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Simple analytical devices suitable for the analysis of various biochemical and immunechemical markers are highly desirable and can provide laboratory diagnoses outside standard hospitals. This study focuses on constructing an easily reproducible do-it-yourself ELISA plate reader biosensor device, assembled from generally available and inexpensive parts. The colorimetric biosensor was based on standard 96-well microplates, 3D-printed parts, and a smartphone camera as a detector was utilized here as a tool to replace the ELISA method, and its function was illustrated in the assay of TNFα as a model immunochemical marker. The assay provided a limit of detection of 19 pg/mL when the B channel of the RGB color model was used for calibration. The assay was well correlated with the ELISA method, and no significant matrix effect was observed for standard biological samples or interference of proteins expected in a sample. The results of this study will inform the development of simple analytical devices easily reproducible by 3D printing and found on generally available electronics.

Urine test strips for urinalysis are a common diagnostic tool with minimal costs and are used in various situations including homecare and hospitalization. The coloration scaled by the naked eye is simple, but it is suitable for semiquantitative analysis only. In this paper, a colorimetric assay is developed based on a smartphone digital camera and urine test strips. Assays of pH, albumin, glucose, and lipase activity were performed as a tool for the diagnosis of aciduria, alkaluria, glycosuria, proteinuria, and leukocyturia. The RGB color channels were analyzed in the colorimetric assay, and the assay exerted good sensitivity, and all the particular diagnoses proved to be reliable. The limits of detection for glucose (0.11 mmol/L), albumin (0.15 g/L), and lipase (2.50 U/μL) were low enough to cover the expected physiological concentration, and the range for pH was also satisfactory. The urine test strips with a camera as an output detector proved applicability to spiked urine samples, and the results were also well in comparison to the standard assays which confirms the practical relevance of the presented findings.

Environmental screening is essential due to the increased occurrence of harmful substances in the environment. Open Meter Duo (OMD) is an open-source field photo/fluorimeter that uses an RGB diode that imitates a color according to the selected wavelength and uses a UV LED from the security kit diode as an excitation light source. The prepared PCB shield with a 3D-printed aperture was connected to Arduino UNO R4 WiFi. This system was used for the fluorescent detection of cholinesterase activity with the indoxyl acetate method. Carbofuran—a toxic pesticide—and donepezil—a drug used to treat Alzheimer’s disease—were tested as model inhibitors of cholinesterase activity. The limit of detection of indoxyl acetate was 11.6 μmol/L, and the IC50 values of the inhibitors were evaluated. This system is optimized for wireless use in field analysis with added cloud support and power source. The time of analysis was 5 min for the fluorimetric assay and 20 min for the optional photometric assay. The time of field operation was approximately 4 h of continuous measurement. This system is ready to be used as a cheap and easy control platform for portable use in drug control and point-of-care testing.

Chemical, Biological, Radiological and Nuclear (CBRN) weapons still represent a relevant threat despite international treaties and regulations. International projects like the Permanent Structured Cooperation (PESCO) of European Union (EU) appears as a suitable way of the further development when considered both complexity and expensiveness of the countermeasures to the CBRN. In this work, basic principles of PESCO, limitations, opportunities and expected directions of innovations were searched. The final recommendations on PESCO projects in the CBRN field are founded on the reached findings. Recommendations and conclusions on limited PESCO efficacy on one side and opportunity to resolve weakness of the Army of the Czech Republic (AČR) in the CBRN by collaboration in the EU on the other were particularly described in this paper. Without the collaboration, the AČR will hardly reflect the progress on CBRN defense.

Excessive use of pesticides could potentially harm the environment for a long time. The reason for this is that the banned pesticide is still likely to be used incorrectly. Carbofuran and other banned pesticides that remain in the environment may also have a negative effect on human beings. In order to provide a better chance for effective environmental screening, this thesis describes a prototype of a photometer tested with cholinesterase to potentially detect pesticides in the environment. The open-source portable photodetection platform uses a color-programmable red, green and blue light-emitting diode (RGB LED) as a light source and a TSL230R light frequency sensor. Acetylcholinesterase from Electrophorus electricus (AChE) with high similarity to human AChE was used for biorecognition. The Ellman method was selected as a standard method. Two analytical approaches were applied: (1) subtraction of the output values after a certain period of time and (2) comparison of the slope values of the linear trend. The optimal preincubation time for carbofuran with AChE was 7 min. The limits of detection for carbofuran were 6.3 nmol/L for the kinetic assay and 13.5 nmol/L for the endpoint assay. The paper demonstrates that the open alternative for commercial photometry is equivalent. The concept based on the OS3P/OS3P could be used as a large-scale screening system.

An immunosensor for the assay of toxic biological warfare agents is a biosensor suitable for detecting hazardous substances such as aflatoxin, botulinum toxin, ricin, Shiga toxin, and others. The application of immunosensors is used in outdoor assays, point-of-care tests, as a spare method for more expensive devices, and even in the laboratory as a standard analytical method. Some immunosensors, such as automated flow-through analyzers or lateral flow tests, have been successfully commercialized as tools for toxins assay, but the research is ongoing. New devices are being developed, and the use of advanced materials and assay techniques make immunosensors highly competitive analytical devices in the field of toxic biological warfare agents assay. This review summarizes facts about current applications and new trends of immunosensors regarding recent papers in this area.

