Citrinin-producing capacity of Monascus purpureus in response to low −frequency magnetic fields

Citrinin is a type of mycotoxin that negatively affects Monascus products. Application of a low-frequency magnetic field (LF-MF) decreased citrinin production by M. purpureus in liquid-state fermentation during shake flask culture. Under 30 °C incubation, six different magnetic field induction intensity (MF-II), four different exposure times and five exposure time periods were tested to discover optimal treatment conditions. The cultures were exposure to a MF-II of 1.6 mT from 0 to 2 d of incubation time. With LF-MF treatment, peak citrinin production decreased by 46.7% while yellow, red, orange pigments and monacolin K production increased by 31.3%, 40.3%, 41.7% and 29.3%, respectively, compared with control groups at 12 d of incubation. Moreover, the relative expression levels of the citrinin biosynthesis genes pksCT and ctnA were 0.46 and 0.43 times lower, respectively, than the control values relative to the GAPDH reference gene. This study provides evidence that LF-MF is a preferable way to alter M. purpureus metabolism to reduce citrinin production and to increase pigments and monacolin K production without affecting cell growth. Therefore, LF-MF may be used as a tool to process Monascus products to obtain important functional food additive while reducing the adverse effects of citrinin.     

Growth kinetics of biopigment production by Thai isolated <Emphasis Type="Italic">Monascus purpureus</Emphasis> in a stirred tank bioreactor

Monascus purpureus is a biopigment-producing fungi whose pigments can be used in many biotechnological and food industries. The growth kinetics of biopigment production were investigated in a liquid fermentation medium in a 5-l stirred tank bioreactor at 30°C, pH 7, for 8 days with 100 rpm agitation and 1.38 × 10⁵ N/m² aeration. Thai Monascus purpureus strains TISTR 3002, 3180, 3090 and 3385 were studied for color production, growth kinetics and productivity. Citrinin as a toxic metabolite was measured from the Monascus fermentation broth. The biopigment productions were detected from fermentation broth by scanning spectra of each strain produced. Results showed a mixture of yellow, orange and red pigments with absorption peaks of pigments occurring at different wavelengths for the four strains. It was found that for each pigment color, the color production from the strains increased in the order TISTR 3002, 3180, 3090, 3385 with 3385 production being approximately 10 times that of 3002. Similar results were found for growth kinetics and productivity. HPLC results showed that citrinin was not produced under the culture conditions of this study. The L*, a* and b* values of the CIELAB color system were also obtained for the yellow, orange and red pigments produced from the TISTR 3002, 3180, 3090 and 3385 strains. The colors of the pigments ranged from burnt umber to deep red.     

The role of intracellular sodium in the regulation of NMDA-receptor-mediated channel activity and toxicity

Sodium (Na⁺) is the major cation in extracellular space and, with its entry into cells, may act as a critical intracellular second messenger that regulates many cellular functions. Through our investigations of mechanisms underlying the activity-dependent regulation of N-methyl-d-aspartate (NMDA) receptors, we recently characterized intracellular Na⁺ as a possible signaling factor common to processes underlying the upregulation of NMDA receptors by non-NMDA glutamate channels, voltage-gated Na⁺ channels, and remote NMDA receptors. Furthermore, although Ca²⁺ influx during the activation of NMDA receptors acts as a negative feedback mechanism that downregulates NMDA receptor activity, Na⁺ influx provides an essential positive feedback mechanism to overcome Ca²⁺-induced inhibition, thereby potentiating both NMDA receptor activity and inward Ca²⁺ flow. NMDA receptors may be recruited to cause excitoxicity through a Na⁺-dependent mechanism. Therefore, the further characterization of mechanisms underlying the regulation of NMDA receptors by intracellular Na⁺ is essential to understanding activity-dependent neuroplasticity in the nervous system.     

