In Vitro Percutaneous Permeation and Skin Accumulation of Finasteride Using Vesicular Ethosomal Carriers

In order to develop a novel transdermal drug delivery system that facilitates the skin permeation of finasteride encapsulated in novel lipid-based vesicular carriers (ethosomes)finasteride ethosomes were constructed and the morphological characteristics were studied by transmission electron microscopy. The particle size, zeta potential and the entrapment capacity of ethosome were also determined. In contrast to liposomes ethosomes were of more condensed vesicular structure and they were found to be oppositely charged. Ethosomes were found to be more efficient delivery carriers with high encapsulation capacities. In vitro percutaneous permeation experiments demonstrated that the permeation of finasteride through human cadaver skin was significantly increased when ethosomes were used. The finasteride transdermal fluxes from ethosomes containing formulation (1.34 ± 0.11 μg/cm²/h) were 7.4, 3.2 and 2.6 times higher than that of finasteride from aqueous solution, conventional liposomes and hydroethanolic solution respectively (P < 0.01).Furthermore, ethosomes produced a significant (P < 0.01) finasteride accumulation in the skin, especially in deeper layers, for instance in dermis it reached to 18.2 ± 1.8 μg/cm². In contrast, the accumulation of finasteride in the dermis was only 2.8 ± 1.3 μg/cm² with liposome formulation. The study demonstrated that ethosomes are promising vesicular carriers for enhancing percutaneous absorption of finasteride.     

Design and development of ethosomal transdermal drug delivery system of valsartan with preclinical assessment in Wistar albino rats

Abstract

Valsartan (VLT) is a highly selective and orally active antihypertensive drug. However, its oral administration is associated with drawbacks like low bioavailability. The objective of this study was to design and develop a transdermal delivery system for VLT using ethosomal carriers to investigate their enhanced transdermal delivery potential. VLT ethosomes were prepared by cold method. VLT ethosomes were characterized by scanning electron microscopy. The prepared ethanolic liposomes were characterized to be spherical having low polydispersity of nano-size range with good entrapment efficiency. ETC5 ethosomal suspension with 4% of phospholipon 90H and 40% of ethanol was found to have highest entrapment efficiency, i.e. 80.230?±?0.8748%. The permeation study of ethosomes was evaluated by ex vivo diffusion study through rat abdominal skin using Franz’s diffusion cells and ETC5 ethosomal suspension was found to have highest permeation with flux of 92.819?±?1.539?µg/cm²/h, when compared to the permeation profiles of drug solutions either in water or in a water–ethanol mixture. Transdermal application of ethosomal VLT on Wistar rats showed better and prolonged antihypertensive activity in comparison to orally administered VLT suspension by virtue of transdermal permeation through Wistar rat skin. Histopathological study of skin applied with ETC5 showed intercellular permeation across skin by dissolving intercellular lipids in epidermis without causing any rigorous changes in the skin cellular structure. In conclusion, ethosomes enabled the transdermal permeation of VLT, which amply proves its superiority over oral administration for antihypertensive treatment.     

Microemulsion-based gel of terbinafine for the treatment of onychomycosis: optimization of formulation using D-optimal design

The aim of the present investigation was to evaluate microemulsion as a vehicle for dermal drug delivery and to develop microemulsion-based gel of terbinafine for the treatment of onychomycosis. D-optimal mixture experimental design was adopted to optimize the amount of oil (X ₁), Smix (mixture of surfactant and cosurfactant; X ₂) and water (X ₃) in the microemulsion. The formulations were assessed for globule size (in nanometers; Y ₁) and solubility of drug in microemulsion (in milligrams per milliliter; Y ₂). The microemulsion containing 5.75% oil, 53.75% surfactant–cosurfactant mixture and 40.5% water was selected as the optimized batch. The globule size and solubility of the optimized batch were 18.14 nm and 43.71 mg/ml, respectively. Transmission electron microscopy showed that globules were spherical in shape. Drug containing microemulsion was converted into gel employing 0.75% w/w carbopol 934P. The optimized gel showed better penetration and retention in the human cadaver skin as compared to the commercial cream. The cumulative amount of terbinafine permeated after 12 h was 244.65 ± 18.43 μg cm⁻² which was three times more than the selected commercial cream. Terbinafine microemulsion in the gel form showed better activity against Candida albicans and Trichophyton rubrum than the commercial cream. It was concluded that drug-loaded gel could be a promising formulation for effective treatment of onychomycosis.     