The negative effects of the current agricultural practices include erosion, acidification, loss of soil organic matter (dehumification), loss of soil structure, soil contamination by risky elements, reduction of biological diversity and land use for non-agricultural purposes. All these effects are a huge risk to the further development of soil quality from an agronomic point of view and its resilience to projected climate change. Organic matter has a crucial role in it. Relatively significant correlations with the quality or the health of soil parameters and the soil organic matter or some fraction of the soil organic matter have been found. In particular, Ctot, Cox, humic and fulvic acids, the C/N ratio, and glomalin. Our work was focused on glomalin, a glycoprotein produced by the hyphae and spores of arbuscular mycorrhizal fungi (AMF), which we classify as Glomeromycota. Arbuscular mycorrhiza, and its molecular pathways, is not a well understood phenomenon. It appears that many proteins are involved in the arbuscular mycorrhiza from which glomalin is probably one of the most significant. This protein is also responsible for the unique chemical and physical properties of soils and has an ecological and economical relevance in this sense and it is a real product of the mycorrhiza. Glomalin is very resistant to destruction (recalcitrant) and difficult to dissolve in water. Its extraction requires specific conditions: high temperature (121°C) and a citrate buffer with a neutral or alkaline pH. Due to these properties, glomalin (or its fractions) are very stable compounds that protect the soil aggregate surface. In this review, the actual literature has been researched and the importance of glomalin is discussed.

Biosensors containing cholinesterase are analytical devices suitable for the assay of neurotoxic compounds. In the research on biosensors, a new platform has appeared some years ago. It is the digital photography and scoring of coloration (photogrammetry). In this paper, a colorimetric biosensor is constructed using 3D-printed multiwell pads treated with indoxylacetate as a chromogenic substrate and gold nanoparticles with the immobilized enzyme butyrylcholinesterase. A smartphone camera served for photogrammetry. The biosensor was tested for the assay of carbofuran and paraoxon ethyl as two types of covalently binding inhibitors: irreversible and pseudoirreversible. The biosensor exerted good sensitivity to the inhibitors and was able to detect carbofuran with a limit of detection for carbofuran 7.7 nmol/l and 17.6 nmol/l for paraoxon ethyl. A sample sized 25 μl was suitable for the assay lasting approximately 70 minutes. Up to 121 samples can be measured contemporary using one multiwell pad. The received data fully correlated with the standard spectrophotometry. The colorimetric biosensor exerts promising specifications and appears to be competitive to the other analytical procedures working on the principle of cholinesterase inhibition. Low-cost, simple, and portable design represent an advantage of the assay of the biosensor. Despite the overall simplicity, the biosensor can fully replace the standard spectroscopic methods.

Coronavirus disease 2019 (COVID-19) is a highly virulent infection that has caused a pandemic since 2019. Early diagnosis of the disease has been recognized as one of the important approaches to minimize the pathological impact and spread of infection. Point-of-care tests proved to be substantial analytical tools, and especially lateral flow immunoassays (lateral flow tests) serve the purpose. In the last few years, biosensors have gained popularity. These are simple but highly sensitive and accurate analytical devices composed from a selective molecule such as an antibody or antigen and a sensor platform. Biosensors would be an advanced alternative to current point-of-care tests for COVID-19 diagnosis and standard laboratory methods as well. Recent discoveries related to point-of-care diagnostic tests for COVID-19, the development of biosensors for specific antibodies and specific virus parts or their genetic information are reviewed.

Analytical devices for point-of-care diagnoses are highly desired and would improve quality of life when first diagnoses are made early and pathologies are recognized soon. Lateral flow tests (LFTs) are such tools that can be easily performed without specific equipment, skills, or experiences. This review is focused on the use of LFT in point-of-care diagnoses. The principle of the assay is explained, and new materials like nanoparticles for labeling, new recognition molecules for interaction with an analyte, and new additional instrumentation like signal scaling by a smartphone camera are described and discussed. Advantages of the LFT devices as well as their limitations are described and discussed here considering actual papers that are properly cited.

Background. Alzheimer’s disease (AD) is a multifactorial progressive and irreversible neurodegenerative disorder affecting mainly the population over 65 years of age. It is becoming a global health and socioeconomic problem, and the current number of patients reaching 30–50 million people will be three times higher over the next thirty years. Objective. Late diagnosis caused by decades of the asymptomatic phase and invasive and cost-demanding diagnosis are problems that make the whole situation worse. Electrochemical biosensors could be the right tool for less invasive and inexpensive early diagnosis helping to reduce spend sources— both money and time. Method. This review is a survey of the latest advances in the design of electrochemical biosensors for the early diagnosis of Alzheimer’s disease. Biosensors are divided according to target biomarkers. Conclusion. Standard laboratory methodology could be improved by analyzing a combination of currently estimated markers along with neurotransmitters and genetic markers from blood samples, which make the test for AD diagnosis available to the wide public.

C-reactive protein (CRP) is an important part of the immune system’s reaction to various pathological impulses such as bacterial infections, systemic inflammation, and internal organ failures. An increased CRP level serves to diagnose the mentioned pathological states. Both standard laboratory methods and simple point-of-care devices such as lateral flow tests and immunoturbidimetric assays serve for the instrumental diagnoses based on CRP. The current method for CRP has many flaws and limitations in its use. Biosensor and bioassay analytical devices are presently researched by many teams to provide more sensitive and better-suited tools for point-of-care tests of CRP in biological samples when compared to the standard methods. This review article is focused on mapping the diagnostical relevance of CRP, the applicability of the current analytical methods, and the recent innovations in the measurement of CRP level.

