Isolation of bovine plasma albumin by liquid chromatography and its polymerization for use in immunohematology

The aim of the method described here is to remove hemoglobin, the major contaminant in the bovine plasma obtained from slaughterhouses, by adding a mixture of 19% cold ethanol and 0.6% chloroform, followed by fibrinogen and globulin precipitation by the Cohn method and nonspecific hemagglutinin by thermocoagulation. The experimental volume of bovine plasma was 2,000 ml per batch. Final purification was performed by liquid chromatography using the ion-exchange gel DEAE-Sepharose FF. The bovine albumin thus obtained presented > or = 99% purity, a yield of 25.0 ± 1.2 g/l plasma and >71.5% recovery. N-acetyl-DL-tryptophan (0.04 mmol/g protein) and sodium caprylate (0.04 mmol/g protein) were used as stabilizers and the final concentration of albumin was adjusted to 22.0% (w/v), pH 7.2 to 7.3. Viral inactivation was performed by pasteurization for 10 h at 60°C. The bovine albumin for the hemagglutination tests used in immunohematology was submitted to chemical treatment with 0.06% (w/v) glutaraldehyde and 0.1% (w/v) formaldehyde at 37°C for 12 h to obtain polymerization. A change in molecular distribution was observed after this treatment, with average contents of 56.0% monomers, 23.6% dimers, 12.2% trimers and 8.2% polymers. The tests performed demonstrated that this polymerized albumin enhances the agglutination of Rho(D)-positive red cells by anti-Rho(D) serum, permitting and improving visualization of the results.

Regulation of the renal proximal tubule second sodium pump by angiotensins

For several years it was believed that angiotensin II (Ang II) alone mediated the effects of the renin-angiotensin system. However, it has been observed that other peptides of this system, such as angiotensin-(1-7) (Ang-(1-7)), present biological activity. The effect of Ang II and Ang-(1-7) on renal sodium excretion has been associated, at least in part, with modulation of proximal tubule sodium reabsorption. In the present review, we discuss the evidence for the involvement of Na+-ATPase, called the second sodium pump, as a target for the actions of these compounds in the regulation of proximal tubule sodium reabsorption.

Reaction of diphenyl diselenide with hydrogen peroxide and inhibition of delta-aminolevulinate dehydratase from rat liver and cucumber leaves

The interaction of the product of H2O2 and (PhSe)2 with delta-aminolevulinate dehydratase (delta-ALA-D) from mammals and plants was investigated. (PhSe)2 inhibited rat hepatic delta-ALA-D with an IC50 of 10 µM but not the enzyme from cucumber leaves. The reaction of (PhSe)2 with H2O2 for 1 h increased the inhibitory potency of the original compound and the IC50 for animal delta-ALA-D inhibition was decreased from 10 to 2 µM. delta-ALA-D from cucumber leaves was also inhibited by the products of reaction of (PhSe)2 with H2O2 with an IC50 of 4 µM. The major product of reaction of (PhSe)2 with H2O2 was identified as seleninic acid and produced an intermediate with a lambdamax at 265 nm after reaction with t-BuSH. These results suggest that the interaction of (PhSe)2 with mammal delta-ALA-D requires the presence of cysteinyl residues in close proximity. Two cysteine residues in spatial proximity have been recently described for the mammalian enzyme. Analysis of the primary structure of plant delta-ALA-D did not reveal an analogous site. In contrast to (PhSe)2, seleninic acid, as a result of the higher electrophilic nature of its selenium atom, may react with additional cysteinyl residue(s) in mammalian delta-ALA-D and also with cysteinyl residues from cucumber leaves located at a site distinct from that found at the B and A sites in mammals. Although the interaction of organochalcogens with H2O2 may have some antioxidant properties, the formation of seleninic acid as a product of this reaction may increase the toxicity of organic chalcogens such as (PhSe)2.

