Phenylalanine 4-monooxygenase: the "sulfoxidation polymorphism".

1. Consistent differences in the proportion of an orally administered dose of S-carboxymethyl-l-cysteine subsequently excreted in the urine as S-oxide metabolites were reported 40 years ago. This observation suggested the existence of inter-individual variation in the ability to undertake the enzymatic S-oxygenation of this compound. Pedigree studies and investigations employing twin pairs indicated a genetically controlled phenomenon overlaid with environmental influences. It was reproducible and not related to gender or age.2. Studies undertaken in several healthy volunteer cohorts always provided similar results that were not significantly different when statistically analysed. However, when compared to these healthy populations, a preponderance of subjects exhibiting the characteristic of poor sulfoxidation of S-carboxymethyl-l-cysteine was found within groups of patients suffering from various disease conditions. The most striking of these associations were witnessed amongst subjects diagnosed with neurodegenerative disorders; although, underlying mechanisms were unknown.3. Exhaustive investigation has identified the enzyme responsible for this S-oxygenation reaction as the tetrahydrobiopterin-dependent aromatic amino acid hydroxylase, phenylalanine 4-monooxygenase classically assigned the sole function of converting phenylalanine to tyrosine. The underlying principle is discussed that enzymes traditionally associated solely with intermediary metabolism may have as yet unrecognised alternative roles in protecting the organism from potential toxic assault.

Effect of CArbocisteine in Prevention of exaceRbation of chronic obstructive pulmonary disease (CAPRI study): An observational study.

BACKGROUND: Chronic Obstructive Pulmonary Disease (COPD) is a chronic and progressive lung disease characterized by irreversible airflow obstruction, airway inflammation, oxidative stress and, often, mucus hypersecretion. The aim of this study is to determine if carbocisteine, a mucolytic and antioxidant agent, administered daily for 12 months, can reduce exacerbation frequency in COPD patients. METHODS: This observational study was conducted in Naples (population approximately 1000,000), Italy. It included 85 out-patients (mean age of 67.8 ± 8.6 years) followed by Clinic of Respiratory Diseases of the University Federico II. Every patient underwent spirometry demonstrating airflow obstruction not fully reversible according to ERS/ATS criteria for COPD diagnosis (Tiffenau index less than 70% after administration of salbutamol, a beta2 agonist drug). Patients enrolled had diagnosed COPD since 2 years and suffered at least one exacerbation in the previous year. None of the patients had been treated with carbocisteine or other mucolytic agent for a longer period of time than 7 days and no more than 4 times in the previous year to the enrollment. All of them assumed daily 2.7 g of carbocisteine lysine salt for a year in addition to their basic therapy. RESULTS: The comparison of exacerbation frequency between the previous year (T0) and the end of study treatment (T12), documents a statistically significant reduction of exacerbations(number of exacerbations at T0: 2 [1,3] vs number of exacerbations at T12: 1 [1,2]; p < 0.001).Quality of life was also reported and showed a statistically significant improvement at the end of the study (p < 0.001).We did not find correlation between reducing exacerbation frequency and exposure to cigarette smoking, passive smoking exposure in childhood, the use of inhaled steroids, the level of education of our patients and the GOLD stadium. INTERPRETATION: Daily administration of a mucolytic drug such as carbocisteine for prolonged periods in addition to the bronchodilator therapy can be considered a good strategy for reducing exacerbation frequency in COPD.

An investigation into possible xenobiotic-endobiotic inter-relationships involving the amino acid analogue drug, S-carboxymethyl-L-cysteine and plasma amino acids in humans.