Glomalin is a soil protein resembling heat shock protein (HSP) 60 and exerting high affinity to metals, causing retention of water in the environment and improving mechanical stability of soil. Currently, glomalin is determined in the soil or other samples by combination of autoclaving extraction and total protein determination typically by the Bradford method. In this paper, a piezoelectric biosensor was prepared to determine glomalin in a label-free measurement. The biosensor contained antibodies immobilized on quartz crystal microbalance (QCM), and the recognition layer was stabilized by iron oxide nanoparticles. The assay was tested on real soil samples and compared with the standard Bradford assay. Limit of detection of the assay was equal to 2.4 µg/g for a soil extract with a volume of 50 µl. The assay takes approximately half of an hour and was fully correlated to the Bradford assay. The biosensor had significant advantages than the other methods: it worked in a label-free mode and was fully applicable for practical samples.

The onset of modern-style plate tectonics remains under debate. Here, Xu et al. report a cold thermal-gradient recorded in an eclogite xenolith in Paleoproterozoic carbonatite from orogen, and propose that modern-style subduction has operated since at least the Paleoproterozoic.

Soil samples (n = 11) were collected in the chernozem areas of the Czech Republic (the Central Europe) from the topsoil and used as representative samples. All sampling areas have been used for agricultural purposes (arable soil) and they were selected as typical representatives of agricultural soil. These samples represented the soil with same genesis (to reduction differencies between soil types) but with different soil properties (physical and chemical). Complete chemical and physical analyses were made for confirmation of copper adsorption on solid phase: we analysed the particle size distribution, content of oxidizable carbon (Cox), the cation exchange capacity (CEC), supply of exchange calcium, magnesium, sodium, phosphorus and potassium, soil reaction and the total supply of Fe, Al, Mn, Ca, Mg, K, P and N. The strongest simple correlation between analysed soil properties and copper concentration had content of available magnesium (r = 0.44) and available phosphorus (r = −0.51). In the case of multiple correlations (i. e. collective influence of multiple soil properties) had the strongest influence combination of clay, soil reaction, total content of phosphorus, available magnesium and available phosphorus. The main influence of phosphorus and magnesium is evident. We suppose that copper and phosphorus enter into specific complex. Influence of these five soil properties can explain 92.7 % (r = 0.927) changes in the content of copper changes in the experiment.

Lipases play a crucial role in metabolism of microbes, fungi, plants, and animals, and in analytical chemistry, they are often used in detection of fats and triglycerides. Determination of lipase activity is also important in toxicology, when lipase activity can be both increased and decreased by organophosphates and other pesticides and in medicine for diagnosis of heart diseases. The standard method for lipase activity determination is based on cleaving ester bonds in lipase buffer containing Tween. Our aim was to find a method with faster and more sensitive response. It is known that acetylcholinesterase belongs to the same group of hydrolases enzymes as lipases and it cleaves indoxyl acetate, so we assume indoxyl acetate could report a similar reaction with lipase. Our method is based on indoxyl acetate as a substrate for lipase, where indoxyl acetate is cleaved by lipase to indoxyl and acetate moiety and blue indigo is created. The method was optimized for different times and amount of enzyme and compared with the standard Tween assay. The calibration curve measured in reaction time 20 minutes with 10 μl of lipase exhibited the best analytical parameters, and it showed Michaelis–Menten response with the Michaelis–Menten constant equal to 8.72 mmol/l. The indoxyl acetate-based method showed faster and more sensitive response than the standard method for lipase activity determination, so it has great potential in biosensor construction and it could be used in industry, medicine, toxicology, and common practice where the activity of lipases is need to be measured.

Glycated hemoglobin (HbA1c) is a product of the spontaneous reaction between hemoglobin and elevated glucose levels in the blood. It is included among the so-called advanced glycation end products, of which is the most important for the clinical diagnosis of diabetes mellitus, and it can serve as an alternative to glycemia measurement. Compared to the diagnosis of diabetes mellitus by glycemia, the HbA1c level is less influenced by a short-term problem with diabetes compensation. Mass spectroscopy and chromatographic techniques are among the standard methods of HbA1c level measurement. Compared to glycemia measurement, there is lack of simple methods for diabetes mellitus diagnosis by means of the HbA1c assay using a point-of-care test. This review article is focused on the surveying of facts about HbA1c and its importance in diabetes mellitus diagnosis, and surveying standard methods and new methods suitable for the HbA1c assay under point-of-care conditions. Various bioassays and biosensors are mentioned and their specifications are discussed.

Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) can serve as biochemical markers of various pathologies like liver disfunction and poisonings by nerve agents. Ellman’s assay is the standard spectrophotometric method to measure cholinesterase activity in clinical laboratories. The authors present a new colorimetric test to assess AChE and BChE activity in biological samples using chromogenic reagents, treated 3D-printed measuring pads and a smartphone camera as a signal detector. Multiwell pads treated with reagent substrates 2,6-dichlorophenolindophenyl acetate, indoxylacetate, ethoxyresorufin and methoxyresorufin were prepared and tested for AChE and BChE. In the experiments, 3D-printed pads containing indoxylacetate as a chromogenic substrate were optimal for analytical purposes. The best results were achieved using the red (R) channel, where the limit of detection was 4.05 µkat/mL for BChE and 4.38 µkat/mL for AChE using a 40 µL sample and a 60 min assay. The major advantage of this method is its overall simplicity, as samples are applied directly without any specific treatment or added reagents. The assay was also validated to the standard Ellman’s assay using human plasma samples. In conclusion, this smartphone camera-based colorimetric assay appears to have practical applicability and to be a suitable method for point-of-care testing because it does not require specific manipulation, additional education of staff or use of sophisticated analytical instruments.