Light-induced extracellular calcium and sodium concentration changes in the retina of Calliphora: involvement in the mechanism of light adaptation

Summary Ion-selective microelectrodes inserted into the compound eyes of Calliphora were used to monitor the changes in extracellular concentration of Ca²⁺ and Na⁺ (Ca_o, Na^o) brought about by a 1-min exposure to white light (maximal luminous intensity 0.1 cd/cm²).Using Ringer solution as the reference (Ca²⁺ = 1 mM), the dark concentration of the calcium in the retina was found to be (1.4 ± 0.4) mM (n=12). Stimulation with light reduces Ca_o. At intensities near maximal the Ca_o signal is phasic, reaching a transient minimum about 6 s after light onset and then rising to a nearly stable plateau below the dark level (-3.3% ± 2.6%). Ca_o signals measured in the white-eyed mutant (chalky), which lacks pigment granules, are comparable to those in the wild type.Conclusions: (a) There are no extracellular Ca²⁺ binding sites that regulate light adaptation, such as were postulated by Hochstrate and Hamdorf (1985). (b) Ca²⁺ influx into the photoreceptors seems to be necessary for light adaptation, (c) The pigment granules have no major function in intracellular calcium regulation.The time course of the Na_o signals resembles that of the Ca_o signals. Because the percentage concentration change is small, light-induced extracellular Na⁺-depletion cannot contribute to a reduced response amplitude at light adaptation.Abbreviations Ca _i intracellular Ca²⁺ concentration - Ca _o extracellular Ca²⁺ concentration - Kino extracellular K⁺ concentration - Na _o extracellular Na⁺ concentration     

Characterization of a non-pigment producing Monascus purpureus mutant strain

A characterization of a non-pigment producing mutant Monascus purpureus M₁₂ compared with its parental strain Monascus purpureus Went CBS 109.07 has been performed aiming to investigate the relation between pigment biosynthesis and other characteristics of these fungi. A comparison has been made of morphological features, some physiological properties and biochemical activities of both strains. The albino mutant exhibits an anamorph life cycle, high conidia forming capability, slower radial growth rate and temperature sensitivity. The assimilation capacity of both strains for mono-, disaccharides and some alcohols is in the same range (Y_X/C 0.2 – 0.35), while the red strain has a higher fermentation capacity. In a selected albino mutant, the growth rate, metabolic activity and capacity for production of typical for Monascus fungi secondary metabolites were reduced considerably. Hydrolytic activity towards natural substrates expressed through glucoamylase and protease was approximately 10 fold lower in the non pigment producing strain (0.05 – 0.08 U/mg protein and 0.01 – 0.07 U/mg protein respectively) compared with the red one. Important qualitative differences between both strains was found in fatty acid composition and in the production of citrinin and monacolin. The mutant strain possessed C₁₇, C₂₀ and C₂₂ fatty acids and did not produce citrinin. This revised version was published online in June 2006 with corrections to the Cover Date.     

Ameliorative Effects of a Static Magnetic Field on Hyssop (Hyssopus officinalis L.) Growth and Phytochemical Traits Under Water Stress

The present study was carried out in order to evaluate the promoting effect of a static magnetic field (SMF) on drought tolerance and medicinal properties in Hyssopus officinalis. In the current work, the effect of seed priming with SMF (45, 90, 200, and 250 mT for 5 min) was investigated in 60-day-old hyssop (H. officinalis) plants that were irrigated every 8 days. The assessments consisted of total dry mass, membrane integrity, photosynthetic pigment concentrations, polyphenol content, antioxidant enzyme activities, and antioxidant capacity. Compared with exclusively water stress, magnetopriming, particularly at 200 mT, significantly altered these parameters in the grown plants. At this intensity, the level of total dry mass, total chlorophyll, and polyphenol content increased by 94%, 2.5- and 7.7-fold, respectively. Also, the level of electrolyte leakage and malondialdehyde decreased by 35% and 33%. The reducing power, DPPH (1,1-diphenyl-2-picrylhydrozyl), and superoxide anion-scavenging activities were highly augmented as well. Magnetopriming at 200 mT increased catalase (+92%) and ascorbate peroxidase (+2.3-fold) activities. However, the highest activity of guaiacol peroxidase was recorded at 90 mT. Generally, the present study illustrated the positive effect of magnetopriming (200 mT) on improvement of drought tolerance in H. officinalis through protection of cellular membrane integrity, maintenance of photosynthetic pigment content, and alternation of antioxidant enzyme activities. Furthermore, the data showed this treatment (200 mT) not only had no negative effect on medicinal properties of H. officinalis, but also improved it via increasing total phenolic content and antioxidant capacity. Bioelectromagnetics. 2020;41:403–412. © 2020 Bioelectromagnetics Society.     

