Effect of addition of Syrian thyme (Thymus syriacus) on physiochemical and sensory quality of Sudanese Mudaffara cheese during storage.

This study was designed to introduce Syrian thyme (Thymus syriacus) as a new additive flavor for Mudaffara cheese. Mudaffara cheese was prepared from cow's milk using the commonly used black cumin as control and Syrian thyme (0.3 and 0.5%) as treatment. The physiochemical properties and the sensory attributes were evaluated. The results indicated that Mudaffara cheese samples flavored with 0.3% Syrian thyme were significantly (P < 0.05) higher in protein and acidity content compared to the other cheeses. During the storage period, significant (P < 0.05) differences were obtained for all the studied physicochemical parameters except the ash content. Also the interaction of additives and storage period showed significant (P < 0.05) effect on the protein and fat content of Mudaffara cheese samples. However the additives had no significant effect on all sensory characteristics except the general acceptability. According to the panelist test, the overall acceptability of Mudaffara cheese sample flavored with 0.5% Syrian thyme showed the highest numerical score compared to the others Mudaffara cheese samples. During the storage period, Mudaffara cheese samples revealed significant (P < 0.05) variations in texture, acidity, flavor, taste and general acceptability scores. This study concluded that Mudaffara cheese can be flavored with Syrian thyme at a rate of 0.3 and 0.5%. Supplementary Information: The online version contains supplementary material available at 10.1007/s13197-022-05634-7.

Association of the polymorphism of the Toll-like receptor (TLR)-3 and TLR-9 genes with hepatitis C virus-specific cell-mediated immunity outcomes among Egyptian health-care workers.

Variations in the immune response could explain resistance to hepatitis C virus (HCV) infection. Toll-like receptor gene (TLR)-3 is an innate detector of dsRNA viruses, and the TLR-9 gene recognizes bacterial and viral unmethylated cytosine-phosphate-guanosine (CpG) motifs. We previously reported that the TLR-3.rs3775290 CC genotype was associated with HCV chronicity and that the TLR-9 gene played no major role in this infection. This study identified the role of TLR-3.rs3775290 (c.1377C/T), TLR-9.rs5743836 (-1237T→C) and TLR-9.rs352140 (G2848A) gene polymorphisms in predicting the outcome of HCV-specific cell-mediated immunity (CMI) among Egyptian health-care workers (HCWs). We enrolled 265 HCWs in this study and divided them into four groups. Group 1: 140 seronegative-aviraemic HCWs; group 2: 20 seronegative-viraemic HCWs; group 3: 35 subjects with spontaneously resolved HCV infection; and group 4: 70 chronic HCV HCWs (patients). All subjects were genotyped by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis for the TLR-3.rs3775290, TLR-9.rs5743836 and TLR-9.rs352140 single nucleotide polymorphisms (SNPs). We also quantified HCV-specific CMI in the four groups using an interferon (IFN)-γ enzyme-linked immunospot (ELISPOT) assay in response to nine HCV genotype 4a, overlapping 15mer peptide pools covering the whole viral genome. No statistically significant difference was found between CMI-responding subjects with different HCV states and TLR-3.rs3775290 or TLR-9.rs352140 genotypes. However, there was a significant relationship between the outcome of the HCV-specific CMI and the TLR-9.rs5743836 genotype among the responding subjects (P = 0·005) and the chronic HCV patients (P = 0·044). In conclusion, TLR-9.rs5743836 SNP, but not TLR-3.rs3775290 or TLR-9.rs352140 genotypes, could predict the outcome of HCV-specific CMI responses among Egyptians infected with genotype-4.

Multiple pathways of the reaction of 2,2-diphenyl-1-picrylhydrazyl radical (DPPH·) with (+)-catechin: evidence for the formation of a covalent adduct between DPPH· and the oxidized form of the polyphenol.