Factorial design based curcumin ethosomal nanocarriers for the skin cancer delivery: in vitro evaluation

Abstract

Melanoma is the most deadly and life-threatening form of skin cancer with progressively higher rates of incidence worldwide. The objective of the present investigation is to develop and to statistically optimize and characterize curcumin (CUR) loaded ethosomes for treatment of melanoma. A two factor, three level (3²) factorial design approach was employed for the optimization of ethosomes. The prepared ethosomes were evaluated for size, zeta potential, entrapment efficiency, in vitro skin permeation and deposition ability. The optimized ethosomal formulation was evaluated for in vitro cytotoxicity and cellular uptake studies using A375 human melanoma cells. The optimized formulation has imperfect round shaped unilamellar structures with a mean vesicle size of 247?±?5.25?nm and an entrapment efficiency of 92.24?±?0.20%. The in vitro skin permeation studies proved the superiority of ethosomes over the traditional liposomes in terms of the amount of drug permeated and deposited in skin layers. Fluorescence microscopy showed the enhanced penetration of ethosomes into the deeper layers of the skin. In vitro cytotoxicity and cellular uptake studies revealed that curcumin ethosomes have significantly improved cytotoxicity and cellular uptake in A375 human melanoma cell lines. The colony formation assay results showed that curcumin ethosomes have a superior antiproliferative effect as they effectively inhibit the clonogenic ability of A375 cells. The flow cytometry results indicate that curcumin ethosomes induce cell death in A375 cells by apoptosis mechanism. The present study provides a strong rationale and motivation for further investigation of newly developed curcumin ethosomes as a potential therapeutic strategy for melanoma treatment.     

Transdermal absorption enhancement of rice bran bioactive compounds entrapped in niosomes

Niosomes composed of Tween 61 and cholesterol at 1:1 molar ratio were entrapped with the mixture of the three semi-purified rice (Oryza sativa L., Family Gramineae) bran bioactive compounds [ferulic acid (F), γ-oryzanol (O), and phytic acid (P)] at 0.5%, 1.5%, and 1.5%, respectively, by the supercritical CO₂ technique. The transdermal absorption by vertical Franz diffusion cells of the compounds entrapped in niosomes (Nio FOP), the unentrapped compounds (Mixed FOP), the compounds incorporated in gel and cream (Gel FOP and Cream FOP), and the compounds entrapped in niosomes and incorporated in gel and cream (Gel nio and Cream nio) was investigated. At 6 h, F and P from Nio FOP gave lower cumulative amount in viable epidermis and dermis (VED) than from Mixed FOP of 1.1 and 1.6 times, respectively, while O from Nio FOP exhibited higher cumulative amount in VED than from Mixed FOP of 2.4 times. The highest cumulative amount in VED of F, O, and P were from Gel nio, Cream nio, and Mixed FOP at 1.564 ± 0.052, 15.972 ± 0.273, and 25.857 ± 0.025 ng/cm², respectively. Niosomes enhanced the transdermal absorption of the hydrophobic compound O, while retarded the hydrophilic compounds F and P indicating the less systemic risk of F and P than O when entrapped in niosomes. Thus, transdermal absorption of F, O, and P appeared to depend on niosomal size, lipophilicity of the bioactive compounds, and types of formulations. These preclinical results can be applied for the design of the clinical study of the developed rice bran niosomal topical products.     