Nitrergic modulation of vasopressin, oxytocin and atrial natriuretic peptide secretion in response to sodium intake and hypertonic blood volume expansion

The central nervous system plays an important role in the control of renal sodium excretion. We present here a brief review of physiologic regulation of hydromineral balance and discuss recent results from our laboratory that focus on the participation of nitrergic, vasopressinergic, and oxytocinergic systems in the regulation of water and sodium excretion under different salt intake and hypertonic blood volume expansion (BVE) conditions. High sodium intake induced a significant increase in nitric oxide synthase (NOS) activity in the medial basal hypothalamus and neural lobe, while a low sodium diet decreased NOS activity in the neural lobe, suggesting that central NOS is involved in the control of sodium balance. An increase in plasma concentrations in vasopressin (AVP), oxytocin (OT), atrial natriuretic peptide (ANP), and nitrate after hypertonic BVE was also demonstrated. The central inhibition of NOS by L-NAME caused a decrease in plasma AVP and no change in plasma OT or ANP levels after BVE. These data indicate that the increase in AVP release after hypertonic BVE depends on nitric oxide production. In contrast, the pattern of OT secretion was similar to that of ANP secretion, supporting the view that OT is a neuromodulator of ANP secretion during hypertonic BVE. Thus, neurohypophyseal hormones and ANP are secreted under hypertonic BVE in order to correct the changes induced in blood volume and osmolality, and the secretion of AVP in this particular situation depends on NOS activity.

Effects of hypertonic sodium chloride solution on the electrophysiologic alterations caused by bupivacaine in the dog heart

The effects of various hypertonic solutions on the intraventricular conduction, ventricular repolarization and the arrhythmias caused by the intravenous (iv) injection of bupivacaine (6.5 mg/kg) were studied in sodium pentobarbital-anesthetized mongrel dogs. Hypertonic solutions, given iv 5 min before bupivacaine, were 7.5% (w/v) NaCl, 5.4% (w/v) LiCl, 50% (w/v) glucose (2,400 mOsm/l, 5 ml/kg), or 20% (w/v) mannitol (1,200 mOsm/l, 10 ml/kg). Bupivacaine induced severe arrhythmias and ventricular conduction and repolarization disturbances, as reflected by significant increases in QRS complex duration, HV interval, IV interval and monophasic action potential duration, as well as severe hemodynamic impairment. Significant prevention against ventricular electrophysiologic and hemodynamic disturbances and ventricular arrhythmias was observed with 7.5% NaCl (percent increase in QRS complex duration: 164.4 ± 21.8% in the non-pretreated group vs 74.7 ± 14.1% in the pretreated group, P<0.05; percent increase in HV interval: 131.4 ± 16.1% in the non-pretreated group vs 58.2 ± 7.5% in the pretreated group, P<0.05; percent increase in monophasic action potential duration: 22.7 ± 6.8% in the non-pretreated group vs 9.8 ± 6.3% in the pretreated group, P<0.05; percent decrease in cardiac index: -46 ± 6% in the non-pretreated group vs -28 ± 5% in the pretreated group, P<0.05). The other three hypertonic solutions were ineffective. These findings suggest an involvement of sodium ions in the mechanism of hypertonic protection.

Oral administration of L-arginine decreases blood pressure and increases renal excretion of sodium and water in renovascular hypertensive rats

The two-kidney, one-clip renovascular (2K1C) hypertension model is characterized by a reduction in renal flow on the clipped artery that activates the renin-angiotensin system. Endothelium dysfunction, including diminished nitric oxide production, is also believed to play a role in the pathophysiology of this model. Some studies have shown an effect of L-arginine (L-Arg, a nitric oxide precursor) on hypertension. In the present study we determined the ability of L-Arg (7 days of treatment) to reduce blood pressure and alter renal excretions of water, Na+ and K+ in a model of 2K1C-induced hypertension. Under ether anesthesia, male Wistar rats (150-170 g) had a silver clip (0.20 mm) placed around the left renal artery to produce the 2K1C renovascular hypertension model. In the experimental group, the drinking water was replaced with an L-Arg solution (10 mg/ml; average intake of 300 mg/day) from the 7th to the 14th day after surgery. Sham-operated rats were used as controls. At the end of the treatment period, mean blood pressure was measured in conscious animals. The animals were then killed and the kidneys were removed and weighed. There was a significant reduction of mean blood pressure in the L-Arg-treated group when compared to control (129 ± 7 vs 168 ± 6 mmHg, N = 8-10 per group; P<0.05). Concomitantly, a significant enhancement of water and Na+ excretion was observed in the 2K1C L-Arg-treated group when compared to control (water: 13.0 ± 0.7 vs 9.2 ± 0.5 ml/day, P<0.01; Na+: 1.1 ± 0.05 vs 0.8 ± 0.05 mEq/day, respectively, P<0.01). These results show that orally administered L-Arg acts on the kidney, possibly inducing changes in renal hemodynamics or tubular transport due to an increase in nitric oxide formation.