The amino acid derivative, S-carboxymethyl-L-cysteine, is an anti-oxidant agent extensively employed as adjunctive therapy in the treatment of human pulmonary conditions. A major biotransformation route of this drug, which displays considerable variation in capacity in man, involves the oxidation of the sulfide moiety to the inactive S-oxide metabolite. Previous observations have indicated that fasted plasma L-cysteine concentrations and fasted plasma L-cysteine/free inorganic sulfate ratios were correlated with the degree of sulfoxidation of this drug and that these particular parameters may be used as endobiotic biomarkers for this xenobiotic metabolism. It has been proposed also that the enzyme, cysteine dioxygenase, was responsible for the drug sulfoxidation. Further in this theme, the degree of S-oxidation of S-carboxymethyl-L-cysteine in 100 human volunteers was investigated with respect to it potential correlation with fasted plasma amino acid concentrations. Extensive statistical analyses showed no significant associations or relationships between the degree of drug S-oxidation and fasted plasma amino acid concentrations, especially with respect to the sulfur-containing compounds, methionine, L-cysteine, L-cysteine sulfinic acid, taurine and free inorganic sulfate, also the derived ratios of L-cysteine/L-cysteine sulfinic acid and L-cysteine/free inorganic sulfate. It was concluded that plasma amino acid levels or derived ratios cannot be employed to predict the degree of S-oxidation of S-carboxymethyl-L-cysteine (or vice versa) and that it is doubtful if the enzyme, cysteine dioxygenase, has any involvement in the metabolism of this drug.

A prospective study of the effects of carbocysteine lysine salt on frequency of exacerbations in COPD patients treated with or without inhaled steroids.

OBJECTIVE: COPD is one of the major causes of morbidity and mortality worldwide and represents one of the most important issues for public health. Frequent exacerbations induce a faster decline in lung function and poorer quality of life, increase mortality, and have a socio-economic impact with a high burden in terms of resources and healthcare costs. The clinical trials evaluated the effect of mucolytics in COPD and showed that the long-term carbocysteine, associated with bronchodilators, anticholinergics, and steroids, reduces the frequency of exacerbations and improves the quality of life. PATIENTS AND METHODS: The aim of this prospective real-life study was to evaluate the long-term impact on exacerbations (at 1 year) in COPD patients treated with carbocysteine lysine salt (single dose of 2.7 g once a day) in addition to background therapy with or without inhaled steroids. RESULTS: In a total of 155 evaluable patients, our study showed that the addition of a single dose of carbocysteine lysine salt to background therapy determines a statistically significant reduction of the average number of exacerbations vs. the number observed in the previous year (from 1.97±0.10 to 1.03±0.11; p<0.01), irrespective of treatment with or without inhaled steroids. In particular, in patients with ≥2 exacerbations in the previous year, the addition of carbocysteine lysine salt resulted in a statistically significant reduction in the exacerbations rate from 69% to 33% and from 58% to 25%, respectively (p<0.01) in patients with or without inhaled steroids. CONCLUSIONS: In summary, our data highlighted the efficacy of long-term administration of a single daily dose of carbocysteine lysine salt (2.7 g/day) in reducing the number and rate of exacerbations in COPD patients, independently from the use of inhaled steroids.

S-CMC-Lys protective effects on human respiratory cells during oxidative stress.

The mucoactive drug S-carbocysteine lysine salt monohydrate (S-CMC-Lys) stimulates glutathione (GSH) efflux from respiratory cells. Since GSH is one of the most important redox regulatory mechanisms, the aim of this study was to evaluate the S-CMC-Lys effects on GSH efflux and intracellular concentration during an oxidative stress induced by the hydroxyl radical (xOH). Experiments were performed on cultured human respiratory WI-26VA4 cells by means of patch-clamp experiments in whole-cell configuration and of fluorimetric analyses at confocal microscope. xOH exposure induced an irreversible inhibition of the GSH and chloride currents that was prevented if the cells were incubated with S-CMC-Lys. In this instance, the currents were inhibited by the specific blocker CFTR(inh)-172. CFT1-C2 cells, which lack a functional CFTR channel, were not responsive to S-CMC-Lys, but the stimulatory effect of the drug was restored in LCFSN-infected CFT1 cells, functionally corrected to express CFTR. Fluorimetric measurements performed on the S-CMC-Lys-incubated cells revealed a significant increase of the GSH concentration that was completely hindered after oxidative stress and abolished by CFTR(inh)-172. The cellular content of reactive oxygen species was significantly lower in the S-CMC-Lys-treated cells either before or after xOH exposure. As a conclusion, S-CMC-Lys could exert a protective function during oxidative stress, therefore preventing or reducing the ROS-mediated inflammatory response.

