Total Chemiluminescence Research Articles

Features of the Kinetics of the Fenton Reaction

The Fenton reaction is of considerable interest due to the fact that it can be applied in chemical, biochemical and biomedical research. The Fenton reaction makes it possible to estimate the content of hydroperoxides. Hydroperoxides in a large number of cases are the trace that remains after the occurrence of redox reactions. To perform such studies, it is necessary to know the characteristics of the reaction at different ratios between the concentrations of hydroperoxides and ferrous iron. The reaction yield is determined by registering chemiluminescence. The number of oxidation events of ferrous iron N(Fe2+ ® Fe3+) in the Fenton reaction is calculated in this work for the cases [Fe2+] > [H2 O2 ] and [Fe2+] < [H2 O2 ] at concentrations of Fe2+ and H2 O2 ranging from 10-3 to 10-7 mol/L. It is shown that the peroxide concentration can be unambiguously determined only for the case [Fe2+] > [H2 O2 ]. In the case of [Fe2+] < [H2 O2 ], due to a strong decrease in the reaction rate, it is not always possible to register the total chemiluminescence yield. The kinetics of the formation of luminous products in the Fenton reaction after the introduction of luminol has been calculated. Conditions are considered when chemiluminescence with luminol is proportional to the concentration of hydroperoxides. The considered features of the Fenton reaction must be taken into account when performing specific research work.

Enhanced Photon Emission of Chemiluminescent Luminophore for Ultra-Fast and Semi-Automatic Immunoassay toward Single Molecule Detection.

Optical single molecule detection is normally achieved via amplifying the total emission of photons of luminophores and is strongly anticipated to extend the commercialized application of chemiluminescence (CL). To overcome the limited CL photons of molecule luminophores, herein, a nanocrystal (NC) luminophore self-amplified strategy is proposed to repetitively excite CL luminophores for amplifying the total CL photons per luminophore, which can be exploited to perform CL immunoassays (CLIAs) toward single molecule detection via employing KMnO4 as the CL triggering agent and the dual-stabilizer-capped CdTe NCs as the CL luminophore. KMnO4 can oxidize the S element from each stabilizer of mercaptopropionic acid (MPA) and release enough energy to excite the CdTe core for flash CL. The substantial MPA around each CdTe core enables every CdTe luminophore to be repetitively excited and give off amplified total CL photons in a self-enhanced way. The CL of CdTe NCs/KMnO4 can release all photons rapidly, and the collection of all these photons can be utilized to determine the model analyte of thyroid-stimulating hormone antigen (TSH) with a limit of detection of 5 ag/mL (S/N = 3), which is corresponding to about 2-4 TSH molecules in a 20 μL sample. The whole immunologic operating process can be terminated within 6 min. This strategy of repetitively breaking the CL reaction involving chemical bonds within one luminophore is promising for semi-automatic as well as fully automatic single molecule detection and extends the commercialized application of CL immunodiagnosis.

Vitamin D in COVID-19

Severe acute respiratory syndrome coronavirus 2(SARS-CoV-2) causes coronavirus disease 2019 (COVID-19) with clinical outcome ranging from asymptomatic to severe disease, and even death to some. It posed a terrifying challenge to healthcare system worldwide. Several observational and clinical trials has reported that, Vitamin D deficiency has contributed to acute respiratory distress syndrome. Case fatality rate increases with age and comorbidities, both of which are associated with decreased Vitamin D level. Therefore, this study is done to study the prevalence of 25(OH)Vitamin D in RT-PCR positive COVID-19 cases and RT-PCR negative controls.This is a hospital based cross-sectional study conducted at Jawaharlal Nehru Institute of Medical Sciences, Manipur on 88 RT-PCR positive Covid-19 cases and 88 COVID-19 negative controls over a period of 2 years. Analysis of the sample was done by Liaison 25(OH) Vitamin D Total Chemiluminescence assay(CLIA).In this study, statistically significant (p-0.018) lower plasma 25(OH)Vitamin D level is seen in COVID-19 positive cases (median 28±20.47) when compared to Covid-19 negative controls(median 33.50±10.66). The number of 25(OH) Vitamin D deficient is higher in Covid-19 positive cases when compared to Covid-19 negative controls -46(52.3%) and 30(34%) respectively with a high statistically significant value (p-0.015).COVID-19 positive cases have higher tendency to have suboptimal plasma 25(OH) Vitamin D level which may contribute to the high hospitalization risk in COVID-19 infection. This finding is important as it can identify population at risk, and contribute to interventions in reducing the risk of hospitalization associated with COVID-19 infection.