The pathways of the reaction of 2,2-diphenyl picrylhydrazyl radicals (DPPH·) with (+)-catechin were studied in alcoholic solvents. The reaction mixtures were analysed by using reversed-phase liquid chromatography (HPLC) and electrospray ionization mass spectrometry (ESI-MS). The intermediate o-quinone of catechin, yellow dimers, trimers and, interestingly, an adduct of the oxidized form of catechin with DPPH radicals were identified. The mass of this adduct was 681 Da, suggesting that one molecule of the DPPH radical complexes with the oxidized form of catechin. It is concluded that once the intermediate o-quinone is formed, the reaction proceeds in two pathways, either the o-quinone reacts with catechin to form a hydrophilic dimer (type B), which is further oxidized to hydrophobic dimers (type A) and consequently to oligomers of higher molecular weights; or the A-ring of the o-quinone is further oxidized by a DPPH radical and that this oxidized intermediate then reacts with another DPPH radical to form the observed adduct. The identification of the latter mechanism could explain the contradictory results reported in the literature for the reaction of polyphenols with DPPH radicals.

Thyroid Dysfunctions in Sudanese Patients with Vitiligo

Introduction: Vitiligo is a chronic acquired skin condition that causes loss of pigment; resulting in irregular pale patches of skin. The precise cause of vitiligo is not fully understood. The autoimmune base of the disease is supported by the frequent observation that several autoimmune disorders; particularly thyroid diseases; are associated with vitiligo. Objective: To determine the frequency of thyroid dysfunctions in Sudanese patients with vitiligo. Methods: Two groups; i.e. vitiligo patients and control; were collected with simple random collection. The control group included individuals free of vitiligo. 5 ml of venous blood was taken from every individual in both groups and the ELISA test was done for thyroid hormones; i.e. T3; T4 and TSH; using the DRG-USA kits. Results: The number of patients with vitiligo in the study was 46; while the control group was 45. Nine (19.56) patients were found to have abnormal levels of thyroid hormones. No abnormal levels in the control group. Mean T3 level in patients was 1.463ng/l; while in control group it was 1.467ng/l. Mean T4 level in patients was 102.761 nmol/l; while in control group it was 90.844 nmol/l. Mean TSH level in patients was 0.841 ?IU/l; while in control group it was 1.50 ?IU/l. The t-test was done to determine the significance of difference between means of T3; T4; and TSH between the patients and control groups. The P-values were found to be significant.Conclusion: There is a strong pathogenetic relationship between vitiligo in Sudanese patients and thyroid dysfunctions

The psychological impact of vitiligo in adult Sudanese patients.

OBJECTIVE: Vitiligo is a chronic skin disease that causes loss of pigment, resulting in irregular pale patches of skin. The disease has profound psychological consequences. These effects range from mild embarrassment to a severe loss of self-confidence and social anxiety, especially for those who have lesions on exposed skin. The study sought to determine the psychological impact of vitiligo in Sudanese patients. METHOD: This study is a cross-sectional, clinical-epidemiological and hospital-based study, undertaken in Khartoum Dermatologic Hospital (KDH). The data was collected between June 2007 and November 2007. 111 adult patients were enrolled sequentially during the study period and they were tested using the 12-item General Health Questionnaire (GHQ-12). RESULTS: Psychological disturbances as a consequence of vitiligo were found in 36 (31 %) adult patients. Patients with mild psychological disturbances were found in 20 of these patients and severe disturbances in 16. CONCLUSION: Psychological consequences are common in patients with vitiligo.

Diurnal distribution of airborne bacteria and fungi in the atmosphere of Helwan area, Egypt.

Airborne bacterial and fungal composition in an industrial town of Helwan, Egypt, was studied using a slit impactor sampler during the period from March 2006 to February 2007. Airborne bacterial concentrations were usually higher than fungi. Bacteria and fungi had similar diurnal variation patterns. Airborne microorganisms reached their concentration peaks in the evening and gradually decreased during the night time. The hourly concentration peaks of the bacteria and fungi appeared at 20:00h. A significant difference (P < or =0.05) was found between the hourly mean concentrations of airborne fungi in winter compared to other seasons. Fungi concentrations were significantly higher (P< or =0.05) on working weekdays than weekends. Aspergillus, Penicillium, Alternaria and Cladosporium were the most predominant airborne fungal genera. Aspergillus showed double peak patterns whereas Penicillium, Alternaria and Cladosporium showed one peak pattern. The diurnal variations of the bacteria and fungi could be divided into four periods: 1) the morning maximum concentration (6:00h-10:00h), 2) midday to afternoon pattern (10:00h-16:00h), 3) the evening concentration peak (18:00h-20:00h) and 4) the gradual decrease of night time concentration (22:00h-24:00h). Geographical location, human activity, growth cycle of organisms and meteorological factors were the main criteria controlling the temporal variations of the air microorganisms in the Wadi Hof area.