NMR Reinvestigation of the Caffeine–Chlorogenate Complex in Aqueous Solution and in Coffee Brews

Caffeine complexation by chlorogenic acid (3-caffeoylquinic acid, CAS Number [327-97-9]) in aqueous solution as well as caffeine–chlorogenate complex in freshly prepared coffee brews have been investigated by high-resolution ¹H-NMR. Caffeine and chlorogenic acid self-associations have also been studied and self-association constants have been determined resorting to both classical isodesmic model and a recently introduced method of data analysis able to provide also the critical aggregation concentration (cac). Furthermore, caffeine–chlorogenate association constant was measured. For the caffeine, the average value of the self-association constant determined by isodesmic model (K _i = 7.6 ± 0.5 M⁻¹) is in good agreement with the average value (K _a = 10 ± 1.8 M⁻¹) determined with the method which permits the determination of the cac (8.43 ± 0.05 mM). Chlorogenic acid shows a slight decreased tendency to aggregation with a lower average value of association constants (K _i = 2.8 ± 0.6 M⁻¹; K _a = 3.4 ± 0.6 M⁻¹) and a critical concentration equal to 24 ± 1 mM. The value of the association constant of the caffeine–chlorogenate complex (30 ± 4 M⁻¹) is compatible with previous studies and within the typical range of reported association constants for other caffeine–polyphenol complexes. Structural features of the complex have also been investigated, and the complex conformation has been rediscussed. Caffeine chemical shifts comparison (monomeric, complexed, coffee brews) clearly indicates a significant amount of caffeine is complexed in beverage real system, being chlorogenate ions the main complexing agents.     

Growth and survival performance of the gametophyte of Gigartina skottsbergii (Rhodophyta, Gigartinales) under defined nutrient conditions in laboratory culture

Gigartina skottsbergii is a commercially important carrageenan producer that has been suffering severe extraction pressure in Chile’s Magellan Region and Cape Horn Archipelago since 1998. In order to create baseline information for its cultivation and repopulation, we studied the effects of agricultural fertilizers on growth of G. skottsbergii early developmental stages. The culture media utilized were: a) seawater + Bayfoland, b) seawater + Superphosphate, c) seawater + Urea, d) seawater + Provasoli and e) seawater as a control. The culture conditions were: a) 12L:12D photoperiod; b) temperature 8 ± 1°C and c) irradiance at 45 μmol photons m⁻² s⁻¹. After 60 days, higher relative growth rates between treatments were observed; the treatments that included Bayfoland and Provasoli showed greater growth (382 ± 55 and 378 ± 50 μm, respectively,) compared to Superphosphate (88 ± 16 μm), control (78 ± 10 μm) and Urea (70 ± 11 μm) treatments, after 81 days. The Urea treatment and the control had inhibitory effects on G. skottsbergii germlings growth and survival, as evidenced by progressive loss of pigmentation and death after 60 days. These results showed that Bayfoland was an excellent alternative to develop cultures.     

Heterologous expression and characterization of Choline Oxidase from the soil bacterium Arthrobacter nicotianae

In the course of a microbial screening of soil samples for new oxidases, different enrichment strategies were carried out. With choline as the only carbon source, a microorganism was isolated and identified as Arthrobacter nicotianae. From this strain, a gene coding for a choline oxidase was isolated from chromosomal DNA. This gene named codA was cloned in Escherichia coli BL21-Gold and the protein (An_CodA) heterologously overexpressed as a soluble intracellular protein of 59.1 kDa. Basic biochemical characterization of purified protein revealed a pH optimum of 7.4 and activity over a broad temperature range (15–70 °C). Specific activities were determined toward choline chloride (4.70 ± 0.12 U/mg) and the synthetic analogs bis(2-hydroxyethyl)-dimethylammonium chloride (0.05 ± 0.45 × 10^–2 U/mg) and tris-(2-hydroxyethyl)-methylammonium methylsulfate (0.01 ± 0.12 × 10^–2 U/mg). With increasing number of oxidizable groups, a significant decrease in activity was noted. Determination of kinetic parameters in atmorspheric oxygen resulted in K _M = 1.51 ± 0.09 mM and V _max = 42.73 ± 0.42 mU/min for choline chloride and K _M = 4.77 ± 0.76 mM and V _max = 48.40 ± 2.88 mU/min for the reaction intermediate betaine aldehyde respectively. Nuclear magnetic resonance spectroscopic analysis of the products formed during the enzyme reaction with choline chloride showed that in vitro the intermediate betaine aldehyde exists also free in solution.     