In animal models, there was observed alteration of various physiological processes caused by microtraumas. Here reported experiment was aimed on the research of link between injection and development of an oxidative imbalance. Laboratory guinea pig was chosen as a suitable model for examining of the oxidative stress. Markers indicating oxidative homeostasis were assayed in the frontal, temporal and occipital brain lobe, cerebellum, liver, kidney, spleen and heart one hour after an intramuscular injection. Common biochemical parameters were measured in plasma samples as well. The most extensive effect was observed in the heart where the thiobarbituric acid reactive substances value was more than twice increased after the injection. The level of carbonylated proteins was significantly elevated in the kidney and ferric reducing antioxidant power value was increased in the brain compartments. The enzyme activities in the organs were not influenced except the activity of superoxide dismutase, which was moderately decreased in the brain. In the plasma samples, there was observed increase of the blood urea nitrogen. The results showed significant the influence of the intramuscular injection on a development of an oxidative insult. The injection can be considered as an adverse effect with quite extensive stress consequences.

Background: Alcohol variants such as ethanol and methanol are simple organic compounds widely used in foods, pharmaceuticals, chemical synthesis, etc. Both are becoming an emerging health problem; abuse of ethanol containing beverages can lead to disparate health problems and methanol is highly toxic and unfit for consumption. Methods and Results: This review summarizes the basic knowledge about ethanol and methanol toxicity, the effect mechanism on the body, the current care of poisoned individuals and the implication of alcohols in the development of diseases. Alcohol related dementia, stroke, metabolic syndrome and hepatitis are discussed as well. Besides ethanol, methanol toxicity and its biodegradation pathways are addressed. Conclusions: The impact of ethanol and methanol on the body is shown as case reports, along with a discussion on the possible implication of alcohol in Alzheimer's disease and antidotal therapy for methanol poisoning. The role of ethanol in cancer and degenerative disorders seems to be underestimated given the current knowledge. Treatment in case of poisoning is another issue that remains unresolved even though effective protocols and drugs exist.

An old saying states that ‘’children are not little adults” and this certainly holds true for celiac disease, as there are many peculiar aspects regarding its epidemiology, diagnosis, clinical presentations, associated diseases, and response to treatment in pediatric compared to adult populations, to such an extent that it merits a description of its own. In fact, contrary to the past when it was thought that celiac disease was a disorder predominantly affecting childhood and characterized by a malabsorption syndrome, nowadays it is well recognized that it affects also adult and elderly people with an impressive variability of clinical presentation. In general, the clinical guidelines for diagnosis recommend starting with specific serologic testing in all suspected subjects, including those suffering from extraintestinal related conditions, and performing upper endoscopy with appropriate biopsy sampling of duodenal mucosa in case of positivity. The latter may be omitted in young patients showing high titers of anti-transglutaminase antibodies. The subsequent management of a celiac patient differs substantially depending on the age at diagnosis and should be based on the important consideration that this is a lifelong condition.

Acetylcholinesterase is an enzyme responsible for termination of excitatory transmission at cholinergic synapses by the hydrolyzing of a neurotransmitter acetylcholine. Nowadays, other functions of acetylcholinesterase in the organism are considered, for example its role in regulation of apoptosis. Cholinergic nervous system as well as acetylcholinesterase activity is closely related to pathogenesis of Alzheimer disease. The mostly used therapy of Alzheimer disease is based on enhancing cholinergic function using inhibitors of acetylcholinesterase like rivastigmine, donepezil or galantamine. These drugs can influence not only the acetylcholinesterase activity but also other processes in treated organism. The paper is aimed mainly on possibility of increased expression and protein level of acetylcholinesterase caused by the therapy with acetylcholinesterase inhibitors.

Melatonin is a hormone with strong antioxidant properties. In this experiment, Freund’s complete adjuvant was used as a stressogenic substance given to laboratory outbred mice, whereas melatonin was investigated as a protectant against the stressogenic effect. Levels of low molecular weight antioxidants, thiobarbituric acid reactive substances, and tumor necrosis factor α and activity of glutathione reductase were determined in blood from the animals. Surprisingly, melatonin was not involved in direct regulation of antioxidants, thiobarbituric acid reactive substances and tumor necrosis factor α. On the other hand, melatonin regulated glutathione reductase activity. We can conclude on regulation of metabolism caused by melatonin in the model. The effect was more important than the expected regulation of immunity and basal oxidative homeostasis.

Smartphones are popular devices frequently equipped with sensitive sensors and great computational ability. Despite the widespread availability of smartphones, practical uses in analytical chemistry are limited, though some papers have proposed promising applications. In the present paper, a smartphone is used as a tool for the determination of cholinesterasemia i.e., the determination of a biochemical marker butyrylcholinesterase (BChE). The work should demonstrate suitability of a smartphone-integrated camera for analytical purposes. Paper strips soaked with indoxylacetate were used for the determination of BChE activity, while the standard Ellman’s assay was used as a reference measurement. In the smartphone-based assay, BChE converted indoxylacetate to indigo blue and coloration was photographed using the phone’s integrated camera. A RGB color model was analyzed and color values for the individual color channels were determined. The assay was verified using plasma samples and samples containing pure BChE, and validated using Ellmans’s assay. The smartphone assay was proved to be reliable and applicable for routine diagnoses where BChE serves as a marker (liver function tests; some poisonings, etc.). It can be concluded that the assay is expected to be of practical applicability because of the results’ relevance.

