Ultra-Sensitive Isopropanol Biochemical Gas Sensor (Bio-Sniffer) for Monitoring of Human Volatiles

Our groups have previously developed a biochemical gas sensor to measure isopropanol (IPA) in exhaled air and have applied it for breath IPA investigation in healthy subjects and diabetes patients. In this study, the original bio-sniffer was modified with a series of components that improved the limit of detection (LOD). First, the modified IPA bio-sniffer used a C8855-type photomultiplier tube (PMT) that performed well in the photon sensitivity at the peak wavelength of nicotinamide adenine dinucleotide (NADH) fluorescence. Second, the multi-core bifurcated optical fiber, which incorporated 36 fibers to replace the previous dual-core type, enhanced the fluorescence collection. Third, the optical fiber probe was reinforced for greater width, and the flow-cell was redesigned to increase the area of the enzyme-immobilized membrane in contact with the air sample. These modifications lowered the detection limit to 0.5 ppb, a significant increase over the previous 1.0 ppb. Moreover, the modified bio-sniffer successfully analyzed the IPA concentration in exhaled air from a volunteer, which confirmed its capability for real-world sample detection. The modified bio-sniffer is more applicable to breath measurement and the detection of other extremely-low-concentration samples.

PATTERNS OF CHANGES IN FLUORESCENCE OF NADH AND FAD COENZYMES AND THEIR RELATIONSHIP IN SKELETAL MUSCLE IN EARLY POST-MORTEM PERIOD (EXPERIMENTAL STUDY)

The paper presents changes and mathematical models of autofluorescence of reduced nicotinamide adenine dinucleotide (NADH) and flavin adenine dinucleotide (FAD) and their redox ratio (RR) in the rat skeletal muscle during the first 24 hours after death. Aim. To establish a pattern of change in the fluorescence intensity of NADH and FAD coenzymes, as well as their relationship in the skeletal muscle during the first 24 hours after death using mathematical modeling. Material and methods. The experiment was performed on Sprague Dawley rats. The fluorescence intensity of FAD and NADH coenzymes in skeletal muscle was measured in situ in living rats under general anesthesia, 5 minutes after euthanasia, and at intervals of 1.5-3 h during the 24-hour postmortem period. To estimate the fluorescence intensity of NADH and FAD coenzymes and to calculate the RR, the Lasma MC-3 fluorescence measuring device with special software was used. The obtained data were analyzed by non-linear regression analysis. The summary, accuracy estimation and significance of the regression equation coefficients were assessed using SigmaPlot 10.0 software. The significance of the regression model was tested using the Fisher F-criterion. Results. During the first 3 hours of the postmortem period, an increase in the mean values of RR and NADH fluorescence was detected, starting from 4.5 and till 24 hours post-mortem their gradual decrease was observed. The relationship between NADH, RR and time after death is characterized by the Weibull equation. The statistical significance of NADH and RR models, based on the obtained equations was considered high. Conclusion. The analysis of the obtained data allowed to create mathematical models describing the relationship between RR and NADH fluorescence intensity and time after death, which confirms non-randomness and regularity of the discovered patterns.time of death; NADH; FAD; auto-fluorescence; redox ratio; early post-mortem changes

The Effect of a 7-Week Training Period on Changes in Skin NADH Fluorescence in Highly Trained Athletes

The study aimed to evaluate the changes of nicotinamide adenine dinucleotide (NADH) fluorescence in the reduced form in the superficial skin layer, resulting from a 7-week training period in highly trained competitive athletes (n = 41). The newly, non-invasive flow mediated skin fluorescence (FMSF) method was implemented to indirectly evaluate the mitochondrial activity by NADH fluorescence. The FMSF measurements were taken before and after an exercise treadmill test until exhaustion. We found that athletes showed higher post-training values in basal NADH fluorescence (pre-exercise: 41% increase; post-exercise: 49% increase). Maximum NADH fluorescence was also higher after training both pre- (42% increase) and post-exercise (47% increase). Similar changes have been revealed before and after exercise for minimal NADH fluorescence (before exercise: 39% increase; after exercise: 47% increase). In conclusion, physical training results in an increase in the skin NADH fluorescence levels at rest and after exercise in athletes.