A new manganese superoxide dismutase identified from Beauveria bassiana enhances virulence and stress tolerance when overexpressed in the fungal pathogen

A superoxide dismutase (SOD) was characterized from Beauveria bassiana, a fungal entomopathogen widely applied to insect control. This 209-aa enzyme (BbSod2) showed no more than 71% sequence identity to other fungal Mn-SODs, sharing all conserved residues with the Mn-SOD family and lacking a mitochondrial signal. The SOD activity of purified BbSod2 was significantly elevated by Mn²⁺, suppressed by Cu²⁺ and Zn²⁺ but inhibited by Fe³⁺. Overexpressing the enzyme in a BbSod2-absent B. bassiana strain enhanced its SOD activity (107.2 ± 6.1 U mg⁻¹ protein) by 4–10-fold in different transformants analyzed. The best BbSod2-transformed strain with the SOD activity of 1,157.9 ± 74.7 U mg⁻¹ was 93% and 61% more tolerant to superoxide-generating menadione in both colony growth (EC₅₀ = 2.41 ± 0.03 versus 1.25 ± 0.01 mM) and conidial germination (EC₅₀ = 0.89 ± 0.06 versus 0.55 ± 0.07 mM), and 23% more tolerant to UV-B irradiation (LD₅₀ = 0.49 ± 0.02 versus 0.39 ± 0.01 J cm⁻²). Its virulence to Spodoptera litura larvae was enhanced by 26% [LT₅₀ = 4.5 (4.2–4.8) versus 5.7 (5.2–6.4) days]. Our study highlights for the first time that the Mn²⁺-cofactored, cytosolic BbSod2 contributes significantly to the virulence and stress tolerance of B. bassiana and reveals possible means to improving field persistence and efficacy of a fungal formulation by manipulating the antioxidant enzymes of a candidate strain.     

Superior quality agar from <Emphasis Type="Italic">Gracilaria</Emphasis> species (Gracilariales,Rhodophyta) collected from the Gulf of Mannar,India

Gracilaria edulis, G. crassa, G. foliifera, and G. corticata are naturally occurring agarophytes of Indian waters. These agarophytes were evaluated for their agar contents using an improved process recently reported by us (US Patent 2005/0267296A1). The effect of different concentrations of NaOH in the alkali treatment was studied for optimizing the extraction conditions. These Gracilaria species of Indian waters produced agars, both native and alkali treated, with different properties confirming the heterogeneity of the agar polymers in this genera, as one would expect. Among these, G. edulis and G. crassa produced agar polymers having high gel strengths of 490 ± 8.16 and 800 ± 15.4 g cm⁻², respectively, with 8% NaOH treatment as opposed the low gel strength agars that have been reported in the literature to date.     

Density of tiger and leopard in a tropical deciduous forest of Mudumalai Tiger Reserve,southern India,as estimated using photographic capture–recapture sampling

Density of tiger Panthera tigris and leopard Panthera pardus was estimated using photographic capture–recapture sampling in a tropical deciduous forest of Mudumalai Tiger Reserve, southern India, from November 2008 to February 2009. A total of 2,000 camera trap nights for 100 days yielded 19 tigers and 29 leopards within an intensive sampling area of 107 km². Population size of tiger from closed population estimator model M_b Zippin was 19 tigers (SE = ±0.9) and for leopards M_h Jackknife estimated 53 (SE = ±11) individuals. Spatially explicit maximum likelihood and Bayesian model estimates were 8.31 (SE = ±2.73) and 8.9 (SE = ±2.56) per 100 km² for tigers and 13.17 (SE = ±3.15) and 13.01 (SE = ±2.31) per 100 km² for leopards, respectively. Tiger density for MMDM models ranged from 6.07 (SE = ±1.74) to 9.72 (SE = ±2.94) per 100 km² and leopard density ranged from 13.41 (SE = ±2.67) to 28.91 (SE = ±7.22) per 100 km². Spatially explicit models were more appropriate as they handle information at capture locations in a more specific manner than some generalizations assumed in the classical approach. Results revealed high density of tiger and leopard in Mudumalai which is unusual for other high density tiger areas. The tiger population in Mudumalai is a part of the largest population at present in India and a source for the surrounding Reserved Forest.     