Anti-CCP2 is an adjunct to, not a surrogate for, rheumatoid factor in the diagnosis of rheumatoid arthritis: diagnostic utility of anti-CCP2 antibodies in Egyptian patients with rheumatoid arthritis.

OBJECTIVES: To examine the diagnostic utility of the second generation of anti-cyclic citrullinated peptide (anti-CCP2) antibodies versus rheumatoid factor (RF) in rheumatoid arthritis (RA), and to study the association between anti-CCP2 and RA disease parameters. METHODS: Fifty consecutive Egyptian patients with RA, 37 patients with other rheumatic diseases, and 10 healthy controls were recruited for testing for anti-CCP2 and immunoglobulin M (IgM) rheumatoid factor (RF). Assessment measures included the Disease Activity Score (DAS28) for disease activity, the Health Assessment Questionnaire - Disability Index (HAQ-DI) for disability and the Short Erosion Narrowing Score (SENS) for radiological damage. RESULTS: The sensitivities of anti-CCP2 and IgM-RF in RA patients were 70% and 52%, with specificities of 91.5% and 89.4%, respectively. There was 73.2% agreement between anti-CCP2 and RF for all groups tested (kappa = 0.42, p<0.001) but agreement was only 66% for RA patients (kappa = 0.31, p<0.05). Anti-CCP2 had superior diagnostic properties [sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV)] than RF, but using both RF and anti-CCP2 enhanced the sensitivity to 78%, when either test was positive, and the specificity to 100%, with a PPV of 1, when both tests were positive. Anti-CCP2 titre was significantly correlated with disease severity [rheumatoid nodules, rheumatoid factor (RF), and radiological damage] and HAQ-DI (p<0.05) but not with parameters of disease activity. CONCLUSION: Anti-CCP2 has superior diagnostic and prognostic properties in RA compared with RF. It should not replace RF as a serological test; however, since using both tests modestly increases sensitivity and markedly enhances specificity, so that diagnosis of RA is highly probable when both tests are positive.

Comparison of key enzymes in the zebra mussel, Dreissena polymorpha, the earthworm Allolobophora chlorotica and Chironomus riparius larvae.

The levels of the enzymes, glutathione S-transferase, catalase, NAD(P)H-cytochrome c reductases, and DT-diaphorase were determined and compared in the tissues of three invertebrates commonly used in monitoring environmental quality: a freshwater mussel, Dreissena polymorpha, the earthworm Allolobophora chlorotica and the fourth instar of Chironomus riparius. It was found that the activities of GST, catalase, and NAD(P)-cytochrome c reductases were comparable in A. chlorotica and C. riparius, whereas comparatively a higher GST and a lower catalase activity was determined in the mussel tissues. DT-diaphorase was not detectable in A. chlorotica and the C. riparius larvae tissues, whereas this enzyme is present in the gills and the rest of soft mussel tissues (soft mussel tissues minus gills). It is suggested that the relatively low catalase activity observed in the tissues of the latter organism might be compensated by the presence of the antixidant role of DT-diaphorase. In addition, the inducibility of DT-diaphorase in D. polymorpha, by butylated hydroxyanisole (BHA) and lead (Pb) was investigated. Despite the bioaccumulation of both BHA (5.2+/-0.14 microgg(-1) wet weight) and Pb (233.7+/-0.95 mgkg(-1) dry weight) in the soft mussel tissues, the mussel DT-diaphorase was not induced. Although the activity of NADPH-cytochrome c (P-450) reductase was also not affected by these reagents, its activity was 2-fold higher in the gills than the rest of soft mussel tissues.

Differential responses of biomarkers in tissues of a freshwater mussel, Dreissena polymorpha, to the exposure of sediment extracts with different levels of contamination.

