β-Adrenoreceptor participation in the formation of the thermoregulatory and immune responses under the effect of rapid deep cooling

The effect of β-adrenoantagonist (obzidan) iontophoresis to skin on the thermoregulatory response and immune response to antigen was analyzed to elucidate the significance of β-adrenoceptors in formation of these responses at deep rapid cooling in rats.

On the background of β-adrenoceptors blockade in thermoneutral conditions the skin and core temperatures decreases; at rapid cooling non-shivering thermogenesis is attenuated and shivering thermogenesis is considerably enhanced.

Administration of β-adrenoantagonist affect the modulating influence of cold exposure on the immune response—the immunosuppressive effect of deep cooling on the immune response is abolished. This concerns both antigen binding function of spleen and peritoneal cells and antibody formation.

The results support the idea that β-adrenoceptors participates in the processes of the stimulation of thermogenesis and suppression of the immune response to antigen at rapid deep cooling.     

In vitro culture of the seagrass Halophila decipiens

The seagrass Halophila decipiens Ostenfeld was grown axenically in a culture medium consisting of 20% artificial seawater, f/4 nutrients (except that glutamic acid was the nitrogen source), and 1% sucrose (w:v). The culture medium was adjusted to pH 5.0. A root–rhizome layer was created by solidifying a portion of the medium with 0.9% agar (w:v) and 1% activated charcoal (w:v). The rhizome layer also contained the following vitamins: 0.5 mg l⁻¹ nicotinic acid, 0.5 mg l⁻¹ pyridoxine, 0.5 mg l⁻¹ biotin, 0.5 mg l⁻¹ cyanocobalamin and 0.1 mg l⁻¹ of thiamine HCl. The liquid overlay (without vitamins or charcoal) was poured onto the agar-solidified root–rhizome layer. Growth of H. decipiens was not improved by the addition of the auxins indoleacetic acid (IAA), indolebutyric acid (IBA) or naphthaleneacetic acid (NAA) at either of the tested concentrations (10 and 50 μM). At a concentration of 10 μM, the cytokinins 6-(γ,γ-dimethylallylamino) purine (2iP) and benzylaminopurine (BA) stimulated shoot and branch production compared to controls with no cytokinins. Among the tested nitrogen sources, growth was best on 1.7 mM glutamic acid. Cultures grown on 1.7 mM NH₄Cl showed the same growth rates as those grown on glutamic acid, but the leaves were smaller and curled, suggesting incipient ammonium toxicity. Use of nitrate or urea led to mortality of the cultures. Long term axenic culture of H. decipiens appears to require the added vitamins. Hence, H. decipiens is the first seagrass known to need exogenous vitamins. Cultures of H. decipiens died when grown suspended in liquid cultures or in a biphasic medium system without activated charcoal in the root–rhizome layer. The use of more highly charged κ-carrageenan could not replace the use of activated charcoal and agar in the root–rhizome layer.     

Arachidonic acid production by Mortierella alpina with growth-coupled lipid synthesis

The fungus Mortierella alpina LPM 301, a producer of arachidonic acid (ARA), was found to possess a unique property of a growth-coupled lipid synthesis. An increase in specific growth rate (μ) from 0.03 to 0.05 h⁻¹ resulted in a two-fold increase in the specific rate of lipid synthesis (milligram lipid (gram per lipid-free biomass) per hour). Under batch cultivation in glucose-containing media with urea or potassium nitrate as nitrogen sources, the ARA content was 46.0 and 60.4% of lipid; 16.4 and 18.8% of dry biomass; and 4.2 and 4.5 g l⁻¹, respectively. Under continuous cultivation of the strain, the productivity of ARA synthesis was 16.2 and 19.2 mg l⁻¹ h⁻¹ at μ=0.05 and 0.03 h⁻¹, respectively.     

Molecular cloning and expression of a cDNA for human kidney cysteine conjugate β-lyase

Kidney cysteine conjugate β-lyase (glutamine transaminase K, kyneurenine aminotransferase, EC 2.6.1.64) metabolises the cysteine conjugates of certain halogenated alkenes and alkanes to form reactive metabolites which can produce nephrotoxicicity and neurotoxicicity in experimental animals and man. Using a combination of hybridisation screening and PCR techniques we have isolated a full-length cDNA for human kidney cysteine conjugate β-lyase. Comparison of the deduced amino acid sequence with that of the rat enzyme indicated an 82% overall similarity, with 90% similarity around the pyridoxal phosphate binding site, many of the changes being conservative in nature. Expression of the cDNA in Cos-1 cells resulted in the production of a cytosolic enzyme which showed both cysteine conjugate β-lyase and glutamine transaminase K activity. Preliminary mapping of the gene for human cysteine conjugate β-lyase by PCR analysis of genomic DNA from human-rodent hybrid cells indicated that it is located on human chromosome 9.     