Production of pigments byMonascus purpureus using sugar-cane bagasse in roller bottle cultures

Solid-state fermentation, using sugar-cane bagasse, and submerged fermentation, using a semi-synthetic medium, were performed for pigment production byMonascus purpureus in both stationary and rotary conditions. Rotary cultures gave higher yields of crude red and yellow pigments than stationary cultures whereas twice the amount was synthesized at an earlier time (day 8) in liquid medium (1,285U yellow pigment/bottle, 1,728U red pigment/bottle). Supplementing the liquid medium with 0.6% (v/v) corn oil doubled the extracellular pigment yield but halved fungal growth.     

Repeated-batch production of pigments by immobilised Monascus purpureus

Extracellular pigment production by immobilised Monascus purpureus C322 has been studied in repeated-batch processes using different immobilising carriers such as Ca-alginate, polyurethane sponge, active carbon and pearlite. With Ca-alginate, pigment production was maximum (30.5 UA₄₇₀ as process mean production, three batches) while the cell leakage was negligible (0.4 g l⁻¹ free biomass) and the bead mechanical stability good; with this carrier, an extended repeated-batch fermentation (nine batches, 55 days) was carried out: the process pigment productivity was 3.87 UA₄₇₀ day⁻¹.     

23Na multiple quantum filtered NMR characterisation of Na+ binding and dynamics in animal cells: a comparative study and effect of Na+/Li+ competition

Double quantum and triple quantum filtered ²³Na nuclear magnetic resonance techniques were used to characterise in detail the isotropic and anisotropic binding and dynamics of intra- and extracellular Na⁺ in different cellular systems, in the absence and presence of Li⁺. The kinetics of Li⁺ influx by different cell types was evaluated. At steady state, astrocytes accumulated more Li⁺ than red blood cells (RBCs), while a higher intracellular Li⁺ concentration was found in chromaffin than in SH-SY5Y cells. Anisotropic and isotropic motions were detected for extracellular Na⁺ in all cellular systems studied. Isotropic intracellular Na⁺ motions were observed in all types of cells, while anisotropic Na⁺ motions in the intracellular compartment were only detected in RBCs. ²³Na triple quantum signal efficiency for intracellular Na⁺ was SH-SY5Y > chromaffin > RBCs, while the reverse order was observed for the extracellular ions. ²³Na double quantum signal efficiency for intracellular Na⁺ was non-zero only in RBCs, and for extracellular Na⁺ the order RBCs > chromaffin > SH-SY5Y cells was observed. Li⁺ loading generally decreased intracellular Na⁺ isotropic movements in the cells, except for astrocytes incubated with a low Li⁺ concentration and increased anisotropic intracellular Na⁺ movements in RBCs. Li⁺ effects on the extracellular signals were more complex, reflecting Li⁺/Na⁺ competition for isotropic and anisotropic binding sites at the extracellular surface of cell membranes and also at the surface of the gel used for cell immobilisation. These results are relevant and contribute to the interpretation of the in vivo pharmacokinetics and sites of Li⁺ action.     

Na+ Sensitivity of ROMK1 K+ Channel: Role of the Na+/H+ Antiporter

To examine the extracellular Na⁺ sensitivity of a renal inwardly rectifying K⁺ channel, we performed electrophysiological experiments on Xenopus oocytes or a human kidney cell line, HEK293, in which we had expressed the cloned renal K⁺ channel, ROMK1 (Kir1.1). When extracellular Na⁺ was removed, the whole-cell ROMK1 currents were markedly suppressed in both the oocytes and HEK293 cells. Single-channel ROMK1 activities recorded in the cell-attached patch on the oocyte were not affected by removal of Na⁺ from the pipette solution. However, macro-patch ROMK1 currents recorded on the oocyte were significantly suppressed by Na⁺ removal from the bath solution. A blocker of Na⁺/H⁺ antiporters, amiloride, largely inhibited the Na⁺ removal-induced suppression of whole-cell ROMK1 currents in the oocytes. The pH-insensitive K80M mutant of ROMK1 was much less sensitive to Na⁺ removal. Na⁺ removal was found to induce a significant decrease in intracellular pH in the oocytes using H⁺-selective microelectrodes. Coexpression of ROMK1 with NHE3, which is a Na⁺/H⁺ antiporter isoform of the kidney apical membrane, conferred increased sensitivity of ROMK1 channels to extracellular Na⁺ in both the oocytes and HEK293 cells. Thus, it is concluded that the ROMK1 channel is regulated indirectly by extracellular Na⁺, and that the interaction between NHE transporter and ROMK1 channel appears to be involved in the mechanism of Na⁺ sensitivity of ROMK1 channel via regulating intracellular pH. Received: 13 April 1999/Revised: 15 July 1999     