In this study, zebra mussels, D. polymorpha, were exposed to extracts of sediments obtained from two sites, a contaminated lake (Ketelmeer, Km) and a relatively clean lake (Drontenmeer, Dm). The main objective of this work was to investigate whether six selected biomarkers could discriminate between the two sediments. The selected biomarkers included phase I enzymes such as DT-diaphorase, NADPH-cytochrome c reductase, NADH-cytochrome c reductase, a phase II enzyme (glutathione S-transferase, GST), an antioxidant enzyme, catalase, and the total glutathione, reduced (GSH) and oxidized (GSSG). After a short (24 h) and a long-term (7 days) exposure, the levels of these biomarkers were measured in gills and the rest of soft mussel tissues (soft mussel tissue minus gills) and compared with control values. A decrease of GST level by 20% (P = 0.004) and a 4-fold decrease of total glutathione concentration relative to the control, were observed in the gills of mussels exposed to the more contaminated Km extract. No significant differences in the GST activities were observed in the gills of control and Dm extract-treated mussels (P = 0.23). Although the levels of catalase and NADH-cytochrome c reductase were, in the short-term exposure, unaffected, both activities were, in the long-term exposure, reduced in the gills of the mussels exposed to the contaminated Km extract, compared with control values, by 43% and 20%, respectively. The activities of DT-diaphorase and NADPH-cytochrome c reductase remained unaffected in all exposure conditions. However, the level of NADPH-cytochrome c reductase was found higher in gills than in the rest of soft mussel tissues. This difference in the ratio of the two reductases between the two tissues could account for the observed differential responses of the biomarkers.

ABTS radical-driven oxidation of polyphenols: isolation and structural elucidation of covalent adducts.

The formation of covalent adducts obtained from the reaction of the polyphenols, trans-3,3',4',5,7-pentahydroxyflavan (catechin) and 1,3,5-trihydroxybenzene (phloroglucinol), with ABTS radicals is reported. Two adducts derived from (+)-catechin and three adducts from phloroglucinol were isolated and identified using reversed-phase high performance liquid chromatography (HPLC) and electrospray ionization mass spectrometry (ESI-MS). The molecular masses of the (+)-catechin-derived adducts (I(c) and II(c)) were found to be 802 and 559 Da, respectively, whereas the masses of phloroglucinol-derived adducts (I(p), II(p), and III(p)) were 638, 395, and 381 Da, respectively. The initially formed adducts (I(c), I(p)) were unstable and degraded to secondary adducts (II(c), II(p), and III(p)) releasing part of the ABTS molecule. The structures of these adducts were elucidated by interpreting the results of MS/MS analysis of prominent ions generated by both positive and negative ion ESI-MS. The adducts were found to scavenge ABTS radicals, an observation that could explain the complex kinetic behaviour manifested by the reactions of ABTS radicals with polyphenols. A mechanism, which accounts for both the formation of the adducts and the degradation products of ABTS radicals, is proposed.

Isolation and the characterization of the degradation products of the mediator ABTS-derived radicals formed upon reaction with polyphenols.

Two degradation products were obtained from the incubation of the widely used 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid), ABTS, radical cations with the polyphenols, (+)-catechin, (-)-epicatechin, and phloroglucinol in acetate buffer (pH 5). The products were purified by reversed-phase chromatography and characterized by UV-visible detection, mass spectrometry, and (1)H NMR spectroscopy. The data allowed us to identify the degradation products as 3-ethyl-6-sulfonate benzothiazolinone imine and the corresponding sulfoxide, 3-ethyl-6-sulfonate benzothiazolone. Elemental composition strongly supported the proposed structures. Our results unequivocally demonstrated that ABTS radicals are not as stable as usually claimed because they could be degraded upon interaction with polyphenols, in addition to being reduced by these antioxidants back to the parent compound. Therefore, it is concluded that caution must be exercised in using ABTS radicals as a basis for the evaluation of antioxidant capacities of pure compounds and/or complex mixtures.

In vivo exposure of Dreissena polymorpha mussels to the quinones menadione and lawsone: menadione is more toxic to mussels than lawsone.

