Hyperventilation syndrome and reflexologic methods of its correction]

Acupuncture has been studied for possibility to be used for correction of hyperventilation disorders. Diagnostics of hyperventilation syndrome (HVS) was carried out on the basis of data of clinical, laboratory and functional methods. Acupuncture was carried out using the first (strong) variant of classical inhibitive procedure of acupuncture. It was shown that reflex therapy decreased electroencephalographic characters of dysrhythmia and exaltation and paroxysmal activity as well as inter-hemisphere asymmetry in patients with HVS. Parallel with a decrease in the degree of negative subjective sensations, minute respiratory volume, non-informity of regional ventilation were observed to decrease and partial oxygen pressure in alveolar air--to increase. The positive results of reflex therapy of HVS indicate that it is possible to correct hyperventilation disorders using no pharmacotherapy.     

Improved production of (R)-2-octanol via asymmetric reduction of 2-octanone with Oenococcus oeni CECT4730 in a biphasic system

Abstract

Oenococcus oeni CECT4730, which catalyses the asymmetric reduction of 2-octanone to (R)-2-octanol with high enantioselectivity, was further studied to exploit its potential for production of (R)-2-octanol in an aqueous/organic solvent biphasic system. Variables such as the volume ratio of aqueous to organic phase (V_a/V_o), buffer pH, reaction temperature, shaking speed, co-substrates and the ratio of biocatalyst to substrate were examined with respect to the molar conversion, the initial reaction rate and the product enantiomeric excess (e.e.). Under the optimized conditions (V_a/V_o=1:1 (v/v), buffer pH=8.0, reaction temperature=30°C, shaking speed=150 rev/min, ratio of glucose to biomass=5.4:l (w/w), ratio of biocatalyst to substrate=0.51:l (g/mol)), the highest space time yield of (R)-2-octanol, 24 mmol L^?1 per h, and >98% product e.e. were obtained at a substrate concentration close to 1.0 mol L^?1 after 24 h reduction.     

Cardiac output variations in supine resting subjects during head-out cold water immersion

Five men, aged 31.2 years (SD 2.3), under semi-nude conditions and resting in a dorsal reclining position, were exposed to thermoneutral air for 30 min, followed immediately by a cold water (15°C) immersion for 60 min. Cardiac output was measured using a dualbeam Doppler flow meter. During immersion in cold water, cardiac frequency (f _c) showed an initial bradycardia. The lowest values were reached at about 10 min after immersion, 58.3 (SD 2.5) to 48.3 (SD 7.8) beats min^–1 (P < 0.05). By the 20th min of exposure,f _c had gradually risen to 70.0 beats min^–1 (SD 6.6,P < 0.05). This change could be due to the inhibition of the initial vagal reflex by increased catecholamine concentration. Stroke volume (V _s) was significantly increased (P < 0.05) during the whole cold immersion period. Cardiac output, increased from 3.57 (SD 0.50) to 6.26 (SD 1.33)1 min^–1 (P < 0.05) and its change with time was a function of bothV _s andf _c. On the other hand, systolic flow acceleration was unchanged during the period of immersion. The changes in the respiratory variables (ventilation, oxygen uptake, carbon dioxide output and respiratory exchange ratio) during immersion showed an initial hyperventilation followed, as immersion proceeded, by a slower metabolic increase due to shivering.     

Unusual Slow Gating of Gap Junction Channels in Oocytes Expressing Connexin32 or Its COOH-Terminus Truncated Mutant

Gap junction channels are gated by a chemical gate and two transjunctional voltage (V _j)-sensitive gates: fast and slow. Slow V _j gate and chemical gate are believed to be the same. The slow gate closes at the negative side of V _j and is mostly inactive without uncouplers or connexin (Cx) mutations. In contrast, our present data indicate otherwise. Oocytes expressing Cx32 were subjected to series of −100 mV V _j pulses (12-s duration, 30-s intervals). Both peak (PK) and steady-state (SS) junctional conductances (G _j), measured at each pulse, decreased exponentially by 50−60% (tau = ∼1.2 min). G _jPK dropped more dramatically, such that G _jSS/G _jPK increased from 0.4 to 0.6, indicating a drop in V _j sensitivity. Less striking effects were obtained with –60 mV pulses. During recovery, G _j, measured by applying 20 mV pulses (2-s duration, 30-s intervals), slowly returned to initial values (tau = ∼7 min). With reversal of V _j polarity, G _jPK briefly increased and G _jSS/G _jPK decreased, suggesting that V _j-dependent hemichannel reopening is faster than hemichannel closing. Similar yet more dramatic results were obtained with COOH-terminus truncated Cx32 (Cx32-D225), a mutant believed to lack fast V _j gating. The data indicate that the slow gate of Cx32 is active in the absence of uncouplers or mutations and displays unusual V _j behavior. Based on previous evidence for direct calmodulin (CaM) involvement in chemical/slow gating, this may also be CaM-mediated.     