Characterization of Na+ transport in normal human fibroblasts and neoplastic H.Ep.2 cells and the role of inhibitin

Summary Na⁺ transport was characterized in normal human fibroblasts and neoplastic H.Ep. 2 cells in order to investigate the role of the endogenous peptidic factor inhibitin that is secreted by a variety of neoplastic cells (including H.Ep. 2) and inhibits Na⁺/Na⁺ exchange in human erythrocytes. Although active (Na⁺, K⁺-ATPase mediated) Na⁺ fluxes were similar in the two cell types, H.Ep. 2 cells maintained higher intracellular Na[su+] concentration (26mm) compared to fibroblasts (12mm). An analysis of passive Na⁺ fluxes showed a difference in the handling of Na⁺ via ouabain and bumetanide-insensitive transport between the two cell types: H.Ep. 2 cells achieved net Na⁺ influx via an amiloride-sensitive pathway that was only demonstrated in fibroblasts when 10% fetal calf serum (FCS) was present. Kinetic studies were undertaken to investigate the interaction between Na⁺ flux via Na⁺/H⁺ and Na⁺/Na⁺ exchanges. for this purpose, an outwardly directed Na⁺ gradient was created by loading the cells with Na⁺ (Na _i >100mm) to activate the reverse functioning of Na⁺/H⁺ exchange (i.e., Na _out ⁺ H _in ⁺ ). The rates of ouabain-and bumetanide-insensitive Na⁺ efflux were measured over a range of extracellular Na⁺ concentrations (Na _o ⁺ 14–140mm). In the presence of 10% FCS, the two cell types showed different responses: in fibroblasts the Na⁺ efflux rate showed an inverse correlation with extracellular Na⁺ concentration, while H.Ep. 2 cells significantly increased their rate of Na⁺ efflux as extracellular Na⁺ concentration increased. So although the thermodynamic force would direct net Na⁺ efflux when Na _i ⁺ >Na _o ⁺ , H.Ep.2 cells were under kinetic control to perform Na⁺/Na⁺ exchange.When exogenous inhibitin was tested on fibroblasts, the steady-state intracellular Na⁺ concentration increased from 14 to 19mm (p<0.01). In Na⁺-loaded fibroblasts, serum-stimulated Na⁺ efflux was partially inhibitin sensitive and the maximal inhibitory effect was seen when extracellular Na⁺ concentration was 14mm and presumably the Na⁺/H⁺ exchanger operating in the reverse mode. This study demonstrated that, in contrast to fibroblasts, H.Ep.2 cells have a modified Na⁺/H⁺ exchange system whereby it acts in the Na _in ⁺ H _out ⁺ mode without exogenous growth factor activation and resists functioning in the reversed mode. It is proposed that inhibitin, is the endogenous modifier of this transport system in H.Ep.2 cells with the result that H.Ep.2 cells maintain a higher concentration of intracellular Na⁺ compared to fibroblasts.     

Intracellular sodium homeostasis in relation to the effective free-energy change of ATP hydrolysis: studies of crayfish muscle fibers