The principal aim of this study was to assess whether the two quinones, menadione (2-methyl-1,4-naphthoquinone) and lawsone (2-hydroxy-1,4-naphthoquinone), elicit differential toxicity in mussels as has been reported for higher organisms. Therefore, the effects of short-term (48 h) and long-term (20 days) exposure of the two quinones at concentrations of 0.56 and 1 mg l(-1) to zebra mussels, Dreissena polymorpha, under laboratory conditions were studied. After the short-term exposure, the specific activities of the two-electron quinone oxidoreductase (DT-diaphorase) and the one-electron catalysing quinone reductases NADPH-cytochrome c reductase and NADH-cytochrome c reductase were determined in the gills and the rest of the soft tissues (soft mussel tissues minus the gills) of both treated and control mussels. At the higher concentrations of menadione and lawsone used, a significant reduction of the activity of NADPH-cytochrome c reductase in the gills and in the rest of the soft mussel tissues (by 33-34% and 31-43%, respectively) was observed. The activities of DT-diaphorase and NADH-cytochrome c reductase were not significantly affected. Interestingly, DT-diaphorase was observed in the gills, an organ requiring protection against antioxidants. Furthermore, a single-cell electrophoretic assay (comet assay) performed with gill cells to assess DNA damage by the quinones did not show any significant difference between the treated and the control organisms. This indicates that the formation of reactive species by the quinone metabolism in vivo in the mussels was possibly suppressed through the concerted action of DT-diaphorase and antioxidant enzymes. The results of in vitro experiments with gill extracts confirmed the protective role of DT-diaphorase. The rate of the two-electron quinone reduction was found to be five times that of the one-electron quinone reduction. The results of the long-term exposure unambiguously demonstrated that in mussels menadione, unlike in higher organisms, is more toxic than lawsone. The lack of detectability of xanthine oxidase in the mussel tissues could explain the comparatively lower toxicity of lawsone in the invertebtrate, lending support to a previous suggestion that xanthine oxidase might be responsible for the mechanism of toxicity of lawsone in higher organisms in vivo.

Menadione enhances oxyradical formation in earthworm extracts: vulnerability of earthworms to quinone toxicity.

NAD(P)H-cytochrome c reductase activities have been determined in the earthworms, L. rubellus and A. chlorotica, extracts. Menadione (0.35 mM, maximum concentration tested) was found to stimulate the rates of NADPH- and NADH-dependent cytochrome c reduction by three- and twofold, respectively. Superoxide dismutase (SOD) inhibited completely this menadione-mediated stimulation, suggesting that *O2- is involved in the redox cycling of menadione. However, SOD had no effect on the basal activity (activity in the absence of quinone) in the case of NADH-dependent cytochrome c reduction, whereas it partially inhibited the basal activity of NADPH-cytochrome c reduction. This indicates direct electron transfer in the former case and the formation of superoxide anion in the latter. DT-diaphorase, measured as the dicumarol-inhibitable part of menadione reductase activity, was not detectable in the earthworms' extracts. In contrast, it was found that DT-diaphorase represents about 70% of the menadione reductase activities in the freshwater mussel, Dreissena polymorpha. The results of this work suggest that earthworms, compared with mussels, could be more vulnerable to oxidative stress from quinones due to lack, or very low level of DT-diaphorase, an enzyme considered to play a significant role in the detoxification of quinones. On the contrary, mussels have efficient DT-diaphorase, which catalyzes two-electron reduction of menadione directly to hydroquinone, thus circumventing the formation of semiquinone.

Evidence for redox cycling of lawsone (2-hydroxy-1,4-naphthoquinone) in the presence of the hypoxanthine/xanthine oxidase system.

This study reports that lawsone (2-hydroxy-1,4-naphthoquinone) undergoes redox cycling in the presence of the hypoxanthine/xanthine oxidase system. The rate of cytochrome c reduction obtained in the presence of 80 microM lawsone was almost three times the rate of cytochrome c reduction measured in its absence. This increase in the rate of cytochrome c reduction was partially inhibited by superoxide dismutase, suggesting the involvement of O(2)(.-) in this process. It is remarkable to note that, even though lawsone is considered to be a non-redox-cycling quinone in vitro, this quinone was shown to be more toxic in vivo in rats than menadione, causing haemolytic anemia of an oxidative nature and renal damage. The view that this quinone is a non-redox-cycling quinone was based on the inability of one-electron-transferring flavoenzymes such as NADPH-cytochrome c reductase to reduce this naphthoquinone. Our finding that lawsone, like menadione, undergoes redox cycling in the presence of the hypoxanthine/xanthine oxidase system could explain the observed oxidative damage of tissues inflicted by this quinone in rats in vivo. Such an observation therefore reconciles the in vivo toxicity results of this naphthoquinone with those of in vitro experiments.