Application of high-speed, species-specific chemiluminescence imaging for laminar flame speed and Markstein length measurements in spherically expanding flames

A high-speed chemiluminescence imaging diagnostic system has been developed and characterized for measuring species-specific properties of spherically expanding flames, with the objective of obtaining synchronized measurements of multiple flame properties from a single flame experiment. The imaging system comprises a high-speed camera coupled to a high-speed image intensifier and optical filters to isolate emission from desired excited species. This study validates the diagnostic scheme specifically for measuring laminar flame speed and Markstein length from chemiluminescence at three wavelength bands corresponding to OH*, CH*, and total chemiluminescence, respectively. Mixtures of methane-air and methane/ethane-air were tested at initial conditions of atmospheric pressure and room temperature. An under-utilized methodology for computing unburned Markstein length from burned Markstein length was implemented. Flame speed modeling of each mixture was conducted in Chemkin using AramcoMech 2.0. No significant differences were observed between the results derived from the three emission wavelength bands for the flame speeds nor the Markstein lengths. For both of these properties, excellent agreement was observed between the experimental results and the values reported in the literature. Because laminar flame speed and Markstein length show no dependency on the imaging wavelength, the present results imply that multiple flame properties, such as laminar flame speed, Markstein length, flame thickness, and excited-species intensity profiles, can all be acquired simultaneously for the same flame using this method.

Distinct SARS-CoV-2 antibody reactivity patterns in coronavirus convalescent plasma revealed by a coronavirus antigen microarray

A coronavirus antigen microarray (COVAM) was constructed containing 11 SARS-CoV-2, 5 SARS-1, 5 MERS, and 12 seasonal coronavirus recombinant proteins. The array is designed to measure immunoglobulin isotype and subtype levels in serum or plasma samples against each of the individual antigens printed on the array. We probed the COVAM with COVID-19 convalescent plasma (CCP) collected from 99 donors who recovered from a PCR+ confirmed SARS-CoV-2 infection. The results were analyzed using two computational approaches, a generalized linear model (glm) and random forest (RF) prediction model, to classify individual specimens as either Reactive or non-reactive against the SARS-CoV-2 antigens. A training set of 88 pre-COVID-19 specimens (PreCoV) collected in August 2019 and102 positive specimens from SARS-CoV-2 PCR+ confirmed COVID-19 cases was used for these analyses. Results compared with an FDA emergency use authorized (EUA) SARS-CoV2 S1-based total Ig chemiluminescence immunoassay (Ortho Clinical Diagnostics VITROS Anti-SARS-CoV-2 Total, CoV2T) and with a SARS-CoV-2 S1-S2 spike-based pseudovirus micro neutralization assay (SARS-CoV-2 reporter viral particle neutralization titration (RVPNT) showed high concordance between the three assays. Three CCP specimens that were negative by the VITROS CoV2T immunoassay were also negative by both COVAM and the RVPNT assay. Concordance between VITROS CoV2T and COVAM was 96%, VITROS CoV2T and RVPNT 93%, and RVPNT and COVAM 91%. The discordances were all weakly reactive samples near the cutoff threshold of the VITROS CoV2T immunoassay. The multiplex COVAM allows CCP to be grouped according to antibody reactivity patterns against 11 SARS-CoV-2 antigens. Unsupervised K-means analysis, via the gap statistics, as well as hierarchical clustering analysis revealed three main clusters with distinct reactivity intensities and patterns. These patterns were not recapitulated by adjusting the VITROS CoV2T or RVPNT assay thresholds. Plasma classified by COVAM reactivity patterns offers potential to improve CCP therapeutic efficacy CoV2T alone. The use of a SARS-CoV-2 antigen array can qualify CCP for administration as a treatment for acute COVID-19, and interrogate vaccine immunogenicity and performance in preclinical, clinical studies, and routine vaccination to identify antibody responses predictive of protection from infection and disease.