We investigated the impact of reductions in the effective free-energy change of ATP hydrolysis (dG/d_ATP) on intracellular sodium homeostasis in bundles of fibers from the abdominal extensor muscle of the crayfish Procambarus clarkii. ³¹P nuclear magnetic resonance (NMR) spectroscopy was used to monitor high-energy phosphate levels and intracellular pH while interleaved ²³Na-NMR spectra were acquired to monitor changes in sodium. Previous work has shown that the bulk of intracellular Na⁺ is NMR visible (see Ivanics et al. 1994). The ²³Na-NMR spectra were diffusion-weighted which effectively filtered out signal contributions from extracellular sodium. The efficacy of this procedure was validated using a relatively non-toxic chemical shift reagent which allowed resolution of extracellular and intracellular Na⁺ signals. Metabolic inhibition (cyanide/iodoacetate) produced pronounced reductions in dG/d_ATP coincident with dramatic increases in intracellular Na⁺ levels ([Na⁺]_i). However, the increases in [Na⁺]_i occurred at dG/d_ATP values well above the threshold value of −46 kJ · mol⁻¹ required by the existing Na⁺ gradient and the membrane potential. These results suggest that the global dG/d_ATP value may not reflect the dG/d_ATP value in the vicinity of the pump. Alternatively, other factors, including low molecular modulators of Na⁺, K⁺-ATPase activity, may be important in this context. Accepted: 15 May 1997     

Nitric oxide and pH modulation in gynaecological cancer

Nitric oxide plays several roles in cellular physiology, including control of the vascular tone and defence against pathogen infection. Neuronal, inducible and endothelial nitric oxide synthase (NOS) isoforms synthesize nitric oxide. Cells generate acid and base equivalents, whose physiological intracellular concentrations are kept due to membrane transport systems, including Na⁺/H⁺ exchangers and Na⁺/HCO₃^? transporters, thus maintaining a physiological pH at the intracellular (~7.0) and extracellular (~7.4) medium. In several pathologies, including cancer, cells are exposed to an extracellular acidic microenvironment, and the role for these membrane transport mechanisms in this phenomenon is likely. As altered NOS expression and activity is seen in cancer cells and because this gas promotes a glycolytic phenotype leading to extracellular acidosis in gynaecological cancer cells, a pro‐inflammatory microenvironment increasing inducible NOS expression in this cell type is feasible. However, whether abnormal control of intracellular and extracellular pH by cancer cells regards with their ability to synthesize or respond to nitric oxide is unknown. We, here, discuss a potential link between pH alterations, pH controlling membrane transport systems and NOS function. We propose a potential association between inducible NOS induction and Na⁺/H⁺ exchanger expression and activity in human ovary cancer. A potentiation between nitric oxide generation and the maintenance of a low extracellular pH (i.e. acidic) is proposed to establish a sequence of events in ovarian cancer cells, thus preserving a pro‐proliferative acidic tumour extracellular microenvironment. We suggest that pharmacological therapeutic targeting of Na⁺/H⁺ exchangers and inducible NOS may have benefits in human epithelial ovarian cancer.     

离子转运蛋白调控钝齿棒杆菌离子和pH稳态促进L-精氨酸合成

L-精氨酸是一种半必需氨基酸,广泛应用于食品、制药、饲料等行业。【目的】当前对L-精氨酸生产菌株的研究,极少涉及离子转运领域。在本研究中,发现在发酵时适量添加外源K~+有利于促进钝齿棒杆菌(Corynebacterium crenatum) SYPA5-5合成L-精氨酸。【方法】在C. crenatum SYPA5-5发酵培养基外源添加0.5 g/L和2.5 g/L的K_3PO_4,取对数期发酵样品进行转录组数据分析,挖掘出K~+转运相关的阳离子转运ATP酶CTAP1以及单价阳离子/H~+逆转运蛋白Mrp1A,研究其在C. crenatum SYPA5-5快速合成L-精氨酸阶段,对菌株生长及L-精氨酸合成的影响。【结果】对基因ctap1和mrp1分别进行敲除和过表达,深入研究突变株对L-精氨酸合成的影响。研究发现同时过表达离子转运蛋白CTAP1和Mrp1A更有利于胞内离子、pH稳态和渗透压调节,最终提高L-精氨酸的产量。在补料分批发酵中分别过表达Mrp1A、CTAP1以及同时过表达Mrp1A和CTAP1的菌株L-精氨酸产量分别达到61.4 g/L、63.9 g/L和65.3 g/L,产率分别为0.383 g/g、0.392 g/g和0.395 g/g,比C. crenatum SYPA5-5分别提高了34.9%、38.0%和39.1%。【结论】CTAP1是特异性的K~+转运ATP酶,可以将培养基中的K~+运输到胞内。同时Mrp1A可将胞内K~+和Na~+等单价阳离子运输到胞外,将胞外H~+运输至胞内,中和胞内L-精氨酸所导致的碱性环境,从而维持胞内pH稳定。CTAP1和Mrp1A的研究为解析离子转运机制和L-精氨酸合成之间的联系奠定了基础。     