Effect of intracerebroventricularly injected insulin on urinary sodium excretion by cerebroventricular streptozotocin-treated rats

Recent evidence suggests that insulin may influence many brain functions. It is known that intracerebroventricular (icv) injection of nondiabetogenic doses of streptozotocin (STZ) can damage insulin receptor signal transduction. In the present study, we examined the functional damage to the brain insulin receptors on central mechanisms regulating glomerular filtration rate and urinary sodium excretion, over four periods of 30 min, in response to 3 µl insulin or 0.15 NaCl (vehicle) injected icv in STZ-treated freely moving Wistar-Hannover rats (250-300 g). The icv cannula site was visually confirmed by 2% Evans blue infusion. Centrally administered insulin (42.0 ng/µl) increased the urinary output of sodium (from 855.6 ± 85.1 to 2055 ± 310.6 delta%/min; N = 11) and potassium (from 460.4 ± 100 to 669 ± 60.8 delta%/min; N = 11). The urinary sodium excretion response to icv insulin microinjection was markedly attenuated by previous central STZ (100 µg/3 µl) administration (from 628 ± 45.8 to 617 ± 87.6 delta%/min; N = 5) or by icv injection of a dopamine antagonist, haloperidol (4 µg/3 µl) (from 498 ± 39.4 to 517 ± 73.2 delta%/min; N = 5). Additionally, insulin-induced natriuresis occurred by increased post-proximal tubule sodium rejection, despite an unchanged glomerular filtration rate. Excluding the possibility of a direct action of STZ on central insulin receptor-carrying neurons, the current data suggest that the insulin-sensitive response may be processed through dopaminergic D1 receptors containing neuronal pathways.

Effect of electrolytic lesion of the dorsal raphe nucleus on water intake and sodium appetite

The present study determined the effect of an electrolytic lesion of the dorsal raphe nucleus (DRN) on water intake and sodium appetite. Male Wistar rats weighing 290-320 g with a lesion of the DRN (L-DRN), performed two days before experiments and confirmed by histology at the end of the experiments, presented increased sensitivity to the dehydration induced by fluid deprivation. The cumulative water intake of L-DRN rats reached 23.3 ± 1.9 ml (a 79% increase, N = 9) while sham-lesioned rats (SL-DRN) did not exceed 13.0 ± 1.0 ml (N = 11, P < 0.0001) after 5 h. The L-DRN rats treated with isoproterenol (300 µg kg-1 ml-1, sc) exhibited an increase in water intake that persisted throughout the experimental period (L-DRN, 15.7 ± 1.47 ml, N = 9 vs SL-DRN, 9.3 ± 1.8 ml, N = 11, P < 0.05). The L-DRN rats also showed an increased spontaneous sodium appetite during the entire period of assessment. The intake of 0.3 M NaCl after 12, 24, 36 and 72 h by the L-DRN rats was always higher than 20.2 ± 4.45 ml (N = 10), while the intake by SL-DRN was always lower than 2.45 ± 0.86 ml (N = 10, P < 0.00001). Sodium- and water-depleted L-DRN rats also exhibited an increased sodium appetite (13.9 ± 2.0 ml, N = 11) compared to SL-DRN (4.6 ± 0.64 ml, N = 11) after 120 min of observation (P < 0.02). The sodium preference of L-DRN rats in both conditions was always higher than that of SL-DRN rats. These results suggest that electrolytic lesion of the DRN overcomes a tonic inhibitory component of sodium appetite.