Optimisation of β-glucuronidase production from a newly isolated Ganoderma applanatum

Media optimisation was attempted for β-glucuronidase production from a newly and locally isolated (Oxfordshire, UK) fungal strain of Ganoderma applanatum. Both fungal growth and β-glucuronidase activity were found to be greatly affected by varying the carbon or the nitrogen source with gum arabic and yeast extracts being the best carbon and nitrogen sources, respectively. Their concentrations were optimised at 8 g L⁻¹ for the former and 2 g L⁻¹ for the latter.

Work then proceeded to enhance the yield of β-glucuronidase in a controlled environment. Control, batch and fed-batch cultivations were performed in 2-L bioreactors using the optimised medium supplemented with cellobiuronic acid as inducer. Time profiles of biomass dry weight, carbohydrate consumption and β-glucuronidase production were obtained and the results showed that production of β-glucuronidase was noticeably increased by the addition of cellobiuronic acid in both batch and fed-batch fermentations. Although the addition did not produce a variation in the pattern of growth seen between control, and induced fermenters, higher levels of the enzyme were attained when adopting a fed-batch process with 1.09 U mL⁻¹ of culture, corresponding to a 5-fold enhancement in β-glucuronidase production rate compared with batch fermentation.     

Evidence for modulation of cell-to-cell electrical coupling by cAMP in mouse islets of Langerhans

The effects of forskolin on electrical coupling among pancreatic β-cells were studied. Two microelectrodes were used to measure membrane potentials simultaneously in pairs of islet β-cells. Intracellular injection of a current pulse (ΔI) elicited a membrane response ΔV₁ in the injected cell and also a response ΔV₂ in a nearby β-cell confirming the existence of cell-to-cell electrical coupling among islet β-cells. In the presence of glucose (7 mM), application of forskolin evoked a transient depolarization of the membrane and electrical activity suggesting that the drug induced a partial inhibition of the β-cell membrane K⁺ conductance. Concomitant with this depolarization of the membrane there was a marked decrease in β-cell input resistance (ΔV₂/ΔI) suggesting that exposure to forskolin enhanced intercellular coupling. Direct measurements of the coupling ratio ΔV₂/ΔV₁ provided further support to the idea that forskolin enhances electrical coupling among islet cells. Indeed, application of forskolin reversibly increased the coupling ratio. These results suggest that cAMP might be involved in the modulation of electrical coupling among islet β-cells.     

Reversible β-hydride elimination of amine ligands forming stable cis-η-iminium hydride Os complexes

Octahedral tetraammineosmium(II) species are generated from their Os^III precursors containing an amine ligand cis to a labile alcohol or triflate. These compounds undergo reversible β-hydride eliminations resulting in the formation of cis-η²-iminium hydride complexes. Judging from NMR data, the η²-iminium group in these products lies parallel to the osmium-hydride bond with the iminium carbon eclipsing the hydride. Attempts to form η²-arene complexes of an Os^II ammine system bearing a stereogenic carbon are also described.     

Immobilization of β-galactosidase from Kluyveromyces lactis on silica and agarose: comparison of different methods

The covalent immobilization of β-galactosidase from Kluyveromyces lactis (β-gal) on to two different porous carriers, CPC-silica and agarose, is reported. CPC-silica was silanizated and activated with glutaraldehyde. The activation of agarose via a cyanylating agent (CDAP) was optimized. Gel-bound protein and gel-bound activity were both measured directly, allowing the determination of apparent specific activities (S.A.). Higher amounts of β-gal were immobilized on the activated CPC-silica (maximum capacity, 23 mg ml⁻¹ of packed support) than on the CDAP-activated agarose. For the lower enzyme loading assayed (12.6 mg ml⁻¹ packed support), 100% of the enzyme was immobilized but only 34% of its activity was expressed. This inactivation during immobilization was confirmed by the S.A. values (22–29 EU mg⁻¹ for the CPC-derivatives and 80 EU mg⁻¹ for soluble β-gal). The K_app (3.4 mM) for the CDAP-derivative with ONPG as substrate was higher than the K_M value for soluble β-gal (2 mM). When the enzyme loading was increased five-fold, the K_app increased four-fold, to 13 mM. The V_app values for the CPC-derivatives were remarkably lower than the V_max for soluble β-galactosidase. CDAP-derivatives showed better thermal stabilities than CPC-derivatives but neither of them enhanced the stability of the soluble enzyme. When stored at 4°C, the activity of both derivatives remained stable for at least 2 months. Both derivatives displayed high percentages of lactose conversion (90%) in packed bed mini-reactors. Glucose production was 3.3-fold higher for the CPC-derivative than for the CDAP-derivative, as a consequence of the higher flow rates achieved.     