Activity-Dependent Adenosine Release May Be Linked to Activation of Na+-K+ ATPase: An In Vitro Rat Study

In the brain, extracellular adenosine increases as a result of neuronal activity. The mechanisms by which this occurs are only incompletely understood. Here we investigate the hypothesis that the Na⁺ influxes associated with neuronal signalling activate the Na⁺-K⁺ ATPase which, by consuming ATP, generates intracellular adenosine that is then released via transporters. By measuring adenosine release directly with microelectrode biosensors, we have demonstrated that AMPA-receptor evoked adenosine release in basal forebrain and cortex depends on extracellular Na⁺. We have simultaneously imaged intracellular Na⁺ and measured adenosine release. The accumulation of intracellular Na⁺ during AMPA receptor activation preceded adenosine release by some 90 s. By removing extracellular Ca²⁺, and thus preventing indiscriminate neuronal activation, we used ouabain to test the role of the Na⁺-K⁺ ATPase in the release of adenosine. Under conditions which caused a Na⁺ influx, brief applications of ouabain increased the accumulation of intracellular Na⁺ but conversely rapidly reduced extracellular adenosine levels. In addition, ouabain greatly reduced the amount of adenosine released during application of AMPA. Our data therefore suggest that activity of the Na⁺-K⁺ ATPase is directly linked to the efflux of adenosine and could provide a universal mechanism that couples adenosine release to neuronal activity. The Na⁺-K⁺ ATPase-dependent adenosine efflux is likely to provide adenosine-mediated activity-dependent negative feedback that will be important in many diverse functional contexts including the regulation of sleep.     

K+-conductance and electrogenic Na+/K+ transport of cultured bovine pigmented ciliary epithelium

Summary Using intracellular microelectrode technique, we investigated the changes in membrane voltage (V) of cultured bovine pigmented ciliary epithelial cells induced by different extracellular solutions. (1)V in 213 cells under steady-state conditions averaged –46.1±0.6 mV (sem). (2) Increasing extracellular K⁺ concentration ([K⁺] _o ) depolarizedV. Addition of Ba²⁺ could diminish this response. (3) Depolarization on doubling [K⁺] _o was increased at higher [K⁺] _o (or low voltage). (4) Removing extracellular Ca²⁺ decreasedV and reduced theV amplitude on increasing [K⁺] _o . (5)V was pH sensitive. Extra-and intracellular acidification depolarizedV; alkalinization induced a hyperpolarization.V responses to high [K⁺] _o were reduced at acidic extracellular pH. (6) Removing K _o ⁺ depolarized, K _o ⁺ readdition after K⁺ depletion transiently hyperpolarizedV. These responses were insensitive to Ba²⁺ but were abolished in the presence of ouabain or in Na⁺-free medium. (7) Na⁺ readdition after Na⁺ depletion transiently hyperpolarizedV. This reaction was markedly reduced in the presence of ouabain or in K⁺-free solution but unchanged by Ba²⁺. It is concluded that in cultured bovine pigmented ciliary epithelial cells K⁺ conductance depends on Ca²⁺, pH and [K⁺] _o (or voltage). An electrogenic Na⁺/K⁺-transport is present, which is stimulated during recovery from K⁺ or Na⁺ depletion. This transport is inhibited by ouabain and in K⁺-or Na⁺-free medium.     

Cation flux in the ehrlich ascites tumor cell evidence for Na-for-Na and K-for-K exchange diffusion