S-CMC-Lys-dependent stimulation of electrogenic glutathione secretion by human respiratory epithelium.

Glutathione (GSH) is one of the most important defense mechanisms against oxidative stress in the respiratory epithelial lining fluid. Considering that GSH secretion in respiratory cells has been postulated to be at least partially electrogenic, and that the mucoregulator S-carbocysteine lysine salt monohydrate (S-CMC-Lys) can cause an activation of epithelial Cl(-) conductance, the purpose of this study was to verify whether S-CMC-Lys is able to stimulate GSH secretion. Experiments have been performed by patch-clamp technique, by high-performance liquid chromatography (HPLC) assay, and by Western blot analysis on cultured lines of human respiratory cells (WI-26VA4 and CFT1-C2). In whole-cell configuration, after cell exposure to 100 microM S-CMC-Lys, a current due to an outward GSH flux was observed, which was inhibitable by 5-nitro-2-(3-phenylpropylamino)-benzoate and glibenclamide. This current was not observed in CFT1-C2 cells, where a functional cystic fibrosis transmembrane conductance regulator (CFTR) is lacking. Inside-out patch-clamp experiments (GSH on the cytoplasm side, Cl(-) on the extracellular side) showed the activity of a channel, which was able to conduct current in both directions: the single channel conductance was 2-4 pS, and the open probability (P(o)) was low and voltage-independent. After preincubation with 100 microM S-CMC-Lys, there was an increase in P(o), in the number of active channels present in each patch, and in the relative permeability to GSH vs Cl(-). Outwardly directed efflux of GSH could also be increased by protein kinase A, adenosine 5'-triphosphate, and cyclic adenosine monophosphate (cAMP) added to the cytoplasmic side (whole-cell configuration). The increased secretion of GSH observed in the presence of S-CMC-Lys or 8-bromoadenosine-3',5'-cyclic monophosphate was also confirmed by HPLC assay of GSH on a confluent monolayer of respiratory cells. Western blot analysis confirmed the presence of CFTR in WI-26VA4 cells. This study suggests that S-CMC-Lys is able to stimulate a channel-mediated GSH secretion by human respiratory cells: electrophysiological and pharmacological characteristics of this channel are similar to those of the CFTR channel.

Disposition and metabolism in 1-(2-chloroethyl)-3-(2',3',4'-tri-o-acetyl, ribopyranosyl)-1-nitrosourea in rats.

The antineoplastic activity in animals of 1-(2-chloroethyl)-3-(2',3',4'-tri-O-acetyl, ribopyranosyl)-1-nitrosourea (RPCNU) has been widely demonstrated. The present study deals with the disposition and the metabolism of three 14C-labeled species of RPCNU. The chemical plasma half-life of the drug was less than 5 min. Within the first min after injections, most of the radioactivity derived from ethyl-14C groups were recovered as volatile products. Among these, 2-chloroethanol was identified as a main component. Analysis of labeled species in urine after administration of [ethyl-14C]RPCNU showed that thiodiacetic acid and its sulfoxide were major metabolites of RPCNU (62% of the urinary radioactivity). Traces of N-acetylcarboxymethyl- and N-acetylhydroxyethylcysteine) were also detected. The only labeled species concentrating in particular tissues was that carrying the chloroethyl moiety. Uptake to high levels of [ethyl-14C]RPCNU did occur in liver, kidney, pancreas, thymus, and Harder's gland.

Phenylalanine 4-monooxygenase and the S-oxidation of S-carboxymethyl-L-cysteine in HepG2 cells.