Enzymatic activity of an extremely halophilic phosphatase from the Archaea Halobacterium salinarum in reversed micelles

Alkaline p-nitrophenylphosphate phosphatase (pNPPase) from the halophilic archaeon Halobacterium salinarum (previously halobium) was solubilized in reversed micelles of cetyltrimethylammonium bromide (CTAB) in cyclohexane with 1-butanol as cosurfactant. The hydrolysis reaction appears to follow Michaelis–Menten kinetics. The dependency of the maximum reaction rate (V_max) on the water content θ (% v/v) (or ω₀ value: molar ratio of water to surfactant concentrations) showed a bell-shaped curve for 0.3 M CTAB, but not for 0.2 M CTAB. The enzyme activity increased with the surfactant concentration at a constant ω₀ value (10.27). When the surfactant concentration was increased at a constant θ, the enzyme activity decreased. The enzyme was more stable in reversed micelles than in aqueous media.     

Inhibition of yeast by lactic acid bacteria in continuous culture: nutrient depletion and/or acid toxicity?

Lactic acid was added to batch very high gravity (VHG) fermentations and to continuous VHG fermentations equilibrated to steady state with Saccharomyces cerevisiae. A 53% reduction in colony-forming units (CFU) ml^–1 of S. cerevisiae was observed in continuous fermentation at an undissociated lactic acid concentration of 3.44% w/v; and greater than 99.9% reduction was evident at 5.35% w/v lactic acid. The differences in yeast cell number in these fermentations were not due to pH, since batch fermentations over a pH range of 2.5–5.0 did not lead to changes in growth rate. Similar fermentations performed in batch showed that growth inhibition with added lactic acid was nearly identical. This indicates that the apparent high resistance of S. cerevisiae to lactic acid in continuous VHG fermentations is not a function of culture mode. Although the total amount of ethanol decreased from 48.7 g l^–1 to 14.5 g l^–1 when 4.74% w/v undissociated lactic acid was added, the specific ethanol productivity increased ca. 3.2-fold (from 7.42×10^–7 g to 24.0×10^–7 g ethanol CFU^–1 h^–1), which indicated that lactic acid stress improved the ethanol production of each surviving cell. In multistage continuous fermentations, lactic acid was not responsible for the 83% (CFU ml^–1) reduction in viable S. cerevisiae yeasts when Lactobacillus paracasei was introduced to the system at a controlled pH of 6.0. The competition for trace nutrients in those fermentations and not lactic acid produced by L. paracasei likely caused the yeast inhibition.     