Permeation and Distribution of Ferulic Acid and Its α-Cyclodextrin Complex from Different Formulations in Hairless Rat Skin

Ferulic acid (FA) is a natural product that occurs in seeds of many plants where it is generally located in the bran. This compound is a multifunctional ingredient endowed with antioxidative, radical scavenging, sunscreening and antibacterial actions. The aim of this study was to analyse the ferulic acid cutaneous permeation and distribution, through and into the skin layers, from different cosmetic vehicles, an O/W emulsion (pH 6.0) and two gel-type formulations at different pH levels (6.0 and 7.4), containing FA alone or an inclusion complex with α-cyclodextrin (CD-FA). In vitro permeation studies were performed in vertical diffusion cells using hairless rat excised skin. At appropriate intervals of time, the amount of permeated sunscreen/radical scavenger was evaluated by high-performance liquid chromatography (HPLC). At the end of experiments, treated skin samples were sectioned with a cryomicrotome and the FA content of the individual slices was analysed by HPLC. FA-containing formulations, O/W emulsion, gels A and B, originated FA fluxes of 8.48 ± 2.31, 8.38 ± 0.89 and 5.72 ± 0.50 μg/cm² h, respectively, thus suggesting the pH influence on FA percutaneous permeation. The use of the inclusion complex, CD-FA, determined in all cases a decrease of FA transdermal permeation while no influence of pH was observed. Gel-type formulations containing FA ensured higher sunscreen storage in the superficial layers if compared with O/W emulsion. When FA was included in α-cyclodextrin, FA amount retained into skin layers decreased markedly.     

In vitro studies on active calcium absorption from ovine rumen

From various in vivo and in vitro studies it has been shown that the rumen represents a significant site of Ca²⁺ absorption in sheep and goats. It was the aim of the present study to further characterize the underlying mechanisms. Unidirectional flux rates of Ca²⁺ across rumen wall epithelia of sheep were measured in vitro by applying the Ussing-chamber technique in the absence of electrochemical gradients. Under these conditions, significant Ca²⁺ net flux rates (J_net) clearly indicate the presence of active mechanisms for Ca²⁺ transport. Short chain fatty acids (SCFAs) caused highest stimulation of Ca²⁺ J_net (6.3 ± 1.9 nmol · cm⁻² · h⁻¹) when used as a mixture of acetate, proprionate and butyrate in physiological proportions (36, 15, 9 mmol · l⁻¹, respectively). The effect of 30 mmol · l⁻¹ butyrate (3.2 ± 0.6 nmol · cm⁻² · h⁻¹) was higher than respective amounts of propionate and acetate (0.6 ± 0.8 nmol · cm⁻² · h⁻¹ and 0.9 ± 0.8 nmol · cm⁻² · h⁻¹, respectively). Eliminating SCFAs resulted in Ca²⁺ J_net of 0.4 ± 1.1 nmol ^. cm^−2 .h⁻¹. Addition of Ca channel blocker verapamil (mucosal 1 mmol · l⁻¹) had no significant effect on SCFA-stimulated J_net of Ca²⁺, whereas application of Na⁺/H⁺ inhibitor amiloride (mucosal 1 mmol · l⁻¹) further enhanced the Ca²⁺ J_net by >65%. The Ca²⁺-pump inhibitor vanadate had no significant effect on J_net of Ca²⁺. Dietary Ca depletion enhanced calcitriol plasma concentrations but had no effect on active Ca²⁺ absorption across the rumen wall of sheep. In addition, no effect on active Ca²⁺ absorption could be observed during early lactation. In conclusion, there is clear evidence for the rumen as a main site for active Ca²⁺ absorption in sheep. Our results suggest the presence of a Ca²⁺/H⁺ exchange mechanism in the apical membrane of rumen epithelial cells which depends on SCFA absorption and which does not seem to be under the control of calcitriol. Basolateral Ca²⁺ extrusion occurs independently from Ca²⁺-pump activity and may be accomplished via Na⁺/Ca²⁺ exchange. Accepted: 29 June 1999     