The kinetics of the active and de-energized transport of O-methyl glucose in Ustilago maydis

The kinetics of the uptake and efflux of 3-O-methyl-glucose in sporidia of Ustilago maydis were measured, both in active cells and in cells whose metabolic activity had been inhibited by azide and iodoacetate.The de-energized transport system proved to be carrier mediated with apparent affinity constants 13 ± 2 mM outside (K₀) and 18 ± 2 mM inside (K₁). The apparent maximum rate constants for the same system were 0.66 ± 0.05 mmol/l cell water per min for uptake (V₊ and 0.53 ± 0.04 mmol/l cell water per min for efflux (V_-. For the active system K₀ = 0.08 ± 0.01, K₁ > 40, V₊ = 9.7 ± 0.5 and V₋ 1.1 ± 0.9 (in equivalent units). These results are discussed in the context of the carrier mechanism as proposed by Regen and Morgan (Regen, D.M. and Morgan, H.E. (1964) Biochim. Biophys. Acta 79, 151–166).The antifungal compound carboxin had no effect on de-energized transport but was shown to decrease both K₀ and V₊ in the active system. Phloretin and phlorizin were also found to be without effect on de-energized cells but the former enhanced while the latter inhibited active uptake.     

Force-Velocity Relationship of Shortening of Blood Vessel of the Common Frog Rana temporaria during Phasic and Tonic Contractions

The relationship between shortening velocity and the applied external load (force-velocity) was studied for vena cava posterior of the frog Rana temporaria. This relationship was determined in 15 s, 2 and 20 min after the beginning of isometric contraction induced by a high KCl concentration (110 mmole/l). It was found that maximal shortening velocity (V ₀) was the highest at the 15th s after the beginning of the isometric contraction and amounted to 0.125 ± 0.009 L ₀/s (n = 4); then it decreased progressively to 0.058 ± 0.005 L ₀/s at the 20th min, respectively. On the contrary, the isometric tension value increased and reached its peak in 5 min after the beginning of contraction, then it decreased slightly. Such phenomenon indicates the occurrence of attached non-cycling bridges. The a/P ₀ parameter was equal to 0.15 ± 0.03 and 0.33 ± 0.06 at the 15th s and the 20th min, respectively.     

Purification,Characterization and Antibacterial Activity of l‐amino Acid Oxidase from Cerastes cerastes

Antibiotic resistance presents a real problem in which new antibacterial molecules from natural secretions could be beneficial in the development of new drugs. In this study, Cerastes cerastes venom was investigated for its antibacterial activity against Gram‐positive and Gram‐negative bacteria. The antibacterial activity was evaluated by measuring the halo inhibition and minimum inhibitory concentration (MIC). An l ‐amino acid oxidase (CcLAAO) was purified from this venom using three chromatographic steps; its homogeneity (60 kDa) was confirmed by SDS‐PAGE. LC–MS/MS analysis of CcLAAO showed similarities with other LAAO enzymes from Echis ocellatus and Viridovipera stejnegeri venoms. CcLAAO presents an antibacterial activity against three bacterial strains (Staphylococcus aureus, Methicillin‐resistant S. aureus, and Pseudomonas aeruginosa) with MIC values of 10, 10, and 20 μg/mL, respectively. However, no effect was observed against Escherichia coli and yeast strains. Kinetic parameters of CcLAAO evaluated on l ‐leucine at pH 8.0 and 20°C were K_m = 0.06 mmol and V_max = 164 mmol/min.     

GSM phone signal does not produce subjective symptoms

The influence of pulsed radiofrequency (RF) electromagnetic fields of digital GSM mobile phones (902 MHz, 217 Hz pulse modulation) on subjective symptoms or sensations in healthy subjects were studied in two single-blind experiments. The duration of the RF exposure was about 60 min in Experiment 1 and 30 min in Experiment 2. Each subject rated symptoms or sensations in the beginning of the experimental session and at the end of both the exposure and the nonexposure conditions. The symptoms rated were headache, dizziness, fatigue, itching or tingling of the skin, redness on the skin, and sensations of warmth on the skin. The results did not reveal any differences between exposure and non-exposure conditions, suggesting that a 30-60 min exposure to this RF field does not produce subjective symptoms in humans.     