The role of 11β-hydroxysteroid dehydrogenase in steroid hormone specificity

11β-hydroxysteroid dehydrogenase (11β-HSD) is thought to confer aldosterone specificity to mineralocorticoid target cells by protecting the mineralocorticoid receptor (MR) from occupancy by endogenous glucocorticoids. In aldosterone target cells the type 2 11β-HSD is present, which, in contrast to the type 1 11β-HSD, has very high affinity for its substrate, is unidirectional and prefers NAD as cofactor. cDNAs encoding 11β-HSD2 have been recently cloned from different species, and the cell-specific expression of its mRNA and protein were determined. 11β-HSD2 is expressed in every aldosterone target tissue. Northern analysis revealed that the rabbit 11β-HSD2 is expressed at high levels in the renal collecting duct and at much lower levels in the colon. RT-PCR experiments demonstrated that 11β-HSD2 mRNA is present only in aldosterone target cells within the kidney. We determined the subcellular localization of the rabbit 11β-HSD2 using a chimera encoding 11β-HSD2 and the green fluorescent protein (GFP). This construct was stably transfected into CHO and MDCK cells. The expressed 11β-HSD2/GFP protein retained high enzymatic activity, and its characteristics were undistinguishable from those of the native enzyme. The intracellular localization of this protein was determined by fluorescence microscopy. 11β-HSD2-associated fluorescence was observed as a reticular network over the cytoplasm whereas the plasma membrane and the nucleus were negative, suggesting endoplasmic reticulum (ER) localization. Co-staining with markers for ER proteins, the Golgi membrane, mitochondria and nucleus confirmed that 11β-HSD2 is localized exclusively to the ER. To determine what structural motifs are responsible for the ER localization, we generated deletion mutants missing the C-terminal 42 and 118 amino acids, and fused them to GFP. Similarly as with the intact 11β-HSD2, these mutants localized exclusively to the ER. Both C-terminal deletion mutants completely lost dehydrogenase activity, independently whether activity was determined in intact cells or homogenates. These results indicate that 11β-HSD2 has a novel ER retrieval signal which is not localized to the C-terminal region. In addition, the C-terminal 118 amino acids are essential for NAD-dependent 11β-HSD activity.     

Influence of ethanol on the hopanoid content and the fatty acid pattern in batch and continuous cultures of Zymomonas mobilis

By thin layer chromatographic, gas-liquid chromatographic, and mass spectrometric methods 1,2,3,4-tetrahydroxypentane-29-hopane (THBH) was shown to occur in Zymomonas mobilis. This compound contributed up to 20% to the total lipids.The fatty acid pattern and the content of hopanoids (hopene, hopanol, and THBH) were determined in batch and continuous cultures. In late exponential cells from batch cultures the relative amount of palmitic acid was increased partially at the expense of cis-vaccenic acid, when the initial glucose concentrations were increased. In a batch culture, THBH reached a maximum value in the early exponential growth phase.In an anaerobic continuous culture with a low glucose feed concentration, the THBH content and the relative amount of cis-vaccenic acid were low. The contribution of both compounds increased strongly with increasing glucose feed concentrations (i.e. at higher steady-state ethanol concentrations). The same result was found with aerobic continuous cultures which produced significant amounts of acetaldehyde and acetic acid, in addition to ethanol and carbon dioxide.It was concluded that stability and permeability of the cytoplasmic membrane of the ethanol producing bacterium Z. mobilis was regulated by variations in the distribution of hopanoids and fatty acids.Abbreviations 14:0 myristic acid - 16:0 palmitic acid - 18:1 cisvaccenic acid - THBH 1,2,3,4-tetrahydroxypentane-29-hopane     

Enhancement of growth rate and β-galactosidase activity, and variation in organic acid profile of Bifidobacterium animalis subsp. lactis Bb 12

The behavior of Bifidobacterium animalis subsp. lactis Bb 12 under batch cultivation, after continuous culturing for up to 12 d, was monitored in skim milk-based media. Previous continuous culture for longer than 6 d affected the physiology of said microorganism. The minimum inhibitory concentrations of lactic and acetic acids increased from 18 to 26 g/l, whereas the molar ratio of acetic to lactic acid increased from 0.8 to 1.55, when the previous continuous culture increased its duration from 1 to 12 d. The specific lactose consumption rate decreased from 0.94 to 0.77 g_lactose/g_{cell dry mass}/h within the batch culture timeframe; this was concomitant with greater amounts of acetic and formic acids, and lower amounts of lactic acid produced. The β-galactosidase activity increased as continuous culturing time increased, and reached 446 units/ml by 12 d; however, the rate of enzyme synthesis decreased concomitantly. Succinic acid was produced during the exponential growth and stationary phases of the batch culture, but the former at exponential growth phase was higher as the continuous culturing time was longer. For comparison purposes, batch cultivation of samples taken from continuous cultures by 1 and 12 d was done using a semi-synthetic medium with glucose as carbon source; a pattern similar to that observed when using skim milk-based media was observed.     