Increased plasma endothelin-1 and cardiac nitric oxide during doxorubicin-induced cardiomyopathy.

The major limiting factor in long-term administration of doxorubicin is the development of cumulative dose-dependent cardiomyopathy and congestive heart failure. Although several mechanisms have been suggested to explain the exact cause of doxorubicin-induced cardiomyopathy, the role of the vascular endothelium-derived vasoactive mediators in the pathophysiology of this toxic effect is still unknown. Accordingly, the present study has been initiated to investigate whether the changes in plasma level of endothelin-1 and nitric oxide along with cardiac nitric oxide are associated with the development of doxorubicin-induced cardiomyopathy. Doxorubicin was injected with a single dose of 5 mg/kg and every other day with a dose of 5 mg/kg, intraperitoneally, to have four cumulative doses of, 10, 15, 20 and 25 mg/kg in five separate groups of male rats. An additional group receiving a single dose of 20 mg/kg and one receiving normal saline were also included in the study. Twenty-four hr after the last dose, the animals were sacrificed and the plasma levels of endothelin-1 and nitric oxide in addition to cardiac nitric oxide were determined. The results show that doxorubicin caused a statistically significant increase of 85%, 76% and 97% in plasma endothelin-1 at a cumulative dose levels of 10, 15 and 20 mg/kg, respectively. However, the level of plasma nitric oxide remained unchanged. Furthermore, doxorubicin treatment resulted in a significant dose-dependent increase in serum lactate dehydrogenase and creatine phosphokinase. In contrast, the increase in nitric oxide production in cardiac tissue by doxorubicin was not dose-dependent with the maximum increase (81%) at a cumulative dose of 10 mg/kg. It is worth mentioning that plasma endothelin-1 and cardiac nitric oxide were significantly increased at 24 hr after the single dose of 20 mg/kg doxorubicin. The increase of plasma endothelin-1 and cardiac nitric oxide with the cardiomyopathy enzymatic indices, may point to the conclusion that both endothelin-1 and cardiac nitric oxide are increased during the development of doxorubicin-induced cardiomyopathy.

A unified theory of enzyme kinetics based upon the systematic analysis of the variations of k(cat), K(M), and k(cat)/K(M) and the relevant DeltaG(0 not equal) values-possible implications in chemotherapy and biotechnology.

To elucidate the kinetic properties of critical enzymatic situations that have previously escaped classification, we performed a systematic analysis of all the possible variations of the kinetic constants k(cat,) K(M,) and k(sp) = k(cat)/K(M,) encompassing all aspects of enzymology. The equation gives a total of thirteen theoretically possible cases, comprising the reference case plus 12 different sets of variations, which can be divided into six principal cases and six specular ones. The six relevant cases are examined individually in the context of each of the main chapters of enzymology, i.e. as regards mechanism of action, specificity of substrate and isoenzyme, reversible and irreversible inhibition, and mutation of residues (enzyme evolution and enzyme engineering). Some critical cases where k(sp) does not hold as a specificity index are classified for the first time. Interestingly, the six possible cases correspond to the five known cases of reversible inhibition (competitive, non-competitive, incompetitive, mixed competitive/non-competitive, and mixed incompetitive/non-competitive) plus an additional case of biphasic nature (activation-inhibition), which is crucial for a full understanding of specificity and which leads us to propose some modification to the definition of enzyme specificity. The systematic approach to enzymology outlined herein could find practical applications in various sectors of biotechnology, including chemotherapy.

Enhanced sensitivity of Cypridina luciferin analogue (CLA) chemiluminescence for the detection of *O2(-) with non-ionic detergents.