Prevalence of SARS-CoV-2 antibodies in a large nationwide sample of patients on dialysis in the USA: a cross-sectional study

BackgroundMany patients receiving dialysis in the USA share the socioeconomic characteristics of underserved communities, and undergo routine monthly laboratory testing, facilitating a practical, unbiased, and repeatable assessment of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) seroprevalence.MethodsFor this cross-sectional study, in partnership with a central laboratory that receives samples from approximately 1300 dialysis facilities across the USA, we tested the remainder plasma of 28 503 randomly selected adult patients receiving dialysis in July, 2020, using a spike protein receptor binding domain total antibody chemiluminescence assay (100% sensitivity, 99·8% specificity). We extracted data on age, sex, race and ethnicity, and residence and facility ZIP codes from the anonymised electronic health records, linking patient-level residence data with cumulative and daily cases and deaths per 100 000 population and with nasal swab test positivity rates. We standardised prevalence estimates according to the overall US dialysis and adult population, and present estimates for four prespecified strata (age, sex, region, and race and ethnicity).FindingsThe sampled population had similar age, sex, and race and ethnicity distribution to the US dialysis population, with a higher proportion of older people, men, and people living in majority Black and Hispanic neighbourhoods than in the US adult population. Seroprevalence of SARS-CoV-2 was 8·0% (95% CI 7·7–8·4) in the sample, 8·3% (8·0–8·6) when standardised to the US dialysis population, and 9·3% (8·8–9·9) when standardised to the US adult population. When standardised to the US dialysis population, seroprevalence ranged from 3·5% (3·1–3·9) in the west to 27·2% (25·9–28·5) in the northeast. Comparing seroprevalent and case counts per 100 000 population, we found that 9·2% (8·7–9·8) of seropositive patients were diagnosed. When compared with other measures of SARS-CoV-2 spread, seroprevalence correlated best with deaths per 100 000 population (Spearman's ρ=0·77). Residents of non-Hispanic Black and Hispanic neighbourhoods experienced higher odds of seropositivity (odds ratio 3·9 [95% CI 3·4–4·6] and 2·3 [1·9–2·6], respectively) compared with residents of predominantly non-Hispanic white neighbourhoods. Residents of neighbourhoods in the highest population density quintile experienced increased odds of seropositivity (10·3 [8·7–12·2]) compared with residents of the lowest density quintile. County mobility restrictions that reduced workplace visits by at least 5% in early March, 2020, were associated with lower odds of seropositivity in July, 2020 (0·4 [0·3–0·5]) when compared with a reduction of less than 5%.InterpretationDuring the first wave of the COVID-19 pandemic, fewer than 10% of the US adult population formed antibodies against SARS-CoV-2, and fewer than 10% of those with antibodies were diagnosed. Public health efforts to limit SARS-CoV-2 spread need to especially target racial and ethnic minority and densely populated communities.FundingAscend Clinical Laboratories.

Antioxidant effect of lutein protects against oxidative damage to porcine spermatozoa

<p>Boar sperm is especially susceptible to peroxidative damage generated by reactive oxygen species (ROS). Chemiluminescence was initiated by incubating porcine semen in an in vitro ascorbate-Fe++ system, a technique that allows the evaluation of oxidative stress in these cells. Lutein is known for its antioxidant effects, chemically it is a dihydric derivative of α-carotene and belongs to the group of xanthophylls. The main objective of this study was to investigate the antioxidant effect of lutein on boar spermatozoa. The effect of lutein was analyzed by two methods: 1) by adding lutein: the sperm samples were placed in an in vitro ascorbate-Fe++ system, during 120 min at 37°C adding increasing amounts of lutein (50, 150 and 250 μg) per mg of protein and 2) by incubation with lutein in an in vitro ascorbate-Fe+2 system, for 120 min at 37°C, using spermatozoa obtained from porcine semen samples previously incubated with lutein (0.15 and 0.25 mg/ml) during 24 h at 15°C. In both methods a control group (without lutein) was used. Peroxidation was measured by chemiluminescence using a liquid scintillation counter, the light emission being quantified in cpm (counts per minute). Analyzing the effect of lutein by the two methods, it was observed that the total amount of cpm/mg of protein originated by chemiluminescence was lower in samples obtained from the lutein group than in the control group (without lutein). Total chemiluminescence (cpm total) was lower in samples obtained from the lutein group than in the control group (without lutein), with a significance of p<0.005. Percent inhibition of peroxidation was not concentration dependent. These results would demonstrate that lutein could act as an antioxidant that would protect the membranes of the sperm from oxidative damage.</p>

Updating the Chemiluminescence Oxygen-Aftereffect Method for Determining the Rate Constant of the Peroxy-Radical Self-Reaction: Oxidation of Cyclohexene.