Calorimetric Glucose Determination by Glucose Oxidase —Methylene Blue

The reaction between glucose and methylene blue, catalyzed by glucose oxidase (GOD)was analysed calorimetrically. The amount of heat produced under saturating methylene blue concentrations ( > 10^?2 mol/1)was measured with glucose concentration and time as parameters (kinetic procedure) Kinetic constants (pseudo one substrate kinetics) were derived from the experimental data: K_M(glucose)= 1.18 × 10^?3 mol/l and V_max = 0.085 J/mg GOD min (3.89 · 10^?6 mol/mg GOD min) Comparison of caloric with optical measurements gave an enthalpy of reaction of 22.52 kJ/mol. Considering the observed substrate inhibition, glucose determinations are possible up to glucose concentrations of 0.1 mol/l.     

Production and characterization of polygalacturonase fromGeotrichum candidum

An extracellular polygalacturonase (EC 3.2.1.15) fromGeotrichum candidum ATCC 34614 grown onsauerkraut brine was produced and characterized. Polygalacturonic acid markedly increased the enzyme yield in the brine. The fungus produced the highest activity (290 U/l) in brine with 0.3% (w/v) polygalacturonic acid. The pH and temperature optima of the enzymes were 4.5 to 5.0 and 30°C, respectively. It was stable from pH 4.0 to 5.8 and at 30°C but lost its activity at higher temperatures. The K_m and V_max values for polygalacturonic acid were 4.2 mg/ml and 0.19mm galacturonic acid/min, respectively. The enzyme was not substrate inhibited.     

Effect of Somatostatin Analog on Postprandial Satiation in Obesity

Objective: Altered satiation may impact postprandial symptoms and potentially change food intake in obesity. Our aim was to compare effects of octreotide and placebo on postprandial symptoms, satiation, and gastric volumes in obesity. Research Methods and Procedures: In a randomized, parallel‐group, double‐blind, placebo‐controlled study, 26 obese but otherwise healthy participants received 100 μg of octreotide or placebo subcutaneously 30 minutes before each study. Studies were performed on 2 separate days and included validated non‐invasive techniques: ^99mTc‐single photon emission computed tomography imaging to measure fasting stomach volume and gastric volume changes after 90 mL of water and 240 mL of Ensure and a standardized nutrient drink test to measure the maximum tolerated volume and postprandial symptoms. Results: Relative to placebo, octreotide increased gastric volume after 90 mL of water; however, fasting and gastric volume change post‐Ensure and maximum tolerated volume of Ensure were not different. Octreotide decreased sensations of fullness (p = 0.035) and bloating (p = 0.05) and tended to reduce aggregate symptoms (p = 0.07) after the fully satiating meal. Discussion: In obese individuals, somatostatin analog significantly reduced postprandial sensations after a satiating meal without altering maximum tolerated meal volume or postnutrient gastric volume, suggesting an effect on upper gut sensation. The role of somatostatin as a permissive factor in the development of obesity by reducing postprandial sensations deserves further study.     

Kinetic characterization of unspecific alkaline phosphatase at different villus sites of rat jejunum

Summary A quantitative histochemical method to determine the apparent Km and V _max values of rat intestinal unspecific alkaline phosphatase at different sites of the villi is described. Naphthol-As-Bi-phosphate (0.025–1.5 mM) is employed as substrate and Fast Blue B as coupling reagent, and the resulting azo-dye in the brush border membrane has an absorbance maximum at 550 nm. The ratio between the absorbance at 550 and 500 nm is constant as calculated from automatically recorded spectra at different intense dye deposits. Its absorbance is a linear function of incubation time up to 3 min and thickness of the slices up to 10 m both with medium (0.5 mM) and high (1.5 mM) substrate concentrations. Using the histochemical assay under comparable conditions in test tube experiments with homogenates of intestinal mucosa an app. Km of 0.26±0.081 mM (weighted regression analysis) and 0.28–0.084 mM (direct linear plotting) is determined, demonstrating an affinity to the histochemical substrate, which is about 10 times higher than for p-Nitro-phenyl-phosphate with the purified enzyme.The results obtained by scanning the total dye deposits along jejunal villi show considerable differences in enzymatic activity between single villi and an increase from the villus base up to the transition between medium and apical villus third. As well in the apical region as at the villus base saturation curves are obtained by determining the relationship between the absorbance and the substrate concentration under standard conditions (slice thickness 10 m, incubation time 3 min, 37°C, pH 8.3). Calculated by weighted regression analysis and direct linear plotting from the absorbance data of six female rats the medium app. kinetic data ±SD from the jejunal villi read as follows. Apical: Km=0.81±0.438 mM, V _max=3.99±1.217 absorbance units (A) and Km=0.87±0.428 mM, V _max=4.02±1.191 A, respectively. Basal: Km=0.82±0.261 mM, V _max=3.26±0.719 A and Km=0.77±0.184 mM, V _max=3.04±0.518 AU, respectively. As demonstrated by factorial analysis of variance only V _max is influenced by the villus position.Supported by the Deutsche Forschungsgemeinschaft GU 184/1     