In a previous study, evidence was presented for an external Na⁺-dependent, ouabain-insensitive component of Na⁺ efflux and an external K⁺-dependent component of K⁺ efflux in the Ehrlich ascites tumor cell. Evidence is now presented that these components are inhibited by the diuretic furosemide and that under conditions of normal extracellular Na⁺ and K⁺ they represent Na⁺-for-Na⁺ and K-⁺for-K⁺ exchange mechanisms. Using ⁸⁶Rb to monitor K⁺ movements, furosemide is shown to inhibit an ouabain-insensitive component of Rb⁺ influx and a component of Rb⁺ efflux, both representing approx. 30% of the total fux. Inhibition of Rb⁺ efflux is greatly reduced by removal of extracellular K⁺. Furosemide does not alter steady-state levels of intracellular K⁺ and it does not prevent cells depleted of K⁺ by incubation in the cold from regaining K⁺ upon warming. Using ²²Na to monitor Na⁺ movements, furosemide is shown to inhibit an ouabain-insensitive component of unidirectional Na⁺ efflux which represents approx. 22% of total Na⁺ efflux. Furosemide does not alter steady-state levels of intracellular Na⁺ and does not prevent removal of intracellular Na⁺ upon warming from cells loaded with Na⁺ by preincubation in the cold. The ability of furosemide to affect unidirectional Na⁺ and K⁺ fluxes but not net fluxes is consistent with the conclusion that these components of cation movement across the cell membrane represent one-for-one exchange mechanisms. Data are also presented which demonstrate that the uptake of α-aminoisobutyrate is not affected by furosemide. This indicates that these components of cation flux are not directly involved in the Na⁺-dependent amino acid transport system A.     

Regulation of intracellular level of Na+, K+ and glycerol in Saccharomyces cerevisiae under osmotic stress

The intracellular level of Na⁺ and K⁺ of S. cerevisiae strain AB1375 revealed that under KCl as well as sorbitol stress, the cationic level was comparable to the level under no stress conditions. On the other hand, there was a sharp drop in the intracellular K⁺ content and increase in the Na⁺ content on addition of NaCl to the medium. However, the total cationic level was close to that under control conditions. In addition to changes in the cationic level, an enhanced production and accumulation of glycerol were also observed under osmotic stress. A regulatory mechanism co-ordinating the intracellular concentration of glycerol as well as Na⁺, K⁺ content under osmotic stress conditions has been proposed.     

Effect of pH on citrinin and red pigments production by Monascus purpureus CCT3802

The production of red pigments and citrinin by Monascus purpureus CCT3802 was investigated in submerged batch cultures performed in two phases: in the first phase, cells were grown on glucose, at pH 4.5, 5.5 or 6.5; after glucose depletion, pH was adjusted, when necessary, to 4.5, 5.5, 6.5, 7.0, 8.0 or 8.5, for a production phase. The highest total red pigments absorbance of 11.3 U was 16 times greater than the lowest absorbance and was achieved with growth at pH 5.5, followed by production at pH 8.5, which causes an immediate reduction of the intra cellular red pigments from 75% to 17% of the total absorbance. The lowest citrinin concentration, 5.5 mg L⁻¹, was verified in the same culture while the highest concentration, 55 mg L⁻¹, was verified in cultures entirely carried out at pH 5.5. An alkaline medium, besides promoting intra cellular red pigments excretion, strongly represses citrinin synthesis.     

Na+ requirement for glutamate-dependent sugar transport byFusobacterium nucleatum ATCC 10953

Resting cells ofFusobacterium nucleatum ATCC 10953, when provided with glutamic acid (Na⁺ salt) as fermentable energy source, rapidly accumulated [¹⁴C]glucose, from the medium. Sugar accumulation was not observed when Na⁺ glutamate was replaced by ammonium glutamate. However, addition of Na⁺ (chloride) to the latter system elicited uptake of [¹⁴C]glucose by the organism. Of other monovalent cations tested, only Li⁺ was found to be slightly stimulatory, but K⁺, Rb⁺, and Cs⁺ ions were ineffective. For determination of the role(s) of Na⁺ in sugar accumulation, the transport of [¹⁴C]glucose and [¹⁴C]glutamic acid by the cells was studied independently, with lysine as an alternate (and Na⁺-independent) energy source. In the presence of lysine, cells ofF. nucleatum 10953 accumulated [¹⁴C]glucose from a Na⁺-free medium, but, in contrast, uptake and fermentation of [¹⁴C]glutamic acid was Na⁺-dependent. The glucose transport system is Na⁺-independent. However, our data indicate dual role(s) for Na⁺ in the transport and intracellular metabolism of glutamic acid. The Na⁺-dependent glutamate fermentation pathway provides the necessary energy for active transport of glucose by the resting cell.