Development,Optimization, and Anti-diabetic Activity of Gliclazide-Loaded Alginate–Methyl Cellulose Mucoadhesive Microcapsules

The purpose of this work was to develop and optimize gliclazide-loaded alginate–methyl cellulose mucoadhesive microcapsules by ionotropic gelation using central composite design. The effect of formulation parameters like polymer blend ratio and cross-linker (CaCl₂) concentration on properties of gliclazide-loaded alginate–methyl cellulose microcapsules like drug encapsulation efficiency and drug release were optimized. The optimized microcapsules were subjected to swelling, mucoadhesive, and in vivo studies. The observed responses coincided well with the predicted values from the optimization technique. The optimized microcapsules showed high drug encapsulation efficiency (83.57 ± 2.59% to 85.52 ± 3.07%) with low T _50% (time for 50% drug release, 5.68 ± 0.09 to 5.83 ± 0.11 h). The in vitro drug release pattern from optimized microcapsules was found to be controlled-release pattern (zero order) with case II transport release mechanism. Particle sizes of these optimized microcapsules were 0.767 ± 0.085 to 0.937 ± 0.086 mm. These microcapsules also exhibited good mucoadhesive properties. The in vivo studies on alloxan-induced diabetic rats indicated the significant hypoglycemic effect that was observed 12 h after oral administration of optimized mucoadhesive microcapsules. The developed and optimized alginate–methyl cellulose microcapsules are suitable for prolonged systemic absorption of gliclazide to maintain lower blood glucose level and improved patient compliance.     

Nanocarrier for the Transdermal Delivery of an Antiparkinsonian Drug

The purpose of the present study was to investigate the potential of nanoemulsions as nanodrug carrier systems for the percutaneous delivery of ropinirole. Nanoemulsions comprised Capryol 90 as the oil phase, Tween 20 as the surfactant, Carbitol as the cosurfactant, and water as an external phase. The effects of composition of nanoemulsion, including the ratio of surfactant and cosurfactant (S _mix) and their concentration on skin permeation, were evaluated. All the prepared nanoemulsions showed a significant increase in permeation parameters such as steady state flux (J _ss) and permeability coefficient (K _p) when compared to the control (p < 0.01). Nanoemulsion composition (NEL3) comprising ropinirole (0.5% w/w), Capryol 90 (5% w/w), S _mix 2:1 (35% w/w), and water (59.5% w/w) showed the highest flux (51.81 ± 5.03 μg/cm²/h) and was selected for formulation into nanoemulsion gel. The gel was further optimized with respect to oil concentration (Capryol 90), polymer concentration (Carbopol), and drug content by employing the Box–Behnken design, which statistically evaluated the effects of these components on ropinirole permeation. Oil and polymer concentrations were found to have a negative influence on permeation, while the drug content had a positive effect. Nanoemulsion gel showed a 7.5-fold increase in skin permeation rate when compared to the conventional hydrogel. In conclusion, the results of the present investigation suggested a promising role of nanoemulsions in enhancing the transdermal permeation of ropinirole.     

<Emphasis Type="Italic">Iridaea cordata</Emphasis> (Gigartinales,Rhodophyta): responses to artificial UVB radiation