Prediction of Pharmacokinetic Profile of Valsartan in Humans Based on in vitro Uptake‐Transport Data

The aim of this study was to evaluate a physiologically based pharmacokinetic (PBPK) model for predicting PK profiles in humans based on a model refined in rats and humans in vitro uptake‐transport data using valsartan as a probe substrate. Valsartan is eliminated unchanged, mostly through biliary excretion, both in humans and rats. It was, therefore, chosen as model compound to predict in vivo elimination based on in vitro hepatic uptake‐transport data using a fully mechanistic PBPK model. Plated rat and human hepatocytes, and cell lines overexpressing human OATP1B1 and OATP1B3 were used for in vitro uptake experiments. A mechanistic two‐compartment model was used to derive the active and passive transport parameters, namely uptake Michaelis–Menten parameters (V_max and K_m,u) together with passive diffusion (P_dif). These transport parameters were then used as input in a whole body physiologically based pharmacokinetic (PBPK) model. The uptake rate of valsartan was higher for rat hepatocytes (K_m,u=28.4±3.7 μM , V_max=1320±180 pmol/mg/min, and P_dif =1.21±0.42 μl/mg/min) compared to human hepatocytes (K_m,u=44.4±14.6 μM , V_max=304±85 pmol/mg/min, and P_dif=0.724±0.271 μl/mg/min). OATP1B1 and ‐1B3 parameters were correlated to human hepatocyte data, using experimentally established relative activity factors (RAF). Resulting PBPK simulations were compared for plasma‐ (humans and rats) and bile‐ (rats) concentration–time profiles following iv bolus administration of valsartan. Plasma clearances (CL_P) for rats and humans were predicted within twofold relative to predictions based on respective in vitro data. The simulations were extended to simulate the impact of either OATP1B1 or ‐1B3 inhibition on plasma profile. The limited data set indicates that the mechanistic model allowed for accurate evaluation of in vitro transport data; and the resulting hepatic uptake transport kinetic parameters enabled the prediction of in vivo PK profiles and plasma clearances, using PBPK modelling. Moreover, the interspecies difference in elimination rate observed in vivo was correctly reflected in the transport parameters determined in vitro.     

Effects of carbon dioxide on the spatially separate electrogenic ion pumps and the growth rate in the hypocotyl ofVigna sesquipedalis

Carbon dioxide, introduced into the gas phase of the experimental chamber, has distinct effects on two spatially separate membrane potentials and the rate of elongation growth in hypocotyl segments ofVigna sesquipedalis Wight. Both membrane potentials (V _ps andV _px=the electric potential difference between the parenchyma symplast and the surface of the hypocotyl, and that between the parenchyma symplast and the xylem, respectively) hyperpolarized rapidly but transiently at the introduction of CO₂. Prolonged exposure of the hypocotyl to high concentrations of CO₂ (above 10%) caused depolarization of membrane potentials above the level before CO₂ introduction. When CO₂ was replaced with air, the membrane potentials exhibited a distinct depolarization response of transient nature. The growth rate of the hypocotyl segments exhibited similar responses to CO₂ as did the membrane potentials (the increase and the decrease of the growth rate were corresponded to the hyperpolarization and the depolarization, respectively), but these responses always followed the changes of the membrane potentials. The CO₂-induced maximum hyperpolarization ofV _ps and the maximum increase of the growth rate were closely correlated. All these responses were strictly dependent on aerobic metabolism. These results indicate that CO₂ may regulate elongation growth in two ways: by affecting the activity of the electrogenic ion pump via intracellular acidification, and also by acting via apoplastic acidification as a wall-loosening acid.Symbols and abbreviations V _sx electric potential difference between the surface (S) and the xylem (X) of the hypocotyl - V _px electric potential difference between the inside of a parenchyma cell (P) andX - V _ps electric potential difference betweenP andS - V _ps (CO₂, max) the maximum value of CO₂-induced hyperpolarization ofV _ps - GR(CO₂, max) the maximum value of CO₂-induced increase of the growth rate - IAA indole-3-acetic acid     