The role of phenylalanine 4-monooxygenase (PAH) in the S-oxidation of S-carboxymethyl-L-cysteine (SCMC) in the rat has now been well established in rat cytosolic fractions in vitro. However, the role of PAH in the S-oxidation of SCMC in human cytosolic fractions or hepatocytes has yet to be investigated. The aim of this investigation was to analyse the kinetic parameters of PAH oxidation of both L-phenylalanine (Phe) and SCMC in the human HepG2 cell line in order to investigate the use of these cells as a model for the cellular regulation of SCMC S-oxidation. The experimentally determined Km and V(max) were 7.14 +/- 0.32 mM and 0.85 +/- 0.32 nmole Tyr formed min(-1) x mg protein(-1) using Phe as substrate. For SCMC the values were 25.24 +/- 5.91 mM and 0.79 +/- 0.09 nmole SCMC (RIS) S-oxides formed min(-1) x mg protein(-1). The experimentally determined Km and V(max) for the cofactor BH4 were 6.81 +/- 0.21 microM and 0.41 +/- 0.004 nmole Tyr formed min(-1) x mg protein(-1) for Phe and 7.24 +/- 0.19 microM and 0.42 +/- 0.002 nmole SCMC (R/S) S-oxides formed min(-1) x mg protein(-1) for SCMC. The use of various PAH inhibitors confirmed that HepG2 cells contained PAH and that the enzyme was capable of converting SCMC to its (R) and (S) S-oxide metabolites in an in vitro PAH assay. Thus HepG2 cells have become a useful additional tool for the investigation of the cellular regulation of PAH in the S-oxidation of SCMC.

Development and validation of a stability indicating method for S-carboxymethyl-L-cysteine and related degradation products in oral syrup formulation.

A stability-indicating method for the determination of S-carboxymethyl-L-cysteine and related degradation impurities in Exputex® 250mg/5mL syrup was developed in anion-exchange liquid chromatography mode. A forced degradation study supported the method development to ensure stability indicating conditions. Aqueous solutions of the active pharmaceutical ingredient and syrup samples at different pH-values were stress-tested in different thermal, light exposure and headspace conditions. One degradation product was detected in thermal stress studies at 60°C and 80°C in the pH range 5.0-7.0 and was identified by mass spectrometry as 5-oxo-thiomorpholine-3-carboxylic acid (lactam of carbocysteine). A second degradation product was only generated in moderately strong oxidizing conditions (0.5% H2O2 aqueous solution) and was identified as S-carboxymethyl-L-cysteine-(R/S)-sulphoxide (carbocysteine sulphoxide). The method was developed on a Zorbax SAX column, in isocratic mode. The mobile phase consisted of 200mM phosphate solution at pH 4.0 and acetonitrile (50:50 v/v) and UV detection was performed at a wavelength of 205nm. The method was linear for carbocysteine (R>0.9982) over a concentration range of 2.5-50µg/mL and 0.4-0.6mg/mL. Linearity for the impurities was shown from the LOQ to 50µg/mL. Specificity was verified and accuracy demonstrated for the active ingredient and its degradation products in syrup samples at 3 levels around their respective specification limits. Repeatability, intermediate precision and inter-laboratory reproducibility were assessed on three commercial batches, analyzed in triplicate by two operators at both the transferring and the receiving site and demonstrated a successful method transfer to the manufacturing quality control laboratory.

Re-evaluation of the metabolism of carbocisteine in a British white population.