The effects of UVB radiation on the different developmental stages of the carrageenan-producing red alga Iridaea cordata were evaluated considering: (1) carpospore and discoid germling mortality; (2) growth rates and morphology of young tetrasporophytes; and (3) growth rates and pigment content of field-collected plant fragments. Unialgal cultures were submitted to 0.17, 0.5, or 0.83 W m⁻² of UVB radiation for 3 h per day. The general culture conditions were as follows: 12 h light/12 h dark cycles; irradiance of 55 μmol photon^.per square meter per second; temperature of 9 ± 1°C; and seawater enriched with Provasoli solution. All UVB irradiation treatments were harmful to carpospores ( 0.17  \textW \textm ^{- 2} = 40.9 ±6.9% 0.17\;{\text{W}}\,{{\text{m}}^{ - 2}} = 40.9 \pm 6.9\% , 0.5  \textW \textm ^{- 2} = 59.8 ±13.4% 0.5\;{\text{W}}\,{{\text{m}}^{ - 2}} = 59.8 \pm 13.4\% , 0.83  \textW \textm ^{- 2} = 49 ±17.4% 0.83\;{\text{W}}\,{{\text{m}}^{ - 2}} = 49 \pm 17.4\% mortality in 3 days). Even though the mortality of all discoid germlings exposed to UVB radiation was unchanged when compared to the control, those germlings exposed to 0.5 and 0.83 W m⁻² treatments became paler and had smaller diameters than those cultivated under control treatment. Decreases in growth rates were observed in young tetrasporophytes, mainly in 0.5 and 0.83 W m⁻² treatments. Similar effects were only observed in fragments of adult plants cultivated at 0.83 W m⁻². Additionally, UVB radiation caused morphological changes in fragments of adult plants in the first week, while the young individuals only displayed this pattern during the third week. The verified morphological alterations in I. cordata could be interpreted as a defense against UVB by reducing the area exposed to radiation. However, a high level of radiation appears to produce irreparable damage, especially under long-term exposure. Our results suggest that the sensitivity to ultraviolet radiation decreases with increased algal age and that the various developmental stages have different responses when exposed to the same doses of UVB radiation.     

Production of hexanoic acid from d-galactitol by a newly isolated Clostridium sp. BS-1

In a study screening anaerobic microbes utilizing d-galactitol as a fermentable carbon source, four bacterial strains were isolated from an enrichment culture producing H₂, ethanol, butanol, acetic acid, butyric acid, and hexanoic acid. Among these isolates, strain BS-1 produced hexanoic acid as a major metabolic product of anaerobic fermentation with d-galactitol. Strain BS-1 belonged to the genus Clostridium based on phylogenetic analysis using 16S rRNA gene sequences, and the most closely related strain was Clostridium sporosphaeroides DSM 1294^T, with 94.4% 16S rRNA gene similarity. In batch cultures, Clostridium sp. BS-1 produced 550 ± 31 mL L⁻¹ of H₂, 0.36 ± 0.01 g L⁻¹ of acetic acid, 0.44 ± 0.01 g L⁻¹ of butyric acid, and 0.98 ± 0.03 g L⁻¹ of hexanoic acid in a 4-day cultivation. The production of hexanoic acid increased to 1.22 and 1.73 g L⁻¹ with the addition of 1.5 g L⁻¹ of sodium acetate and 100 mM 2-(N-morpholino)ethanesulfonic acid (MES), respectively. Especially when 1.5 g L⁻¹ of sodium acetate and 100 mM MES were added simultaneously, the production of hexanoic acid increased up to 2.99 g L⁻¹. Without adding sodium acetate, 2.75 g L⁻¹ of hexanoic acid production from d-galactitol was achieved using a coculture of Clostridium sp. BS-1 and one of the isolates, Clostridium sp. BS-7, in the presence of 100 mM MES. In addition, volatile fatty acid (VFA) production by Clostridium sp. BS-1 from d-galactitol and d-glucose was enhanced when a more reduced culture redox potential (CRP) was applied via addition of Na₂S·9H₂O.     

An initial comparison of tubular netting versus tie–tie methods of cultivation for Kappaphycus alvarezii (Rhodophyta, Solieriaceae) on the south coast of Rio de Janeiro State, Brazil