Smartphones are widely spread and their usage does not require any trained personnel. Recently, smartphones were successfully used in analytical chemistry as a simple detection tool in some applications. This paper focuses on immobilization of acetylcholinesterase (AChE) onto commercially available pH strips with stabilization in the gelatin membrane. AChE degrades acetylcholine into choline and acetic acid which causes color change of acid-base indicator. Smartphone served as a tool for measurement of indicator color change from red to orange while inhibitors blocked this process. AChE inhibitors were measured with limits of detection, 149 nM and 22.3 nM for galanthamine and donepezil, respectively. Organic solvents were measured for method interferences. Measurement procedure was performed on 3D printed holder and digital photography was evaluated using red-green-blue (RGB) channels. The invented assay was validated to the standard Ellman’s test and verified on murine plasma samples spiked with inhibitors. We consider that the assay is fully suitable for practical performance.

Acetylcholinesterase (AChE) inhibitors are widely used for the symptomatic treatment of Alzheimer’s disease and other dementias. More recent use is for myasthenia gravis. Many of these inhibitors interact with the second known cholinesterase, butyrylcholinesterase (BChE). Further, evidence shows that acetylcholine plays a role in suppression of cytokine release through a “cholinergic anti-inflammatory pathway” which raises questions about the role of these inhibitors in the immune system. This review covers research and discussion of the role of the inhibitors in modulating the immune response using as examples the commonly available drugs, donepezil, galantamine, huperzine, neostigmine and pyridostigmine. Major attention is given to the cholinergic anti-inflammatory pathway, a well-described link between the central nervous system and terminal effector cells in the immune system.

The aim of this study was to evaluate the bioactive substances in 19 berry cultivars of edible honeysuckle (Lonicera edulis). A statistical evaluation was used to determine the relationship between the content of selected bioactive substances and individual cultivars. Regarding mineral elements, the content of sodium was measured using potentiometry and spectrophotometry. The content of selected polyphenolic compounds with high antioxidant activity was determined by a HPLC–UV/ED method. The total amount of polyphenols was determined by the Folin-Ciocalteu method. The antioxidant activity was determined using five methods (DPPH, FRAP, ABTS, FR and DMPD) that differ in their principles. The content of 13 amino acids was determined by ion-exchange chromatography. The experimental results obtained for the different cultivars were evaluated and compared by statistical and bioinformatic methods. A unique feature of this study lies in the exhaustive analysis of the chosen parameters (amino acids, mineral elements, polyphenolic compounds and antioxidant activity) during one growing season.

Biosensors are analytical devices combining a physical sensor with a part of biological origin providing sensitivity and selectivity toward analyte. Biological warfare agents are infectious microorganisms or toxins with the capability to harm or kill humans. They can be produced and spread by a military or misused by a terrorist group. For example, Bacillus anthracis, Francisella tularensis, Brucella sp., Yersinia pestis, staphylococcal enterotoxin B, botulinum toxin and orthopoxviruses are typical biological warfare agents. Biosensors for biological warfare agents serve as simple but reliable analytical tools for the both field and laboratory assay. There are examples of commercially available biosensors, but research and development of new types continue and their application in praxis can be expected in the future. This review summarizes the facts and role of biosensors in the biological warfare agents’ assay, and shows current commercially available devices and trends in research of the news. Survey of actual literature is provided.

Lipases are enzymes responsible for the conversion of triglycerides and other esterified substrates, they are involved in the basic metabolism of a wide number of organisms, from a simple microorganism and to mammals. They also have broad applicability in many fields from which industrial biotechnology, the production of cleaning agents, and pharmacy are the most important. The use of lipases in analytical chemistry where it can serve as a part of biosensors or bioassays is an application of growing interest and has become another important use. This review is focused on the description of lipases chemistry, their current applications and the methods for their assay measurement. Examples of bioassays and biosensors, including their physical and chemical principles, performance for specific substrates, and discussion of their relevance, are given in this work.

Flavonoids are a significant group of secondary metabolites in plants. Many of these compounds are potent antioxidants, being an important part in food products derived from the plants. The current status of research on flavonoid compounds in the fruit of Saskatoon berries (Amelanchier alnifolia Nutt.) and their health promoting effects, including recommended utilization, are reviewed. The major classes of flavonoids in the fruit are flavonols (quercetin and rutin), flavanes (proanthocyanidin compounds ranging from dimers through to heptamers and even higher polymers) and finally anthocyanins. The flavonoids represented the group of polyphenols that mostly contributed to the antioxidant activity of Saskatoon berries. High content of the flavoinoids antioxidants in the fruit is responsible for the observed anti-inflammatory, antidiadiabetic and chemo-protective effects.

The enzyme acetylcholinesterase (AChE) is an important part of cholinergic nervous system, where it stops neurotransmission by hydrolysis of the neurotransmitter acetylcholine. It is sensitive to inhibition by organophosphate and carbamate insecticides, some Alzheimer disease drugs, secondary metabolites such as aflatoxins and nerve agents used in chemical warfare. When immobilized on a sensor (physico-chemical transducer), it can be used for assay of these inhibitors. In the experiments described herein, an AChE- based electrochemical biosensor using screen printed electrode systems was prepared. The biosensor was used for assay of nerve agents such as sarin, soman, tabun and VX. The limits of detection achieved in a measuring protocol lasting ten minutes were 7.41 × 10−12 mol/L for sarin, 6.31 × 10−12 mol /L for soman, 6.17 × 10−11 mol/L for tabun, and 2.19 × 10−11 mol/L for VX, respectively. The assay was reliable, with minor interferences caused by the organic solvents ethanol, methanol, isopropanol and acetonitrile. Isopropanol was chosen as suitable medium for processing lipophilic samples.