It has been claimed that the amino acid derivative carbocisteine is predominantly metabolized by sulfoxidation and that this pathway exhibits a genetic polymorphism. Moreover, those subjects with a 'poor metabolizer' phenotype have been thought to have a genetic predisposition to developing certain diseases. We have confirmed the observations of others that this marker drug does not undergo significant S-oxidation. Furthermore, a novel urinary metabolite, S-(carboxymethylthio)-L-cysteine (CMTC) has recently been identified. To determine if a genetic polymorphism for this biotransformation pathway exists, metabolic ratios (% urinary excretion carbocisteine/% urinary excretion CMTC) for 120 healthy volunteers were assessed using high-performance thin-layer chromatography. Urinary excretion of the parent drug ranged from 6% of the dose administered to 56% (mean +/- SD, 23.4 +/- 0.8%). No cysteinyl sulfoxide metabolites were identified in the urine samples. The amount excreted as CMTC exhibited a 12-fold variation but only accounted for mean of 4.4% (1-12%) of the dose given. Two individuals initially had high metabolic ratios (> 30), however, on rechallenge both their MRs were less than 5. Therefore, carbocisteine is not an appropriate probe drug for sulfoxidation. The formation of the novel metabolite CMTC appears to exhibit polymorphism, although the considerable intra-subject variation for its formation does not allow assignment of a phenotype.

S-carbocysteine-lysine salt monohydrate and cAMP cause non-additive activation of the cystic fibrosis transmembrane regulator channel in human respiratory epithelium.

S-Carbocysteine-lysine salt monohydrate (S-CMC-Lys) has been shown to open a Cl- channel in the trachea, thus aiding fluid secretion. The aim of this study was to characterize the channel and the action mechanism on a culture line of human respiratory epithelial cells. The patch-clamp technique (in cell-attached or inside-out configuration) and conventional micro-electrodes were used. The activity and density of a cAMP-dependent Cl- channel, identical to the cystic fibrosis transmembrane regulator (CFTR) channel, proved to be maximally stimulated by 100 microM S-CMC-Lys present in the cAMP-free cell incubation medium for 240-290 min (cell-attached configuration). Subsequent addition of cAMP to the medium did not determine any further activation. S-CMC-Lys acted mostly indirectly as, when placed in direct contact with a membrane patch, activation of the CFTR channel was nil (cytoplasmic side) or limited (external side).

Pharmacokinetic behavior of S-carboxymethylcysteine-Lys in patients with chronic bronchitis.

A mass fragmentographic technique was used to study the pharmacokinetic behavior of SCMC-Lys in patients with acute exacerbations of chronic bronchitis and with dense expectoration. Serum and urine levels, as well as bronchial mucus levels and their correlations, were determined. The data suggest that SCMC-Lys diffuses well into bronchial mucus, a useful feature for a mucolytic drug.

Exhaled Interleukine-6 and 8-isoprostane in chronic obstructive pulmonary disease: effect of carbocysteine lysine salt monohydrate (SCMC-Lys).

Chronic obstructive pulmonary disease (COPD) is characterized by an airways inflammation and by an enhanced generation of reactive oxygen species. The aim of our study was to assess the inflammation and the oxidative stress in airways of COPD patients with acute exacerbation of disease and in stability. Furthermore, we investigated the anti-inflammatory and antioxidant effects of 6 months treatment with carbocysteine lysine salt monohydrate (SCMC-Lys) in COPD. We studied 30 mild acute COPD, 10 mild stable COPD and 15 healthy subjects. 8-isoprostane and Interleukine-6 were measured in their breath condensate through immunoassay. Significantly higher concentrations of exhaled 8-isoprostane and Interleukine-6 were found in acute COPD patients compared to stable COPD and healthy controls (21.8+/-5.1 vs. 13.2+/-2.0 vs. 4.7+/-1.8 pg/ml and 7.4+/-0.9 vs. 5.8+/-0.2 vs. 2.7+/-0.6 pg/ml, p<0.0001). COPD patients treated with SCMC-Lys showed a marked reduction of exhaled 8-isoprostane and Interleukine-6 (8.9+/-1.5 and 4.6+/-0.8 pg/ml, p<0.0001). These findings suggest that there is an increase of 8-isoprostane and Interleukine-6 concentrations in the breath condensate of COPD patients compared to healthy controls especially during acute exacerbations of the disease. Moreover, we showed an anti-inflammatory and antioxidant effect of short-term administration of SCMC-Lys in COPD, suggesting the importance of a further placebo-controlled study that should evaluate the effects of this drug.