Characterisation of the beta-tubulin gene of Cylicocyclus nassatus?

mRNA and genomic DNA were isolated from adult Cylicocyclus nassatus, and the mRNA was reverse transcribed. The cDNA was PCR amplified using degenerate primers designed according to the alignment of the β-tubulin amino acid sequences of other species. To complete the coding sequence, the 3′ end was amplified with the 3′-RACE, and for amplification of the 5′ end the SL1-primer was used. The cDNA of the β-tubulin gene of C. nassatus spans 1429 bp and encodes a protein of 448 amino acids. Specific primers were developed from the cDNA sequence to amplify the genomic DNA sequence and to analyse the genomic organisation of the β-tubulin gene. The complete sequence of the genomic DNA of the β-tubulin gene of C. nassatus has a size of 2652 bp and is organised into nine exons and eight introns. The identities with the exons of the gru-1 β-tubulin gene of Haemonchus contortus range between 79% and 97%.     

3D-structure of human estrogenic 17beta-HSD1: binding with various steroids.

Human estrogenic dehydrogenase (17β-HSD1) catalyses the last step in the biosynthesis of the active estrogens that stimulate the proliferation of breast cancer cells. While the primary substrate for the enzyme is estrone, the enzyme has some activity for the non-estrogenic substrates. To better understand the structure–function relationships of 17β-HSD1 and to provide a better ground for the design of inhibitors, we have determined the crystal structures of 17β-HSD1 in complex with different steroids.

The structure of the complex of estradiol with the enzyme determined previously (Azzi et al., Nature Structural Biology 3, 665–668) showed that the narrow active site was highly complementary to the substrate. The substrate specificity is due to a combination of hydrogen bonding and hydrophobic interactions between the steroid and the enzyme binding pocket. We have now determined structures of 17β-HSD1 in complex with dihydrotestosterone and 20-OH-progesterone. In the case of the C19 androgen, several residues within the enzyme active site make some small adjustments to accommodate the increased bulk of the substrate. In addition, the C19 steroids bind in a slightly different position from estradiol with shifts in positions of up to 1.4 Å. The altered binding position avoids unfavorable steric interactions between Leu 149 and the C19 methyl group (Han et al., unpublished). The known kinetic parameters for these substrates can be rationalized in light of the structures presented. These results give evidence for the structural basis of steroid recognition by 17β-HSD1 and throw light on the design of new inhibitors for this pivotal steroid enzyme.     

Simultaneous production of anthocyanin and triterpenoids in suspension cultures of Perilla frutescens

When cultivated in Murashige & Skoog medium supplemented with 0.2 mg l⁻¹ 2,4-dichlorophenoxy acetic acid and 0.5 mg l⁻¹ 6-benzyladenine, Perilla frutescens cells in suspension culture grew rapidly reaching about 13.6 g dry wt l⁻¹ after 12 days. The cell line produced both anthocyanin 0.9 g l⁻¹ and triterpenoids: 16 mg l⁻¹ oleanolic acid (OA), 25 mg l⁻¹ ursolic acid (UA) and 14 mg l⁻¹ tormentic acid (TA). When P. frutescens cells of 7-day-old cultures were exposed to a yeast elicitor at 0.5–5% (v/v) for 7 days, it was found that anthocyanin content peaked at 10.2% of dry weight with yeast elicitor at 1% (v/v) whereas the maximum production of oleanolic acid and ursolic acid in cultures treated with 2% (v/v) yeast elicitor was 19 and 27 mg l⁻¹, a 46 and 24% increase over the control, respectively. This is the first report of simultaneous production of both anthocyanin and triterpenoids in a single culture system.     

η-Allyl-cobalt (II) complexes

(η³-Cyclooctenyl)Co(bisphosphine) compounds react with HBF₄ in the presence of alkenes with oxidation of the metal to give the novel, paramagnetic organocobalt(II) species [(η³-cyclooctenyl)Co(bisphosphine)]⁺BF₄⁻, (η³-2-RC₃H₄)Co(bisphosphine) complexes react similarly. The Co(II) compounds form adducts with CO and NO (the latter being diamagnetic) and undergo facile chemical and electrochemical reduction.     

Influence of the spacer length on the activity of enzymes immobilised on nylon/polyGMA membranes: Part 2: Non-isothermal conditions

β-Galactosidase has been immobilised through spacers of different length on nylon membranes grafted with glycidyl methacrylate. Hexamethylendiamine, ethylendiamine or hydrazine have been separately used as spacers.

The behaviour of the catalytic membranes has been studied in a bioreactor operating under non-isothermal conditions as a function of the applied temperature difference ΔT.

Comparison of the enzyme reaction rates under isothermal and non-isothermal conditions resulted in percentage activity increases (PAI) and reduction of the production time (τ_r) proportional to the size of the applied ΔT. Both these parameters increased with the increase of the spacer length.