Cytotoxicity of chlorhexidine digluconate to murine macrophages and its effect on hydrogen peroxide and nitric oxide induction

Chlorhexidine, even at low concentrations, is toxic for a variety of eukaryotic cells; however, its effects on host immune cells are not well known. We evaluated in vitro chlorhexidine-induced cytotoxicity and its effects on reactive oxygen/nitrogen intermediate induction by murine peritoneal macrophages. Thioglycollate-induced cells were obtained from Swiss mice by peritoneal lavage with 5 ml of 10 mM phosphate-buffered saline, washed twice and resuspended (10(6) cells/ml) in appropriate medium for each test. Cell preparations contained more than 95% macrophages. The cytotoxicity was determined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide assay and the presence of hydrogen peroxide (H2O2) and nitric oxide (NO) by the horseradish peroxidase-dependent oxidation of phenol red and Griess reaction, respectively. The midpoint cytotoxicity values for 1- and 24-h exposures were 61.12 ± 2.46 and 21.22 ± 2.44 µg/ml, respectively. Chlorhexidine did not induce synthesis or liberation of reactive oxygen/nitrogen intermediates. When macrophages were treated with various sub-toxic doses for 1 h (1, 5, 10, and 20 µg/ml) and 24 h (0.5, 1, and 5 µg/ml) and stimulated with 200 nM phorbol myristate acetate (PMA) solution, the H2O2 production was not altered; however, the NO production induced by 10 µg/ml lipopolysaccharide (LPS) solution varied from 14.47 ± 1.46 to 22.35 ± 1.94 µmol/l and 13.50 ± 1.42 to 20.44 ± 1.40 µmol/l (N = 5). The results showed that chlorhexidine has no immunostimulating activity and sub-toxic concentrations did not affect the response of macrophages to the soluble stimulus PMA but can interfere with the receptor-dependent stimulus LPS.

Vedyn käytön tarkastelu Kemira Chemicals Oy:n Joutsenon tehtailla

Kemira Chemicals Oy:n Joutsenon tehtailla valmistetaan lipeää, suolahappoa, natriumhypokloriittia sekä natriumkloraattia. Lipeää, suolahappoa ja natriumhypokloriittia valmistetaan lipeätehtaassa. Natriumkloraattia valmistetaan kloraattitehtaassa. Kloraatti- ja lipeätehtaan tuotteet valmistetaan elektrolyysimenetelmällä. Elektrolyysien sivutuotteena syntyy vetykaasua, joka voidaan käyttää suolahapon valmistukseen, vetyvoimalaitoksen polttoaineena tai myydä asiakkaalle. Työn tavoitteena oli tarkastella vedyn käyttöä Joutsenon tehtailla. Tarkastelun tavoitteena oli löytää mahdollisia kehitys- tai jatkotutkimuskohteita vety- ja höyryjärjestelmästä. Koska vetyä käytetään myös vetyvoimalaitoksen polttoaineena, joka tuottaa tehtailla tarvittavan prosessihöyryn, tarkasteltiin työssä myös höyryn käyttöä tehtailla. Tarkastelua varten tehtiin Microsoft Excel-pohjainen taselaskentamalli, jolla simuloitiin vedyn ja höyryn käyttöä tehtailla. Työn tuloksena saatiin Excel-pohjainen simulointimalli, jolla pystyttiin tutkimaan vedyn ja höyryn käyttöä. Vedyn ja höyryn käyttöä tutkittiin viidessä eri skenaariossa. Skenaariossa yksi määritettiin pienimmät mahdolliset elektrolyysiin syötettävät sähkövirran arvot, joilla tehtaita on turvallista käyttää. Skenaariossa kaksi määritettiin pienimmät mahdolliset elektrolyysiin syötettävät sähkövirran arvot, joilla voimalaitoksen turbiini pysyisi ajossa. Skenaariossa kolme määritettiin tehtaiden tämän hetkinen maksimi kapasiteetti. Skenaarioissa neljä ja viisi tutkittiin, miten mahdollinen tehtaiden tuotantojen kasvattaminen vaikuttaisi vety- ja höyryjärjestelmään. Työn tuloksien perusteella kehitys- ja jatkotutkimuskohteita olisivat lipeän haihdutuksen höyryn kulutuksen pienentäminen, turbiinin käyttöajan kasvattaminen sekä eri lähteistä saatavan hukkalämmön parempi hyödyntäminen kaukolämmön tuotannossa. Tehtaiden tuotantoja kasvatettaessa on syytä kiinnittää huomioita myös voimalaitoksen pääkattilan ja turbiinin kapasiteettiin.