A new cultivation technique for Kappaphycus alvarezii was used in the Brazilian southeastern coast (23°02′25″ S and 43°53′39″ W), the tubular netting on floating rafts. The tubular net technique (TN) was compared with the current method of tie–tie (TT). After 47 days, the daily growth rate (mean ± SD of TN and TT = 3.4 ± 0.7% day⁻¹), carrageenan yield (mean ± SD of TN and TT = 36 ± 1.3%), gel strength (mean ± SD of TN and TT = 730 ± 114.5 g cm2), and viscosity (mean ± SD of TN and TT = 350 ± 60.4 cP) did not differ between the two techniques (p > 0.05). The amount of time used to plant (TT = 30.2 ± 2.3 min and TN = 13.3 ± 3.4 min) and to harvest the seedlings (TT = 36.2 ± 2.7 min and TN = 17.8 ± 3.7 min) were lower in TN (p < 0.001). It is concluded that TN was more effective than TT, the cultivation management (time rates used to plant and harvest the seedlings) was 53.6% faster, no seedlings were lost, the cost was lower, and an estimated return in 1 year of nearly 20% more than that of the TT technique.     

Change in ammonia-oxidizing microorganisms in enriched nitrifying activated sludge

In this study, sludge was taken from a municipal wastewater treatment plant that contained a nearly equal number of archaeal amoA genes (5.70 × 10⁶ ± 3.30 × 10⁵ copies mg sludge⁻¹) to bacterial amoA genes (8.60 × 10⁶ ± 7.64 × 10⁵ copies mg sludge⁻¹) and enriched in three continuous-flow reactors receiving an inorganic medium containing different ammonium concentrations: 2, 10, and 30 mM NH₄⁺–N (28, 140, and 420 mg N l⁻¹). The abundance and communities of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) in enriched nitrifying activated sludge (NAS) were monitored at days 60 and 360 of the operation. Early on, between day 0 and day 60 of reactor operation, comparative abundance of AOA amoA genes to AOB amoA genes varied among the reactors depending on the ammonium levels found in the reactors. As compared to the seed sludge, the number of AOA amoA genes was unchanged in the reactor with lower ammonium level (0.06 ± 0.04 mgN l⁻¹), while in the reactors with higher ammonium levels (0.51 ± 0.33 and 0.25 ± 0.10 mgN l⁻¹), the numbers of AOA amoA genes were deteriorated. By day 360, AOA disappeared from the ammonia-oxidizing consortiums in all reactors. The majority of the AOA sequences from all NASs at each sampling period fell into a single AOA cluster, however, suggesting that the ammonium did not affect the AOA communities under this operational condition. This result is contradictory to the case of AOB, where the communities varied significantly among the NASs. AOB with a high affinity for ammonia were present in the reactors with lower ammonium levels, whereas AOB with a low affinity to ammonia existed in the reactors with higher ammonium levels.     

Role of a Tetraethylammonium-Sensitive Component of Potassium Currents in High-Frequency Tonic Impulsation Generated by Rat Retinal Ganglion Cells

Using the voltage/current clamp technique in the whole-cell configuration, we studied the role of the highly tetraethylammonium (TEA) -sensitive component of integral potassium current in the generation of high-frequency tonic impulsation by rat retinal ganglion cells (RGCs). Application of 0.5 mM TEA led to a decrease in the frequency of evoked tonic impulsation by RGCs by 63% (from 55 ± 10 sec^–1 in the control to 26 ± 5 sec^–1 in the presence of the blocker; n = 11). In this case, the duration of single action potentials at the level of 50% their amplitude increased by 64% (from 1.1 ± 0.1 to 1.8 ± 0.1 msec; n = 11), the rate of repolarization decreased by 54% (from −101 ± 9 to −46 ± 5 mV/msec; n = 11), and the amplitude of afterhyperpolarization dropped by 62% (from −16 ± 2 to −6 ± 2 mV; n = 11). Upon the action of 0.5 mM TEA, the amplitude of the integral potassium current in RGCs decreased; the current component sensitive to the above blocker was equal to 0.41 ± 0.05 nA (n = 6), while the respective value in the control was 1.62 ± 0.14 nA (n = 12). Thus, a moderate (on average, by 25%) decrease in the amplitude of the above potassium current significantly influenced the characteristics of impulse activity generated by RGCs. The TEA-sensitive component of the current was similar to the Kv3.1/Kv3.2 potassium current described earlier. The obtained data are indicative of the key role of the highly TEA-sensitive component of the potassium current (passed probably via Kv3.1/Kv3 channels) in high-frequency tonic activity generated by RGCs.