Caffeine is an alkaloid with a stimulant effect in the body. It can interfere in transmissions based on acetylcholine, epinephrine, norepinephrine, serotonin, dopamine and glutamate. Clinical studies indicate that it can be involved in the slowing of Alzheimer disease pathology and some other effects. The effects are not well understood. In the present work, we focused on the question whether caffeine can inhibit acetylcholinesterase (AChE) and/or, butyrylcholinesterase (BChE), the two enzymes participating in cholinergic neurotransmission. A standard Ellman test with human AChE and BChE was done for altering concentrations of caffeine. The test was supported by an in silico examination as well. Donepezil and tacrine were used as standards. In compliance with Dixon’s plot, caffeine was proved to be a non-competitive inhibitor of AChE and BChE. However, inhibition of BChE was quite weak, as the inhibition constant, Ki, was 13.9 ± 7.4 mol/L. Inhibition of AChE was more relevant, as Ki was found to be 175 ± 9 µmol/L. The predicted free energy of binding was −6.7 kcal/mol. The proposed binding orientation of caffeine can interact with Trp86, and it can be stabilize by Tyr337 in comparison to the smaller Ala328 in the case of human BChE; thus, it can explain the lower binding affinity of caffeine for BChE with reference to AChE. The biological relevance of the findings is discussed.

Alpha7 nicotinic acetylcholine receptor (α7 nAChR) is an important part of the cholinergic nerve system in the brain. Moreover, it is associated with a cholinergic anti-inflammatory pathway in the termination of the parasympathetic nervous system. Antagonists of α7 nAChR are a wide group represented by conotoxin and bungarotoxin. Even Alzheimer’s disease drug memantine acting as an antagonist in its side pathway belongs in this group. Agonists of α7 nAChR are suitable for treatment of multiple cognitive dysfunctions such as Alzheimer’s disease or schizophrenia. Inflammation or even sepsis can be ameliorated by the agonistic acting compounds. Preparations RG3487, SEN34625/WYE-103914, SEN12333, ABT-107, Clozapine, GTS-21, CNI-1493, and AR-R17779 are representative examples of the novel compounds with affinity toward the α7 nAChR. Pharmacological, toxicological, and medicinal significance of α7 nAChR are discussed throughout this paper.

Assay of acetylcholinesterase (AChE) activity plays an important role in diagnostic, detection of pesticides and nerve agents, in vitro characterization of toxins and drugs including potential treatments for Alzheimer’s disease. These experiments were done in order to determine whether indoxylacetate could be an adequate chromogenic reactant for AChE assay evaluation. Moreover, the results were compared to the standard Ellman’s method. We calculated Michaelis constant Km (2.06 × 10−4 mol/L for acetylthiocholine and 3.21 × 10−3 mol/L for indoxylacetate) maximum reaction velocity Vmax (4.97 × 10−7 kat for acetylcholine and 7.71 × 10−8 kat for indoxylacetate) for electric eel AChE. In a second part, inhibition values were plotted for paraoxon, and reactivation efficacy was measured for some standard oxime reactivators: obidoxime, pralidoxime (2-PAM) and HI-6. Though indoxylacetate is split with lower turnover rate, this compound appears as a very attractive reactant since it does not show any chemical reactivity with oxime antidots and thiol used for the Ellman’s method. Thus it can be advantageously used for accurate measurement of AChE activity. Suitability of assay for butyrylcholinesterase activity assessment is also discussed.

We have evaluated in vitro the potency of 23 oximes to reactivate human erythrocyte acetylcholinesterase (AChE) and plasma butyrylcholinesterase (BChE) inhibited by racemic leptophos-oxon (O-[4-bromo-2,5-dichlorophenyl]-O-methyl phenyl-phosphonate), a toxic metabolite of the pesticide leptophos. Compounds were assayed in concentrations of 10 and 100 µM. In case of leptophos-oxon inhibited AChE, the best reactivation potency was achieved with methoxime, trimedoxime, obidoxime and oxime K027. The most potent reactivators of inhibited BChE were K033, obidoxime, K117, bis-3-PA, K075, K074 and K127. The reactivation efficacy of tested oximes was lower in case of leptophos-oxon inhibited BChE.

Piezoelectric biosensors are a group of analytical devices working on a principle of affinity interaction recording. A piezoelectric platform or piezoelectric crystal is a sensor part working on the principle of oscillations change due to a mass bound on the piezoelectric crystal surface. In this review, biosensors having their surface modified with an antibody or antigen, with a molecularly imprinted polymer, with genetic information like single stranded DNA, and biosensors with bound receptors of organic of biochemical origin, are presented and discussed. The mentioned recognition parts are frequently combined with use of nanoparticles and applications in this way are also introduced. An overview of the current literature is given and the methods presented are commented upon.

Quaternary pyridinium salts with chains ranging from C8 to C20 belong in the large group of cationic surfactants. In this paper, the preparation of such cationic surface active agents based on the pyridinium moiety and differing in the length of the N-alkyl chain is described. Additionally, HPLC technique was established to distinguish each prepared pyridinium analogue. This study represents universal method for preparation and identification of quaternary pyridinium detergents.