Chronic excitotoxic lesion of the dorsal raphe nucleus induces sodium appetite

We determined if the dorsal raphe nucleus (DRN) exerts tonic control of basal and stimulated sodium and water intake. Male Wistar rats weighing 300-350 g were microinjected with phosphate buffer (PB-DRN, N = 11) or 1 µg/0.2 µl, in a single dose, ibotenic acid (IBO-DRN, N = 9 to 10) through a guide cannula into the DRN and were observed for 21 days in order to measure basal sodium appetite and water intake and in the following situations: furosemide-induced sodium depletion (20 mg/kg, sc, 24 h before the experiment) and a low dose of dietary captopril (1 mg/g chow). From the 6th day after ibotenic acid injection IBO-DRN rats showed an increase in sodium appetite (12.0 ± 2.3 to 22.3 ± 4.6 ml 0.3 M NaCl intake) whereas PB-DRN did not exceed 2 ml (P < 0.001). Water intake was comparable in both groups. In addition to a higher dipsogenic response, sodium-depleted IBO-DRN animals displayed an increase of 0.3 M NaCl intake compared to PB-DRN (37.4 ± 3.8 vs 21.6 ± 3.9 ml 300 min after fluid offer, P < 0.001). Captopril added to chow caused an increase of 0.3 M NaCl intake during the first 2 days (IBO-DRN, 33.8 ± 4.3 and 32.5 ± 3.4 ml on day 1 and day 2, respectively, vs 20.2 ± 2.8 ml on day 0, P < 0.001). These data support the view that DRN, probably via ascending serotonergic system, tonically modulates sodium appetite under basal and sodium depletion conditions and/or after an increase in peripheral or brain angiotensin II.

Effect of intraperitoneally administered hydrolyzed whey protein on blood pressure and renal sodium handling in awake spontaneously hypertensive rats

The present study evaluated the acute effect of the intraperitoneal (ip) administration of a whey protein hydrolysate (WPH) on systolic arterial blood pressure (SBP) and renal sodium handling by conscious spontaneously hypertensive rats (SHR). The ip administration of WPH in a volume of 1 ml dose-dependently lowered the SBP in SHR 2 h after administration at doses of 0.5 g/kg (0.15 M NaCl: 188.5 ± 9.3 mmHg vs WPH: 176.6 ± 4.9 mmHg, N = 8, P = 0.001) and 1.0 g/kg (0.15 M NaCl: 188.5 ± 9.3 mmHg vs WPH: 163.8 ± 5.9 mmHg, N = 8, P = 0.0018). Creatinine clearance decreased significantly (P = 0.0084) in the WPH-treated group (326 ± 67 µL min-1 100 g body weight-1) compared to 0.15 M NaCl-treated (890 ± 26 µL min-1 100 g body weight-1) and captopril-treated (903 ± 72 µL min-1 100 g body weight-1) rats. The ip administration of 1.0 g WPH/kg also decreased fractional sodium excretion to 0.021 ± 0.019% compared to 0.126 ± 0.041 and 0.66 ± 0.015% in 0.15 M NaCl and captopril-treated rats, respectively (P = 0.033). Similarly, the fractional potassium excretion in WPH-treated rats (0.25 ± 0.05%) was significantly lower (P = 0.0063) than in control (0.91 ± 0.15%) and captopril-treated rats (1.24 ± 0.30%), respectively. The present study shows a decreased SBP in SHR after the administration of WPH associated with a rise in tubule sodium reabsorption despite an angiotensin I-converting enzyme (ACE)-inhibiting in vitro activity (IC50 = 0.68 mg/mL). The present findings suggest a pathway involving ACE inhibition but measurements of plasma ACE activity and angiotensin II levels are needed to support this suggestion.

Review of water electrolysis technologies and design of renewable hydrogen production systems

