických závěrů bilance CO2., The objective of this study was to verify potential use of chromatographic analysis of gaseous phase composition in an enclosed two-phase gas-liquid system to study course of biodegradation. This contribution is divided into the two parts: in the first part acidobasic equilibrium states of CO2 in a real biological system is discussed; the second part of this contribution includes experimental verification of the theoretical conclusions.

The authors have assessed various sources of recycled PET (washed dry flakes, ground preforms etc.) using the proper methods (TG, viscometry, filtration test of the melt, DSC, ESM etc.). They have evaluated the most harmful impurities. The data presented could help those interested in the other potential applications of the recycled PET materials. This publication is one of the results based on the Grant sponsored by Czech Ministry of Industry and Trade "The new possibilities of the textile application of the recycled PET bottles".