d-glucose uptake in human liver cell cultures

Summary The kinetic parameters ford-glucose uptake were studied in human liver cell cultures under strictly defined experimental conditions. Using a wide concentration range (0.005 to 30 mmol/l), the kinetic data obtained suggested strongly thatd-glucose in human liver cell cultures can be transported by two separate systems. For the high-affinity system, the apparentK _m was 0.645±0.21 mmol/l and the V_max, 12.49±3.74 nmol/mg protein per min. For the low-affinity system, the apparentK _m was 6.91±0.58 mmol/l and the V_max, 79.90±5.27 nmol/mg protein per min. At a concentration of 2.1×10⁻⁷ mol/l, cytochalasin B preferentially inhibited the high-affinityd-glucose site or transport system. The time course ofd-glucose uptake, studied in two cell lines from patients with hereditary fructose intolerance, was significantly higher than for the control lines. This work was supported by Grant I.N.S.E.R.M. CRL 77-5-210-4.     

Prolonged post-hyperventilation apnea in two young adults with hyperventilation syndrome

Background

The prognosis of hyperventilation syndrome (HVS) is generally good. However, it is important to proceed with care when treating HVS because cases of death following hyperventilation have been reported. This paper was done to demonstrate the clinical risk of post-hyperventilation apnea (PHA) in patients with HVS.

Case presentation

We treated two patients with HVS who suffered from PHA. The first, a 21-year-old woman, had a maximum duration of PHA of about 3.5?minutes and an oxygen saturation (SpO₂) level of 60%. The second patient, a 22-year-old woman, had a maximum duration of PHA of about 3?minutes and an SpO₂ level of 66%. Both patients had loss of consciousness and cyanosis. Because there is no widely accepted regimen for treating patients with prolonged PHA related to HVS, we administered artificial ventilation to both patients using a bag mask and both recovered without any after effects.

Conclusion

These cases show that some patients with HVS develop prolonged PHA or severe hypoxia, which has been shown to lead to death in some cases. Proper treatment must be given to patients with HVS who develop PHA to protect against this possibility. If prolonged PHA or severe hypoxemia arises, respiratory assistance using a bag mask must be done immediately.

     

Hemodynamics in the Microcirculation System of the Rat Cerebral Cortex in Nitrite Methemoglobinemia

Effects of sodium nitrite on the blood flow rate (V) in microvessels of rat cerebral cortex (5–15 µm in diameter), arterial pressure (AP), plasma osmotic pressure (P _osm), and hematological parameters were studied. The mean V and AP in 15 min after administration of NaNO₂ (3 mg/100 g mass) decrease by 36% and 45%, while in 30 and 45 min they increase by 14% and 7% and by 20% and 19% as compared with minimal values, respectively. At the end of the experiment, on the background of an elevation of P _osm (3%), a decrease of total hemoglobin concentration and of the number of erythrocytes was accompanied by a decrease of the discocyte number from 81 to 35% and a reduction of cell diameters by 7%. It is concluded that NaNO₂ produces block of local active mechanisms of the cerebral blood flow regulation. Compensatory responses in this case are provided by passive mechanisms (rheological blood properties) and systemic reactions.     