Updating the facile chemiluminescence oxygen-aftereffect method, most suitable for determining the rate constant (kt ) of the peroxy-radical self-reaction (main chemiluminescence channel), pertained to considering the sensitivity of such a method toward a disturbing influence of the peroxy radicals of the initiator of the chain oxidation process. Such a disturbance may derive from the side chemiluminescent reaction, which involves peroxy radicals of both hydrocarbon and initiator. To examine the applicability and limitations of the chemiluminescence method under present scrutiny, cyclohexene was used as the model oxidizable hydrocarbon substrate. Computer simulations of the reaction and chemiluminescence kinetics have demonstrated the validity of the considered methodology at the value of the rate constant of the propagation of the overall chain process by peroxy radicals of the initiator higher than 1 m-1 s-1 . Despite that the chemiluminescence time profile and the stationary level of the total chemiluminescence intensity depend on the kinetics of the side chemiluminescence channel and on the ratio of the excited-state generation yields in the mentioned reaction channel and in the main chemiluminescence process, the value of kt assessed by the oxygen-aftereffect method has been found independent of variation of these characteristics.

Sensors and Biosensors for C-Reactive Protein, Temperature and pH, and Their Applications for Monitoring Wound Healing: A Review.

Wound assessment is usually performed in hospitals or specialized labs. However, since patients spend most of their time at home, a remote real time wound monitoring would help providing a better care and improving the healing rate. This review describes the advances in sensors and biosensors for monitoring the concentration of C-reactive protein (CRP), temperature and pH in wounds. These three parameters can be used as qualitative biomarkers to assess the wound status and the effectiveness of therapy. CRP biosensors can be classified in: (a) field effect transistors, (b) optical immunosensors based on surface plasmon resonance, total internal reflection, fluorescence and chemiluminescence, (c) electrochemical sensors based on potentiometry, amperometry, and electrochemical impedance, and (d) piezoresistive sensors, such as quartz crystal microbalances and microcantilevers. The last section reports the most recent developments for wearable non-invasive temperature and pH sensors suitable for wound monitoring.

Standardizing 25-hydroxyvitamin D data from the HunMen cohort.

Standardization of 25OHD values is still a challenge. We propose standardization by correction of the measured 25OHD values using the linear regression equation derived from the analysis of relationship between total 25OHD values measured by the methodology used by the laboratory and the NIST total target values (TV) reported by the DEQAS for all 5 of the DEQAS samples in a given survey. We applied our approach to standardize total 25OHD values of the HunMen cohort. All 206 samples for the HunMen cohort were evaluated using the automated Liaison DiaSorin total 25OHD chemiluminescence immunoassay (CLIA). The timing of these measurements coincided with that of the October 2015 DEQAS survey using samples 481 to 485. Following standardization, the mean total 25OHD changed from 53 to 62nmol/L and the prevalence of hypovitaminosis D (<75nmol/L) decreased significantly from 84 to 72%. A simple approach readily applicable at the routine diagnostic laboratory could perhaps be a practical solution to the anomaly surrounding non-standardized 25OHD values.

A numerical study on the ability to predict the heat release rate using CH* chemiluminescence in non-sooting counterflow diffusion flames

Numerical studies on 1-D, non-sooting counterflow diffusion flames were performed to determine the precision with which the total heat release rate can be calculated using light emission, namely, chemiluminescence, from the reaction zone. A detailed reaction mechanism, incorporating sub-reaction models for excited state radicals (CH* and OH*, where * denotes the excited state), was employed in this study. A set of 1-D, steady state conservation equations was solved under standard atmospheric conditions over a counterflow configuration utilizing the CHEMKIN-PRO package. A variety of fuels (CH4 and C3H8), velocities (0.1m/s − near the extinction condition), diluents (N2, H2O, CO2, and Ar), detailed reaction mechanisms for C1–C3 hydrocarbons (GRI-Mech 3.0, Hai Wang’s-, and San Diego-mechanism), different sub-reaction models for excited radicals, and excited radical transport properties were examined for the current purpose. It was found that a one-to-one correlation between total chemiluminescence from CH* and the total heat release rate cannot be sustained when the flame experiences a relatively high stretch and dilution, even though the condition is still far away from extinction. This trend is consistent with the different types of fuels, and it is understood that the reduction of the ethynyl radical (C2H), a potential precursor of CH*, is the main cause of the one-to-one correlation not being sustained. To this end, it was concluded that the observable light emission can only be used to predict the total heat release rate when non-sooting diffusion flames exist under velocity conditions from 0.1m/s to 1.5m/s. In other words, the chemiluminescence intensity does not always correlate with the total heat release rate of highly stretched flames found in practical combustors.