Carbocysteine lysine salt monohydrate (SCMC-LYS) is a selective scavenger of reactive oxygen intermediates (ROIs).

Carbocysteine lysine salt monohydrate (SCMC-Lys) is a well-known mucoactive drug whose therapeutic efficacy is commonly related to the ability of SCMC-Lys to replace fucomucins by sialomucins. The aim of this study was to determine if SCMC-Lys could exert an anti-oxidant action by scavenging reactive oxygen intermediates (ROIs). Our results show that SCMC-Lys proved effective as a selective scavenger of hypochlorous acid (HOCl) and hydroxyl radical (OH.), this effect being related to the reactivity of the SCMC tioether group. The scavenger activity of SCMC-Lys was observed in free cellular system as well as in activated human polymorphonuclear neutrophils (PMNs). SCMC-Lys scavenger activity on HOCl was paralleled by a powerful protection from HOCl-mediated inactivation of alpha1-antitripsin (alpha1-AT) inhibitor, the main serum protease inhibitor. Production of interleukin-(IL-)8, a major mediator of PMN recruitment in inflammatory diseases, is known to be mediated by intracellular OH. SCMC-Lys significantly reduced IL-8 production on stimulated human peripheral blood mononuclear cells (PBMCs) in the same range of concentrations affecting OH. activity. It is concluded that SCMC-Lys could exert, in addition to its mucoactive capacity, an anti-oxidant action, thus contributing to the therapeutic efficacy of SCMC-Lys.

Phenotypic variation in xenobiotic metabolism and adverse environmental response: focus on sulfur-dependent detoxification pathways.

Proper bodily response to environmental toxicants presumably requires proper function of the xenobiotic (foreign chemical) detoxification pathways. Links between phenotypic variations in xenobiotic metabolism and adverse environmental response have long been sought. Metabolism of the drug S-carboxymethyl-L-cysteine (SCMC) is polymorphous in the population, having a bimodal distribution of metabolites, 2.5% of the general population are thought to be nonmetabolizers. The researchers developing this data feel this implies a polymorphism in sulfoxidation of the amino acid cysteine to sulfate. While this interpretation is somewhat controversial, these metabolic differences reflected may have significant effects. Additionally, a significant number of individuals with environmental intolerance or chronic disease have impaired sulfation of phenolic xenobiotics. This impairment is demonstrated with the probe drug acetaminophen and is presumably due to starvation of the sulfotransferases for sulfate substrate. Reduced metabolism of SCMC has been found with increased frequency in individuals with several degenerative neurological and immunological conditions and drug intolerances, including Alzheimer's disease, Parkinson's disease, motor neuron disease, rheumatoid arthritis, and delayed food sensitivity. Impaired sulfation has been found in many of these conditions, and preliminary data suggests that it may be important in multiple chemical sensitivities and diet responsive autism. In addition, impaired sulfation may be relevant to intolerance of phenol, tyramine, and phenylic food constituents, and it may be a factor in the success of the Feingold diet. These studies indicate the need for the development of genetic and functional tests of xenobiotic metabolism as tools for further research in epidemiology and risk assessment.

Antioxidant activity of carbocysteine lysine salt monohydrate.