Results have been discussed in the frame of reference of the process of thermodialysis which reduces the limitations to the diffusion of substrate and reaction products across the catalytic membrane, limitations introduced by the grafting and immobilisation process.

The advantages of employing non-isothermal bioreactors in biotechnological productive processes have been outlined.     

Anti-diabetic properties of the Canadian lowbush blueberry Vaccinium angustifolium Ait.

Incidence of type II diabetes is rapidly increasing worldwide. In order to identify complementary or alternative approaches to existing medications, we studied anti-diabetic properties of Vaccinium angustifolium Ait., a natural health product recommended for diabetes treatment in Canada. Ethanol extracts of root, stem, leaf, and fruit were tested at 12.5 μg/ml for anti-diabetic activity in peripheral tissues and pancreatic β cells using a variety of cell-based bioassays. Specifically, we assessed: (1) deoxyglucose uptake in differentiated C2C12 muscle cells and 3T3-L1 adipocytes; (2) glucose-stimulated insulin secretion (GSIS) in β TC-tet pancreatic β cells; (3) β cell proliferation in β TC-tet cells; (4) lipid accumulation in differentiating 3T3-L1 cells; (5) protection against glucose toxicity in PC12 cells. Root, stem, and leaf extracts significantly enhanced glucose transport in C2C12 cells by 15–25% in presence and absence of insulin after 20 h of incubation; no enhancement resulted from a 1 h exposure. In 3T3 cells, only the root and stem extracts enhanced uptake, and this effect was greater after 1 h than after 20 h; uptake was increased by up to 75% in absence of insulin. GSIS was potentiated by a small amount in growth-arrested β TC-tet cells incubated overnight with leaf or stem extract. However, fruit extracts were found to increase ³H-thymidine incorporation in replicating β TC-tet cells by 2.8-fold. Lipid accumulation in differentiating 3T3-L1 cells was accelerated by root, stem, and leaf extracts by as much as 6.5-fold by the end of a 6-day period. Stem, leaf, and fruit extracts reduced apoptosis by 20–33% in PC12 cells exposed to elevated glucose for 96 h. These results demonstrate that V. angustifolium contains active principles with insulin-like and glitazone-like properties, while conferring protection against glucose toxicity. Enhancement of proliferation in β cells may represent another potential anti-diabetic property. Extracts of the Canadian blueberry thus show promise for use as a complementary anti-diabetic therapy.     

Production of α-tocopherol by sequential heterotrophic–photoautotrophic cultivation of Euglena gracilis

Photoautotrophic cultivation of Euglena gracilis results in cells with high α-tocopherol content but the final cell concentration is usually very low due to the difficulty of supplying light efficiently to the photobioreactor. On the other hand, Euglena grows heterotrophically to high cell concentrations, using various organic carbon sources, but the α-tocopherol contents of heterotrophically grown cells are usually very low. Sequential heterotrophic/photoautotrophic cultivation, by which cells are grown heterotrophically to high cell concentrations and then transferred to photoautotrophic culture for accumulation of α-tocopherol was therefore investigated for efficient α-tocopherol production. In batch culture, using glucose as the organic carbon source, the cellular α-tocopherol content increased from 120 μg g⁻¹ at the end of heterotrophic phase to more than 400 μg g⁻¹ at the end of the photoautotrophic phase. By using ethanol as the organic carbon source during the heterotrophic phase, adding corn steep liquor as a nitrogen source and optimizing light supply during the photoautotrophic phase, the α-tocopherol content of the cells at the end of the photoautotrophic phase increased to 1700 μg g⁻¹. A system consisting of a mini-jar fermentor (for the heterotrophic phase) and an internally illuminated photobioreactor (for the photoautotrophic phase) was then constructed for continuous sequential heterotrophic/photoautotrophic cultivation. The cells were continuously cultivated heterotrophically in the mini-jar fermentor and the effluent was continuously passed through the photobioreactor for α-tocopherol accumulation. In this way, it was possible to produce 7 g l⁻¹ cells containing about 1100 μg α-tocopherol per g-cell continuously for more than 420 h. The continuous process resulted in α-tocopherol productivity of 100 μg l⁻¹ h⁻¹ which is about 9.5 and 4.6 times higher than those obtained in batch photoautotrophic culture and batch heterotrophic cultures, respectively.