Combustion and emission characteristics of alcohol fuels in a CAI engine

The combustion and emission characteristics of alcohol fuels in a controlled auto-ignition (CAI) combustion engine have been investigated. Combustion characteristics were studied by heat releases analysis through in-cylinder pressure measurement and auto-ignition reactions through high-speed chemiluminescence imaging. By means of the negative valve overlap, varied amount of residual gas was trapped in the cylinder to achieve the controlled auto-ignition with/without spark-assisted ignition. Total chemiluminescence images were captured with a high speed camera equipped with a high repetition image intensifier. Methanol, ethanol and their blends with gasoline were tested and their combustion and emission characteristics were compared with the results of gasoline.Alcohol fuels, in particular methanol, resulted in advanced auto-ignition and faster combustion than that of gasoline. In addition, their use could lead to substantially lower HC, NOX and CO exhaust emissions. Spark-assisted ignition was most effective on gasoline combustion by advancing ignition timing and initiating flame kernel at the centre of combustion chamber but it had marginal effect on alcohol fuels. Auto-ignition always took place at the perimeter of the chamber and occurred earlier with alcohol fuels because of their greater heat release and fuel reforming reactions during the negative valve overlap period. Chemiluminescence of the gasoline fuel was most visible and it decreased with increasing percentage of oxygenated fuels.

Differentiating between intra‐ and extracellular chemiluminescence in diluted whole‐blood samples

The differentiation between extra- and intracellular production of reactive oxygen species (ROS) in whole blood was measured by luminol- and isoluminol-enhanced chemiluminescence (CL). Azide (total CL inhibition), azide + horseradish peroxidase (HRP, restoring extracellular CL), superoxide dismutase + catalase (depleting extracellular ROS) and HRP (enhancing extracellular CL) were used to modulate luminol- and isoluminol-enhanced CL (10(-6) -10(-3) m luminophores) of 125× diluted whole blood which was activated by both calcium ionophore A23187 (Ca-I) and opsonized zymosan particles (OZP) separately. Both activators stimulated intra- and extracellular production of ROS. Luminol-enhanced CL of Ca-I-activated samples detected the intracellular ROS, and with the addition of HRP detected the extracellular CL as well. CL enhanced with isoluminol in concentrations of 10(-4) m or less was mostly extracellular. There was a mixture of intra- and extracellular CL in OZP-activated samples, probably because of the ingestion of luminophore molecules. Measurement of Ca-I-activated CL enhanced with 10(-4) m luminol is recommended for the detection of intracellular ROS. The addition of HRP leads to the detection of overall ROS production while the OZP-activated system with its addition of HRP can only be used to detect overall ROS production. Ca-I-activated CL enhanced with 10(-4) m isoluminol and with addition of HRP is recommended for the detection of extracellular CL.

In-cylinder studies of hybrid combustion in a direct injection single-cylinder optical engine

Over the last decade, controlled autoignition (CAI) combustion, or homogeneous charge compression ignition (HCCI), has been the subject of numerous studies and has shown its potential to reduce fuel consumption and NO x emissions simultaneously from gasoline engines. However, there are still some fundamental questions that are yet to be addressed, such as combustion control and underlying physical and chemical processes. In the present study, a single-cylinder optical gasoline engine was operated with negative valve overlap to investigate CAI and hybrid spark ignition (SI)–CAI combustion. The effects of direct fuel injection and spark timings were studied by means of a simultaneous in-cylinder heat release study and high-speed visualization of total chemiluminescence, OH and CHO radicals.