Four novel bisquaternary aldoxime cholinesterase reactivators differing in their chemical structure were prepared. Afterwards, their biological activity was evaluated for their ability to reactivate acetylcholinesterase (AChE; EC 3.1.1.7) and butyrylcholinesterase (BuChE; EC 3.1.1.8) inhibited by paraoxon. Their reactivation activity was compared with standard reactivators—pralidoxime, obidoxime and HI-6—which are clinically used at present. As it resulted, none of the prepared compounds surpassed obidoxime, which is considered to be the most potent compound if used for reactivation of AChE inhibited by paraoxon. In case of BuChE reactivation, two compounds (K053 and K068) achieved similar results as obidoxime.

The purpose of the present study was to employ two methods—square wave voltammetry (SWV) performed on screen printed sensors and ferric reducing antioxidant power (FRAP)—as suitable tools for the assay of low-molecular-weight antioxidants (LMWAs). LMWAs were assayed by both methods and the resulting data were statistically compared. Plasma samples from five Cinereous vultures accidentally intoxicated with lead were used to represent real biological matrices with different levels of LMWAs. Blood was collected from the birds prior to and one month after treatment with Ca-EDTA. SWV resulted in two peaks. The first peak, with the potential value of 466 ± 15 mV, was recognized as ascorbic and uric acids, while the second one (743 ± 30 mV) represented glutathione, tocopherol, ascorbic acid and in a minor effect by uric acid, too. Contribution of individual antioxidants was recognized by separate assays of LMWA standards. Correlation between peaks 1 and 2 as well as the sum of the two peaks and FRAP was analysed. While peak 1 and the sum of peaks were in close correlation to FRAP results (correlation coefficient of 0.97), the relation between peak 2 and FRAP may be expressed using a correlation coefficient of 0.64. The determination of thiols by the Ellman assay confirmed the accuracy of SWV. Levels of glutathione and other similar structures were stable in the chosen model and it may be concluded that SWV is appropriate for assay of LMWAs in plasma samples. The methods employed in the study were advantageous in minimal sample volume consumption and fast acquisition of results.

Seven new oxime-based acetylcholinesterase reactivators were compared with three currently available ones (obidoxime, trimedoxime, HI-6) for their ability to lessen cholinesterase inhibition in blood and brain of cyclosarin-treated rats. Oximes were given at doses of 5% their LD50 along with 21 mg/kg atropine five min before the LD50 of cyclosarin (120 ug/kg) was administered. Blood and brain samples were collected 30 minutes later. The greatest difference between acetylcholinesterase inhibition in blood of cyclosarin-treated rats was found after administration of HI-6 (40%), compared to 22% for trimedoxime and 6% for obidoxime. Only two of the seven newly synthesized oximes had any effect (K203 at 7%, K156 at 5%). Effective oximes against cyclosarin-inhibited plasma butyrylcholinesterase were HI-6 (42%), trimedoxime (11%), and K156 (4%). The oximes were less effective in brain than in blood, with reactivation values for HI-6 30% against acetylcholinesterase and 10% against butyrylcholinesterase. Values for newly synthesized oximes were less than 10% for K206, K269 and K203.

A cholinesterase based biosensor was constructed in order to assess the effects of ionizing radiation on exposed AChE. Although the primary objective of the experiment was to investigate the effect of ionizing radiation on the activity of the biosensor, no changes in cholinesterase activity were observed. Current provided by oxidation of thiocholine previously created from acetylthiocholine by enzyme catalyzed reaction was in a range 395–455 nA. No significant influence of radiation on AChE activity was found, despite the current variation. However, a surprising phenomenon was observed when a model organophosphate paraoxon was assayed. Irradiated biosensors seem to be more susceptible to the inhibitory effects of paraoxon. Control biosensors provided a 94 ± 5 nA current after exposure to 1 ppm paraoxon. The biosensors irradiated by a 5 kGy radiation dose and exposed to paraoxon provided a current of 49 ± 6 nA. Irradiation by doses ranging from 5 mGy to 100 kGy were investigated and the mentioned effect was confirmed at doses above 50 Gy. After the first promising experiments, biosensors irradiated by 5 kGy were used for calibration on paraoxon and compared with the control biosensors. Limits of detection 2.5 and 3.8 ppb were achieved for irradiated and non-irradiated biosensors respectively. The overall impact of this effect is discussed.

Cholinesterase activity in blood of laboratory rats was monitored. Rats were intoxicated with paraoxon at dosis of 0 – 65 – 125 – 170 – 250 – 500 nmol. The 250 nmol dose was found to be the LD50. An electrochemical sensor was found useful to provide information about cholinesterase activity. The decrease of cholinesterase activity was correlated to intoxication symptoms and mortality level. It was found that the symptoms of intoxication are not observed while at least 50% of cholinesterase activity in blood remains. The minimal cholinesterase activity essential to survival is around 10%, when compared with the initial state. No changes in levels of low moleculary weight antioxidants were observed.

The therapeutical efficacies of eleven oxime-based acetylcholinesterase reactivators were compared in an in vivo (rat model) study of treatment of intoxication caused by tabun. In this group there were some currently available oximes (obidoxime, trimedoxime and HI-6) and the rest were newly synthesized compounds. The best reactivation efficacy for acetylcholinesterase in blood (expressed as percent of reactivation) among the currently available oximes was observed after administration of trimedoxime (16%) and of the newly synthesized K127 (22432) (25%). The reactivation of butyrylcholinesterase in plasma was also studied; the best reactivators were trimedoxime, K117 (22435), and K127 (22432), with overall reactivation efficacies of approximately 30%. Partial protection of brain ChE against tabun inhibition was observed after administration of trimedoxime (acetylcholinesterase 20%; butyrylcholinesterase 30%) and obidoxime (acetylcholinesterase 12%; butyrylcholinesterase 16%).