Changes of NADH Fluorescence from the Skin of Patients with Systemic Lupus Erythematosus

Introduction. The blood circulation of the skin is an accessible and representative vascular bed for examining the mechanisms of microcirculatory function. Endothelial function is impaired in systemic lupus erythematosus (SLE), which implies disorders in cell metabolism dependent on blood circulation; however, noninvasive monitoring of metabolism at the tissue and cell level is absent in daily clinical practice. Objective. The aim of the study was to examine changes of NADH fluorescence from the epidermis of a forearm measured with the flow mediated skin fluorescence (FMSF) technique in patients with SLE and to investigate whether they are associated with clinical manifestation of the disease. Materials and Methods. The study enrolled 36 patients with SLE and 34 healthy individuals. Changes of NADH fluorescence were measured using FMSF on the forearm in response to blocking and releasing of blood flow. The results were represented as ischemic (IR max and IR auc) and hyperemic response maximum and area under the curve (HR max and HR auc). Results. IR max, IR auc, HR max, and HR auc were all lower in patients with SLE (p<0.05) compared with controls. All four parameters were negatively correlated (p<0.05) with patient age. No difference was found in NADH fluorescence between SLE patients with malar rash, discoid rash, photosensitivity, oral ulcers, nonerosive arthritis, renal disorder, hematologic disorder, or immunologic disorder and those without. No correlation was revealed between the SLEDAI score and NADH fluorescence. Conclusion. Changes of NADH fluorescence indicate the reduction in NADH restoration, observed especially during reperfusion, and suggest the occurrence of disorders in the microcirculation of the skin and/or at the mitochondrial level. Such changes of NADH during reperfusion in patients with SLE could be associated with their possible lower sensitivity to hypoxia and possibly with endothelial dysfunction.

NADH fluorescence imaging and the histological impact of cortical spreading depolarization during the acute phase of subarachnoid hemorrhage in rats

OBJECTIVEAlthough cortical spreading depolarization (CSD) has been observed during the early phase of subarachnoid hemorrhage (SAH) in clinical settings, the pathogenicity of CSD is unclear. The aim of this study is to elucidate the effects of loss of membrane potential on neuronal damage during the acute phase of SAH.METHODSTwenty-four rats were subjected to SAH by the perforation method. The propagation of depolarization in the brain cortex was examined by using electrodes to monitor 2 direct-current (DC) potentials and obtaining NADH (reduced nicotinamide adenine dinucleotide) fluorescence images while exposing the parietal-temporal cortex to ultraviolet light. Cerebral blood flow (CBF) was monitored in the vicinity of the lateral electrode. Twenty-four hours after onset of SAH, histological damage was evaluated at the DC potential recording sites.RESULTSChanges in DC potentials (n = 48 in total) were sorted into 3 types according to the appearance of ischemic depolarization in the entire hemisphere following induction of SAH. In Type 1 changes (n = 21), ischemic depolarization was not observed during a 1-hour observation period. In Type 2 changes (n = 13), the DC potential demonstrated ischemic depolarization on initiation of SAH and recovered 80% from the maximal DC deflection during a 1-hour observation period (33.3 ± 15.8 minutes). In Type 3 changes (n = 14), the DC potential displayed ischemic depolarization and did not recover during a 1-hour observation period. Histological evaluations at DC potential recording sites showed intact tissue at all sites in the Type 1 group, whereas in the Type 2 and Type 3 groups neuronal damage of varying severity was observed depending on the duration of ischemic depolarization. The duration of depolarization that causes injury to 50% of neurons (P50) was estimated to be 22.4 minutes (95% confidence intervals 17.0–30.3 minutes). CSD was observed in 3 rats at 6 sites in the Type 1 group 5.1 ± 2.2 minutes after initiation of SAH. On NADH fluorescence images CSD was initially observed in the anterior cortex; it propagated through the entire hemisphere in the direction of the occipital cortex at a rate of 3 mm/minute, with repolarization in 2.3 ± 1.2 minutes. DC potential recording sites that had undergone CSD were found to have intact tissue 24 hours later. Compared with depolarization that caused 50% neuronal damage, the duration of CSD was too short to cause histological damage.CONCLUSIONSCSD was successfully visualized using NADH fluorescence. It propagated from the anterior to the posterior cortex along with an increase in CBF. The duration of depolarization in CSD (2.3 ± 1.2 minutes) was far shorter than that causing 50% neuronal damage (22.4 minutes) and was not associated with histological damage in the current experimental setting.