A rapid antioxidant assay based on acidic potassium permanganate chemiluminescence

Abstract The total antioxidant capacity of fruit juices and teas was established using flow-injection analysis with acidic potassium permanganate chemiluminescence detection, and the results were compared to those of two conventional assays. The chemiluminescence procedure was rapid (over 100 samples could be run per hour) and precise (relative standard deviation assays was observed for 20 tea samples (r = 0.955). In general, green teas gave larger responses than black teas with both procedures, but there was considerable variation between brands. Moreover, when coupled with chromatographic separation, this chemiluminescence assay can be used to examine the contribution of individual antioxidants and monitor specific changes in foods and beverages during production and storage. Epigallocatechin and epigallocatechin gallate were found to contribute over 65% of the total chemiluminescence signal (integrated peak area) for a green tea sample. In the chromatogram for a black tea, these peaks were present but less prominent, and a small cluster of peaks representing theaflavins was observed.

Neutrophil chemiluminescence in pneumoconiosis

Neutrophilic phagocytic activity and serum opsonic activity were examined in pneumoconiosis patients. One hundred patients aged 43 to 81 were classified into four types (types 1 to 4) according to the degree of chest X-ray findings. A simultaneous multiple measurement system for luminol-dependent chemiluminescence was used. Neutrophilic phagocytic activity and serum opsonic activity were shown by the time length to reach peak chemiluminescence (PT) and the height of peak chemiluminescence (PH). There were no significant differences of neutrophilic phagocytic activities and serum opsonic activities between normal persons and type 1 cases of mild disease. However, these activities were elevated in cases with more severe disease, and the most severe type 4 type cases tended to show lower activities than type 2 and type 3 cases. PT of total chemiluminescence was influenced by both neutrophilic phagocytic activity and serum opsonic activity, but PH only by neutrophilic phagocytic activity. Statistically significant correlation was observed between serum opsonic activity and serum CH50 levels.

Spectroscopic and chemical-kinetic analysis of the phases of HCCI autoignition and combustion for single- and two-stage ignition fuels

The temporal phases of autoignition and combustion in an HCCI engine have been investigated in both an all-metal engine and a matching optical engine. Gasoline, a primary reference fuel mixture (PRF80), and several representative real-fuel constituents were examined. Only PRF80, which is a two-stage ignition fuel, exhibited a “cool-flame” low-temperature heat-release (LTHR) phase. For all fuels, slow exothermic reactions occurring at intermediate temperatures raised the charge temperature to the hot-ignition point. In addition to the amount of LTHR, differences in this intermediate-temperature heat-release (ITHR) phase affect the fuel ignition quality. Chemiluminescence images of iso-octane show a weak and uniform light emission during this phase. This is followed by the main high-temperature heat-release (HTHR) phase. Finally, a “burnout” phase was observed, with very weak uniform emission and near-zero heat-release rate (HRR). To better understand these combustion phases, chemiluminescence spectroscopy and chemical-kinetic analysis were applied for the single-stage ignition fuel, iso-octane, and the two-stage fuel, PRF80. For both fuels, the spectrum obtained during the ITHR phase was dominated by formaldehyde chemiluminescence. This was similar to the LTHR spectrum of PRF80, but the emission intensity and the temperature were much higher, indicating differences between the ITHR and LTHR phases. Chemical-kinetic modeling clarified the differences and similarities between the LTHR and ITHR phases and the cause of the enhanced ITHR with PRF80. The HTHR spectra for both fuels were dominated by a broad CO continuum with some contribution from bands of HCO, CH, and OH. The modeling showed that the CO + O → CO 2 + h ν reaction responsible for the CO continuum emission tracks the HTHR well, explaining the strong correlation observed experimentally between the total chemiluminescence and HRR during the HTHR phase. It also showed that the CO continuum does not contribute to the ITHR and LTHR chemiluminescence. Bands of H 2O and O 2 in the red and IR regions were also detected during the HTHR, which the data indicated were most likely due to thermal excitation. The very weak light emission in the “burnout” phase also appeared to be thermal emission from H 2O and O 2.