BACKGROUND: Reactive oxygen radicals are involved in many respiratory diseases, including chronic obstructive pulmonary disease (COPD). Carbocysteine lysine salt monohydrate (CLS) is a mucoactive drug effective in the treatment of bronchopulmonary diseases characterized by mucus alterations, including COPD. In the present study, the antioxidant activity of CLS was studied in vitro in three different oxygen radical producing systems, i.e. bronchoalveolar lavages (BAL) from patients affected by COPD, ultrasound treated human serum and cultured human lung endothelial cells challenged with elastase. METHODS: BAL, exposed or not to different concentrations of CLS (1.5-30 mM), was assayed for free radical content by fluorometric analysis of DNA unwinding (FADU) or by cytochrome c reduction kinetics. Human serum was treated with ultrasound in the presence or absence of CLS (1.5, 2.5 mM) or N-acetyl cysteine (NAC; 4, 5 mM) and assayed for free radical content by FADU. Human endothelial cells cultured in vitro from pulmonary artery were incubated with elastase (0.3 IU/mL), in the presence or absence of glutathione (GSH; 0.65 mM) or CLS (0.16 mM). The supernatant was tested for cytochrome c reduction kinetics whereas cell homogenates were assessed for xanthine oxidase (XO) content by SDS-PAGE. RESULTS: Results showed that CLS is more effective as an in vitro scavenger in comparison to GSH and NAC. CLS reduced the damage of DNA from healthy donors exposed to COPD-BAL and was able to quench clastogenic activity induced in human serum by exposure to ultrasound at concentrations as low as 2.5 mM. NAC protect DNA from radical damage, starting from 5 mM. In human lung endothelial cells cultured in presence of elastase, CLS (0.16 mM) decreased xanthine oxidase activity. CONCLUSIONS: These results suggest that CLS could act by interfering with the conversion of xanthine dehydrogenase into superoxide-producing xanthine oxidase. The antioxidant activity of CLS could contribute to its therapeutic activity by reducing radical damage to different lung structures.

Synthesis and chromatographic properties of S-(1-carboxyethyl)-L-cysteine and S-(1-carboxypropyl)-L-cysteine.

Details are reported for the synthesis of S-(1-carboxyethyl)-L-cysteine (1-CEC) and S-(1-carboxypropyl)-L-cysteine (1-CPC) from cysteine and 2-bromopropionic acid or 2-bromobutyric acid, respectively. Some analytical data and the behaviour of these two compounds on paper and ion-exchange chromatography are also reported, which allow their identification.

Carboxymethyl-selenopyruvic acid as the product of the oxidative deamination of carboxymethyl-selenocysteine.

CMSeC labeled with 14C in the carboxymethyl moiety was incubated with snake venom L-aminoacid oxidase. As the product of the reaction only one ketoacid was detected, which retained all the radioactivity of the oxidized substrate. This clearly shows that no breakdown of the C-Se bond of CMSeC occurs during its oxidation, and confirms previously reported data indicating that the ketoacid arising from CMSeC is CMSeP.

Oxidative deamination of S-(1-carboxyethyl)-L-cysteine and S-(1-carboxypropyl)-L-cysteine by L-aminoacid oxidase.

S-(1-carboxyethyl)-L-cysteine (1-CEC) and S-(1-carboxypropyl)-L-cysteine (1-CPC) are oxidatively deaminated by L-aminoacid oxidase with consumption of half a mole of oxygen per mole of substrate in the presence of catalase. This reaction gives rise to the corresponding alpha-ketoacids, identified by some chemical and chromatographic tests and by comparison with synthetic compounds. It has been possible, therefore, to demonstrate that S-(1-carboxyethyl)-thiopvruvic acid (1-CETP) and S-(1-carboxypropyl)-thiopvruvic acid (1-CPTP) are the main products of oxidative deamination of 1-CEC and 1-CPC.

Evaluation of the elastic and viscous components of bronchial mucus before and after S-carboxymethylcysteine-Lys treatment.

The rheological profile of bronchial secretions from patients suffering from chronic bronchitis has been evaluated using the forced sinusoidal oscillations technique, which allows dynamic tests to be carried out. In these tests the shear stress can be plotted versus the strain on a XY plotter, to obtain Lissajous figures. As bronchial mucus is a viscoelastic fluid, elliptic figures were obtained that can be broken down into an elastic component separate from the viscous component. By applying this method to the study of the S-carboxymethylcysteine action on bronchial mucus, it is possible to observe that the improvement in the rheological characteristics of the mucus, after treatment, is mainly due to a decrease in viscosity rather than to a decrease in elasticity.