Redistribution of hepatocyte chloride during l-alanine uptake

We used ion-sensitive, double-barrel microelectrodes to measure changes in hepatocyte transmembrane potential (V _m), intracellular K⁺, Cl^-, and Na⁺ activities (a ⁱ _k, a _Cl ⁱ and a _Na ⁱ ), and water volume during l-alanine uptake. Mouse liver slices were superfused with control and experimental Krebs physiological salt solutions. The experimental solution contained 20 m l-alanine, and the control solution was adjusted to the same osmolality (305 mOsm) with added sucrose. Hepatocytes also were loaded with 50 mm tetramethylammonium ion (TMA⁺) for 10 min. Changes in cell water volume during l-alanine uptake were determined by changes in intracellular, steady-state TMA⁺ activity measured with the K⁺ electrode. Hepatocyte control V _m was -33±1 mV. l-alanine uptake first depolarized V _m by 2±0.2 mV and then hyperpolarized V _m by 5 mV to-38±1 mV (n = 16) over 6 to 13 min. During this hyperpolarization, a _Na ⁱ increased by 30% from 19±2 to 25±3 mm (P < 0.01), and a _K ⁱ did not change significantly from 83±3 mm. However, with added ouabain (1 mm) l-alanine caused only a 2-mV increase in V _m, but now a _K ⁱ decreased from 61±3 to 54±5 mm (P < 0.05). Hyperpolarization of V _m by l-alanine uptake also resulted in a 38% decrease of a _Cl ⁱ from 20±2 to 12±3 mm (P < 0.001). Changes in V _m and V _Cl — V _m voltage traces were parallel during the time of l-alanine hyperpolarization, which is consistent with passive distribution of intracellular Cl^– with the V _m in hepatocytes. Added Ba²⁺ abolished the l-alanineinduced hyperpolarization, and a _Cl ⁱ remained unchanged. Hepatocyte water volume during l-alanine uptake increased by 12±3%. This swelling did not account for any changes in ion activities following l-alanine uptake. We conclude that hepatocyte a _K ⁱ is regulated by increased Na⁺-K⁺ pump activity during l-alanine uptake in spite of cell swelling and increased V _m due to increased K⁺ conductance. The hyperpolarization of V _m during l-alanine uptake provides electromotive force to decrease a _Cl ⁱ . The latter may contribute to hepatocyte volume regulation during organic solute transport.This work was supported by grant AA-08867 from the Alcohol, Drug Abuse, and Mental Health Association.     

A COMPARATIVE STUDY ON THE SYNTHESIS OF THE NATRIURETIC HORMONE DOPAMINE IN OK AND LLC-PK1CELLS

TheV_maxvalues (in nmol/mg protein/15 min) for AAAD in OK cells (0.94±0.08) were found to be significantly (P<0.01) lower than those observed in LLC-PK₁cells (4.37±0.08). However, in both cell lines decarboxylation reaction was a saturable process with similarK_mvalues (OK cells=1.1 mm (0.3, 1.8); LLC-PK₁cells=1.8 mm (1.6, 2.1)). Contrariwise to OK cells, decarboxylation ofl -DOPA to dopamine in LLC-PK₁cells followed a linear (7.6±0.1 pmol/mg protein/min) non-saturable kinetics till 120 min of incubation. The formation of dopamine from increasing concentrations ofl -DOPA (10 to 500 μm ) followed a non-linear kinetics in both cell lines; the process ofl -DOPA decarboxylation was saturated at low concentrations ofl -DOPA with an apparentK_mvalue of 11 μm (0.2, 22.6) in OK cells and 27.4 μm (11.1, 43.7) in LLC-PK₁cells. The formation of dopamine in LLC-PK₁cells (V_max=2097±113 pmol/mg protein/6 min) was 13.7-fold that occurred in OK cells (V_max=153±10 pmol/mg protein/6 min). In conclusion, LLC-PK₁cells appear to be endowed with a greater ability to form dopamine from exogenousl -DOPA when compared to OK cells.     

Immobilized glucose dehydrogenase for cofactor regeneration in heme-catalyzed demethylation and hydroxylation reactions

Glucose dehydrogenase (E.C. 1.1.1.47) from B. megaterium M 1286 was immobilized together with mutarotase (E.C. 5.1.3.3) on several organic carriers and by different methods. The storage stability of the enzyme at pH-values > 6 is slightly improved by immobilization and the pH-optimum is shifted from 8.3 to 8.0. Kinetic constants of the immobilized enzyme are: K′_M(NAD⁺) = 5.36 × 10^?4 mol/l K′_M(glucose) = 3.76 · 10^?2 mol/l and V′_max = 5.54 · 10^?5 mol/(l min g carrier) for the most active preparation (2.16 mg enzyme/g carrier). In reactor experiments the immobilized glucose dehydrogenase was used with glucose to regenerate NADPH in NADPH-dependent iron-III-protoporphyrin-IX-imidazole catalyzed hydroxylation and demethylation of model substrates of cytochrome P-450. The advantages of the coupling of both reactions with cofactor recycling are shown and discussed.