Evaluation of the intracellular radical generation by stimulated leukocytes by total luminol chemiluminescence

Luminol-enhanced chemiluminescence (LCL) is a widely used method for quantitative evaluation of reactive oxygen species (ROS) generated by stimulated leukocytes. The chemiluminescent kinetics registered (Total LCL) is an expression of simultaneously running processes of extra- and intracellular radical generation. In literature, model chemiluminescent systems, designed to separate extra- from the intracellular light, have been proposed. These systems, however, change cellular activity. The aim of the present study was to propose an approach for evaluating the intracellular ROS generation by stimulated leukocytes in whole blood. Heparin anti-coagulated blood from healthy volunteers was used. The chemiluminescent kinetics reflecting leukocyte activity to produce ROS were registered by a computerized chemiluminometer. To distinguish extra - from the intracellularly generated light by the cells, some model chemiluminescent systems were used. It was found that the intracellular light generated by stimulated leukocytes was a substantial part of Total LCL. The proportion between the extra- and intracellular light within the Total response depended on the concentration of the particulate stimulant. At a low zymosan concentration (0.25 mg/ml) Total LCL response was found to be intracellular. On the basis of the results obtained we conclude that Total LCL at a low stimulant concentration is of an intracellular origin, which could be used to study the effect of different compounds on the intracellular oxidative leukocyte activity. In the course of some pathological processes accompanied by suppurative foci formation, bacteriemia, ect., the proportion between the number of bacteria and leukocytes considerably changes. A change in the stimulant concentration allows to model these pathological states and to evaluate the extra- and intracellular radical generation by stimulated leukocytes. Scripta Scientifica Medica 2007;39(2):111-114

Effect of smoking on neutrophil oxidative metabolism

Alterations in neutrophil function by tobacco products may play a central role in the pathogenesis of periodontal diseases and several smoking-related systemic diseases. The aim of the study was to evaluate the effect of smoking on neutrophil oxidative metabolism. The study included 17 smoking men free of systemic diseases who were referred for treatment of various odontological diseases to outpatient department of Kaunas University of Medicine Hospital. The age of subjects varied from 22 to 43 years. All subjects answered the questions about smoking habits. Clinical examination included assessment of oral hygiene status according to the OHI-s index and periodontal status according to Russell and Ramfjord indices. To evaluate the oxidative metabolism of neutrophils, luminol- and liucigenin-dependent chemiluminescence and nitroblue tetrazolium test were used. After smoking, extracellular liucigenin-dependent chemiluminescence response was higher as compared to the response before smoking, but total (intra- and extracellular) luminol-dependent chemiluminescence response was the same both before and after smoking. Exposure of neutrophils to smoking caused a significant increase in nitroblue tetrazolium reduction. The release of reactive oxygen species in neutrophils exposed to smoking may alter the pathogenic processes in periodontal diseases.

Melatonin and N-acetyl serotonin inhibit selectively enzymatic and non-enzymatic lipid peroxidation of rat liver microsomes

Melatonin ( N-acetyl-5-methoxytryptamine) and its immediate precursor N-acetyl serotonin in the metabolism of tryptophan are free radical scavengers that have been found to protect against non-enzymatic lipid peroxidation in many experimental models. By contrast, little is known about the antioxidant ability of these indoleamines against NADPH enzymatic lipid peroxidation. The light emission produced by rat-liver microsomes, expressed as total cpm during 180 min of incubation at 37 °C, was two-fold greater in the presence of ascorbate (0.4 mM) when compared with NADPH (0.2 mM). Maximal peaks of light emission produced by microsomes lipid peroxidized with ascorbic-Fe 2+ or NADPH and expressed as cpm were 354,208 (at 60 min) and 135,800 (at 15 min), respectively. During non-enzymatic lipid peroxidation a decrease of total chemiluminescence (inhibition of lipid peroxidation) was observed when increasing concentrations of melatonin were added to liver microsomes. The protective effect was concentration-dependent. The inhibition observed in light emission was coincident with the protection of the most PUFAs. Preincubation of microsomes with N-acetyl serotonin reduced these changes very dramatically. Thus, in the presence of both antioxidants (0.36, 0.75, 1.5 mM), light emission percent inhibition during non-enzymatic (ascorbate-Fe 2+) lipid peroxidation of rat liver microsomes was for melatonin: 6.12, 16.20, 34.88 and for N-acetyl serotonin: 85.10, 88.48, 84.4 respectively. The incubation of rat liver microsomes in the presence of NADPH (0.36, 0.75, 1.5 mM) produce a sudden increase of chemiluminescence that gradually increased and reached a maximal value at about 15 min; however, N-acetyl serotonin reduced these changes very efficiently.