An electrochemical sensor is introduced as a tool applicable for diagnosis of intoxication by cholinesterase inhibitors caused by the well-known nerve agent VX. The traditional Ellman method was chosen for comparison with the sensor's analytical parameters. Both methods are based on estimation of blood cholinesterase inhibition as a marker of intoxication. While Ellman´s method provided a limit of detection of 5.2´10-7 M for blood containing VX, the electrochemical sensor was able to detect 4.0´10-7 M. Good correlation between both methods was observed (R = 0.92). The electrochemical sensor could be considered a convenient tool for a fast yet accurate method, easily available for field as well as laboratory use. Time and cost savings are key features of the sensor-based assay.

An amperometric biosensor based on acetylcholinesterase (AChE) immobilized in gelatin was used to develop an assay for the organophosphate paraoxon. The more traditional manner employing preincubation was used for comparison between measurement procedures, although the aim of the study was to examine the performance of the biosensor for real time monitoring of organophosphates. The biosensor was immersed in a reaction chamber and paraoxon was injected inside. We were able to detect 200 pg of paraoxon within one minute or 2.5 ppb when the biosensor was preincubed in the sample solution for 15 minutes. The practical impact and expectations are discussed.

A carbon paste electrode biosensor for D-lactic acid analysis in biological samples was developed. D-lactate cytochrome c oxidoreductase from baker’s yeast (EC 1.1.2.4) was used as biorecognition element of the electrode in combination with phenazine methosulphate (PMS)–Reinecke’s salt precipitate as mediator. The enzyme was bound on paste due to Enzacryl using glutardialdehyde. The electrode response to D-lactate addition was linear in the concentration range of 0.015–100 mM. Limit of detection was equal to 56 μM when signal vs. noise criterium (S/N=3) is considered. Specificity for only D-lactate was proven. Relative standard deviation of 6.6 % for 12 mM D-lactate was found. The biosensor exerts long-term stability and also good stability during repeated measuring cycles. The electrode has a good potential for use in practical samples.

Piezoelectric biosensor was used for diagnosis of infection by Francisellatularensis subsp. holarctica in European brown hares. Two kinds of experiments wereperformed in this study. First, sera from experimentally infected European brown hares(Lepus europaeus) were assayed by piezoelectric biosensor and the seventh day postinfection was found as the first one when statistically significant diagnosis of tularemia waspossible; all other sera collected from hares later than on day 7 following the infection werefound tularemia positive. Typing to classify the field strain of F. tularensis used for theexperimental infection was confirmed by proteome study. Second, sera from 35 Europeanbrown hare specimens sampled at hunting grounds and tested as tularemia positive by slowagglutination allowed diagnosis of tularemia by the piezoelectric biosensor. All these sera ofnaturally infected hares were found as tularemia positive, too. Efficacy of the piezoelectricbiosensor for the serological diagnosis of tularemia is discussed.

Monoclonal antibodies with specificity against the Francisella tularensis outerlipopolysaccharide (LPS) membrane were prepared and characterized using the piezoelectricimmunosensor with immobilized LPS antigen from F. tularensis. Signals obtained by theimmunosensor were compared with ELISA and similar sensitivity was noticed. Signal ofnegative controls obtained using the biosensor was below 0.5% of the signal obtained for theselected specific antibody clone 4H3B9D3. Testing of cross reactivity based on the sensorswith immobilized LPS from Escherichia coli and Bacillus subtilis confirmed selectivity ofthis antibody. Furthermore, the 4H3B9D3 antibody was successfully isotypized as IgM usingthe piezoelectric sensors with secondary antibodies. Kinetics parameters of antibody wereevaluated in the flow-through arrangement. The kinetic rate constants for the antibody4H3B9D3 were ka = (2.31 ± 0.20)·105 l mol-1s-1 (association) and kd = (0.0010 ±0.00062) s-1(dissociation) indicating very good affinity to the LPS antigen.

Amperometric biosensor based on enzyme acetylcholinesterase (AChE; EC 3.1.1.7) was tested for pesticide methamidophos assay. Biosensor consists from four screen printed platinum electrodes on ceramic strip. AChE was physically adsorbed onto the electrode surface. The measuring principle was based on the inhibition of AChE activity in the presence of methamidophos. The proposed method limit of detection was 2.45 nM, responding to 3.46 pg of methamidophos detected absolutely when we consider the sample volume.

The use of a cell phone as a detection system is easy, simple and does not require trained personnel, which is in contrast to standard laboratory instruments. This paper deals with immobilization of acetylcholinesterase (AChE) in a gelatin matrix, and phenol red, as an indicator of AChE activity, is used in order to establish a method that is easily compatible with a camera device. AChE splits acetylcholine into choline and acetic acid, which changes the pH of a medium, resulting in a phenol red color change. The coloration changed in presence of an AChE inhibitor. Measurements were performed on 3D-printed, tube-shaped holder, and digital photography, with subsequent analysis of red-green-blue (RGB), served for assay purposes. Calibration of AChE inhibitors, tacrine and galantamine, was performed, with limit of detection equal to 1.1 nM and 1.28 µM, respectively. Interferences were also measured, resulting in a proof-of-method stability. The method was further successfully validated for the standard Ellman’s assay, and verified on murine plasma samples spiked with inhibitors.