An electric system based on renewable energy faces challenges concerning the storage and utilization of energy due to the intermittent and seasonal nature of renewable energy sources. Wind and solar photovoltaic power productions are variable and difficult to predict, and thus electricity storage will be needed in the case of basic power production. Hydrogen’s energetic potential lies in its ability and versatility to store chemical energy, to serve as an energy carrier and as feedstock for various industries. Hydrogen is also used e.g. in the production of biofuels. The amount of energy produced during hydrogen combustion is higher than any other fuel’s on a mass basis with a higher-heating-value of 39.4 kWh/kg. However, even though hydrogen is the most abundant element in the universe, on Earth most hydrogen exists in molecular forms such as water. Therefore, hydrogen must be produced and there are various methods to do so. Today, the majority hydrogen comes from fossil fuels, mainly from steam methane reforming, and only about 4 % of global hydrogen comes from water electrolysis. Combination of electrolytic production of hydrogen from water and supply of renewable energy is attracting more interest due to the sustainability and the increased flexibility of the resulting energy system. The preferred option for intermittent hydrogen storage is pressurization in tanks since at ambient conditions the volumetric energy density of hydrogen is low, and pressurized tanks are efficient and affordable when the cycling rate is high. Pressurized hydrogen enables energy storage in larger capacities compared to battery technologies and additionally the energy can be stored for longer periods of time, on a time scale of months. In this thesis, the thermodynamics and electrochemistry associated with water electrolysis are described. The main water electrolysis technologies are presented with state-of-the-art specifications. Finally, a Power-to-Hydrogen infrastructure design for Lappeenranta University of Technology is presented. Laboratory setup for water electrolysis is specified and factors affecting its commissioning in Finland are presented.

Value at Risk in Foreign Exchange Risk Management

This thesis examines the suitability of VaR in foreign exchange rate risk management from the perspective of a European investor. The suitability of four different VaR models is evaluated in respect to have insight if VaR is a valuable tool in managing foreign exchange rate risk. The models evaluated are historical method, historical bootstrap method, variance-covariance method and Monte Carlo simulation. The data evaluated are divided into emerging and developed market currencies to have more intriguing analysis. The foreign exchange rate data in this thesis is from 31st January 2000 to 30th April 2014. The results show that the previously mentioned VaR models performance in foreign exchange risk management is not to be considered as a single tool in foreign exchange rate risk management. The variance-covariance method and Monte Carlo simulation performs poorest in both currency portfolios. Both historical methods performed better but should also be considered as an additional tool along with other more sophisticated analysis tools. A comparative study of VaR estimates and forward prices is also included in the thesis. The study reveals that regardless of the expensive hedging cost of emerging market currencies the risk captured by VaR is more expensive and thus FX forward hedging is recommended

Mechanisms underlying gas exchange alterations in an experimental model of pulmonary embolism

The aim of the present study was to determine the ventilation/perfusion ratio that contributes to hypoxemia in pulmonary embolism by analyzing blood gases and volumetric capnography in a model of experimental acute pulmonary embolism. Pulmonary embolization with autologous blood clots was induced in seven pigs weighing 24.00 ± 0.6 kg, anesthetized and mechanically ventilated. Significant changes occurred from baseline to 20 min after embolization, such as reduction in oxygen partial pressures in arterial blood (from 87.71 ± 8.64 to 39.14 ± 6.77 mmHg) and alveolar air (from 92.97 ± 2.14 to 63.91 ± 8.27 mmHg). The effective alveolar ventilation exhibited a significant reduction (from 199.62 ± 42.01 to 84.34 ± 44.13) consistent with the fall in alveolar gas volume that effectively participated in gas exchange. The relation between the alveolar ventilation that effectively participated in gas exchange and cardiac output (V Aeff/Q ratio) also presented a significant reduction after embolization (from 0.96 ± 0.34 to 0.33 ± 0.17 fraction). The carbon dioxide partial pressure increased significantly in arterial blood (from 37.51 ± 1.71 to 60.76 ± 6.62 mmHg), but decreased significantly in exhaled air at the end of the respiratory cycle (from 35.57 ± 1.22 to 23.15 ± 8.24 mmHg). Exhaled air at the end of the respiratory cycle returned to baseline values 40 min after embolism. The arterial to alveolar carbon dioxide gradient increased significantly (from 1.94 ± 1.36 to 37.61 ± 12.79 mmHg), as also did the calculated alveolar (from 56.38 ± 22.47 to 178.09 ± 37.46 mL) and physiological (from 0.37 ± 0.05 to 0.75 ± 0.10 fraction) dead spaces. Based on our data, we conclude that the severe arterial hypoxemia observed in this experimental model may be attributed to the reduction of the V Aeff/Q ratio. We were also able to demonstrate that V Aeff/Q progressively improves after embolization, a fact attributed to the alveolar ventilation redistribution induced by hypocapnic bronchoconstriction.