Superoxide anion-triggered chemiluminescence of Cypridina luciferin analogue (CLA), 2-methyl-6-phenyl-3,7-dohydroimidazo[1,2-alpha]pyrazin-3-one, is enhanced by non-ionic detergents such as Tween 20, Triton X-100 and Tween 80. At the concentration of 0.6% (v/v) the largest increase (2.7-fold) of CLA light intensity was obtained with Tween 20, followed by Tween 80 and Triton X-100. Using this detergent-amplified CLA chemiluminescence, the detection limits of xanthine and xanthine oxidase were examined at pH 7.4 and reinvestigated at pH 5.5. At pH 5.5, concentrations of xanthine and xanthine oxidase as low as 5 nmol/L and 3.85 x 10(-7) U/mL, respectively, could be accurately determined, whereas, under the experimental conditions used, at pH 7.4 the lowest concentrations of xanthine and xanthine oxidase detectable were 10 nmol/L and 3.85 x 10(-6) U/mL. The lowest detectable values of xanthine and xanthine oxidase obtained at pH 5.5 are about 400 and 10 times lower than those previously reported. The detection limit of xanthine (5 nmol/L) by this chemiluminescent-based method is about 200 and 20 times more sensitive than the determination of xanthine by enzymatic means or by HPLC with detection limits of 1 micromol/L and 0.1 micromol/L, respectively. Our data suggest that this chemiluminescent probe can detect concentrations of superoxide anion below the nanomolar range.

Comparative studies of the chemiluminescent horseradish peroxidase-catalysed peroxidation of acridan (GZ-11) and luminol reactions: effect of pH and scavengers of reactive oxygen species on the light intensity of these systems.

In this study, the chemiluminescent horseradish peroxidase/H(2)O(2)-catalysed oxidation of acridan (GZ-11) substrate was compared with the well-characterized light-producing luminol reaction. p-Iodophenol and p-phenylphenol were used as enhancers, respectively, for the luminol and acridan reactions. These two light-producing systems showed significant differences in relation to the effect of pH, as well as the effect of scavengers of reactive oxygen species, on the light intensity. Light production measured could be as low as pH 2.6 in the acridan reaction, whereas light emission was not detected in the luminol system below pH 5.6. In contrast with the luminol system, it was found that superoxide dismutase does not inhibit the light intensity of the acridan system. This suggests that superoxide anion does not participate in the mechanism of the light-emitting steps of the acridan reaction. Addition of hydroxyl radical scavengers, mannitol and benzoate, to the acridan reaction medium had no appreciable effect on the chemiluminescent intensity, indicating that hydroxyl radicals do not interfere in light-emitting steps. In addition, the peroxidation of the acridan substrate was found to be very slow at pH 5.6 in the absence of the enhancer, p-phenylphenol, whereas in its presence a rapid degradation of the acridan substrate was observed. Therefore, it is suggested that the enhancer might be initially oxidized by the HRP/H(2)O(2) system, resulting in the formation of the enhancer radical, which could be the actual oxidizing agent of the acridan substrate. Together, the data presented in this paper indicate that the chemiluminescent horseradish peroxidase-catalysed peroxidation of acridan (GZ-11) is more specific than the luminol reaction for the reactive oxygen species involved in the light-emitting steps, i. e, H(2)O(2).

Propionyl-L-carnitine as potential protective agent against adriamycin-induced impairment of fatty acid beta-oxidation in isolated heart mitochondria.

Propionyl-L-carnitine (PLC), a natural short-chain derivative of L-carnitine, has been tested in this study as a potential protective agent against adriamycin (ADR)-induced cardiotoxicity in isolated rat heart myocytes and mitochondria. In cardiac myocytes, ADR (0.5 mM) caused a significant (70%) inhibition of palmitate oxidation, whereas, PLC (5 mM) induced a significant (49%) stimulation. Addition of PLC to ADR-incubated myocytes induced 79% reversal of ADR-induced inhibition of palmitate oxidation. In isolated rat heart mitochondria, ADR produced concentration-dependent inhibition of both palmitoyl-CoA and palmitoyl-carnitine oxidation, while PLC caused a more than 2.5-fold increase in both substrates. Preincubation of mitochondria with 5 mM PLC caused complete reversal of ADR-induced inhibition in the oxidation of both substrates. Also ADR induced concentration-dependent inhibition of CPT I which is parallel to the inhibition of its substrate palmitoyl-CoA. In rat heart slices, ADR induced a significant (65%) decrease in adenosine triphosphate (ATP) and this effect is reduced to 17% only by PLC. Results of this study revealed that ADR induced its cardiotoxicity by inhibition of CPT I and beta-oxidation of long-chain fatty acids with the consequent depletion of ATP in cardiac tissues, and that PLC can be used as a protective agent against ADR-induced cardiotoxicity.