Relationships between kernel vitreousness and dry matter degradability for diverse corn germplasm: II. Ruminal and post-ruminal degradabilities

Correlations between kernel vitreousness and ruminal in situ (RDMD) and total tract dry matter (TDMD; sum of ruminal in situ and post-ruminal in vitro measurements) degradabilities were determined for 33 diverse corn germplasm sources. These included a wide range of endosperm characteristics from opaque 2 (o2) types to densely packed flint types, and a number of intermediates. Harvests were done at two growth stages; 1/2 milk-line (ML) and black-layer (BL). Kernels from middle portion of ears were oven dried at 40 °C for 72 h and ground through a Wiley mill (6 mm screen) for measurement of in situ RDMD after 0 and 14 h of incubation using two steers (1.5 g/bag × 8 replicates per time point per steer in 5 cm × 5 cm bags of 50 μm pore size). Residue from the 14 h bags proceeded to an 8 h in vitro enzymatic post-ruminal digestion after which the residue was oven dried at 62 °C for 48 h and dry matter content determined. Inbred by harvest-stage interactions were observed for 0-h disappearance and TDMD. Vitreousness had strong negative correlations with degradability measurements, particularly for more mature (BL) samples (−0.728, −0.770 and −0.603) versus ML (−0.569, −0.541 and −0.338) for 0 h disappearance, RDMD and TDMD, respectively. Vitreousness was highly correlated with corn degradability, especially at a black-layer stage of harvest, in this diverse corn germplasm.     

Thermal and structural properties of unusual starches from developmental corn lines

Starches from exotic corn lines were screened by using differential scanning calorimetry (DSC) to find thermal properties that were significantly different from those exhibited by starches from normal Corn Belt lines. Two independent gelatinization transitions, one corresponding to the melting of a peak at 66 °C and the other to a peak melting at 69 °C, were found in some starches. The melting characteristics were traced to two separate types of granules within the endosperm. Strong correlations were found between DSC properties and proportion of large granules with equivalent diameter ≥17 μm. Starches with a lower peak onset gelatinization temperature (T_oG), had a lower normalized concentration of chains with a degree of polymerization (dp) of 15–24 and/or a greater normalized concentration of chains with a dp of 6–12. These studies will aid in understanding structure–thermal property relationships of starches, and in identifying corn lines of interest for commercial breeding.     

Effects of Acacia nilotica, A. polyacantha and Leucaena leucocephala leaf meal supplementation on performance of Small East African goats fed native pasture hay basal forages

Optimal utilisation of tannin-rich browse tree fodders including Acacia spp. foliages as crude protein (CP) supplements to ruminants in the tropics is limited by less available information on their feed nutritive potential. Two studies were conducted to: (1) determine rate and extent of ruminal dry matter (DM) degradability (DMD) and (2) investigate effect of sun-dried Acacia nilotica (NLM), A. polyacantha (PLM) and Leucaena leucocephala leaf meal (LLM) supplementation on growth performance of 20 growing (7–9 months old) Small East African male goats (14.6 ± 0.68 kg) fed on native pasture hay (NPH) basal diet for 84 days in a completely randomised design experiment in north-western Tanzania. The goats were randomised into four treatment groups consisting of five animals each. Three supplement diets: 115.3 g NLM (T₂), 125.9 g PLM (T₃) and 124.1 g LLM (T₄), which was used as a positive control, were supplemented at 20% of the expected DM intake (DMI; i.e., 3% body weight) to the three animal groups fed on NPH (basal diet) compared to the animals in a control group that were fed on NPH without browse supplementation (T₁).

NPH had significantly the lowest (P < 0.05) CP of 45.5 g kg⁻¹ DM compared to NLM, PLM and LLM (159, 195 and 187 g kg⁻¹ DM, respectively). NPH had higher (P < 0.05) fibre fractions; lower ruminal DM degradability characteristics and ME than NLM, PLM and LLM. Supplementation of the animals with browse resulted to (P < 0.05) higher average daily weight gains (ADG) of 157.1 g day⁻¹ in T₄ than the animals fed on T₂ (114.3 g day⁻¹) and T₃ (42.9 g day⁻¹), and even to those fed on T₁ (control), which lost weight (−71.4 g day⁻¹). Improved weight gains were mainly due to corrected feed nitrogen (N) or CP due to supplementation of the animals with browse fodder. Too low CP of the NPH would not meet the normal requirements of CP (80 g CP kg⁻¹ DM) for optimal rumen microbial function in ruminants. Higher ADG due to LLM (T₄) and NLM (T₂) supplementation suggest optimised weight gains due to browse supplementation (20% of expected DMI); while lower weight gains from supplementation with PLM (T₃) indicate the possible utilisation of A. polyacantha leaves to overcome weight losses especially during dry seasons.     

Ensiling characteristics, nutrient composition, and in situ ruminal and whole tract degradability of brown midrib and leafy corn silage

A study was conducted to compare the ensiling characteristics, chemical composition, and the ruminal and total tract nutrient degradabilities of leafy (Cargill F227) and brown midrib (Mycogen TMF94) corn silage hybrids. Corn was grown in Saint-Jean-sur-Richelieu, Quebec, Canada, harvested at a target 350 g kg(-1) dry matter (DM) content, and ensiled in mini-silos for 0, 2, 4, 8, 16, and 45 d. Two non-lactating Holstein cows fitted with ruminal and proximal duodenal cannulae were used to determine ruminal and whole tract nutrient degradability. Forage from both hybrids went through a rapid fermentation with a sharp decline in pH during the first 2 d of ensiling, pH in both silage being less than 4.0 after 45 d. Lactic acid concentration was however greater for leafy than brown midrib corn. Chemical analysis of silage after 45 d of ensiling revealed that hybrids differed in their composition. Compared to leafy corn, brown midrib corn had lower neutral detergent fiber (NDF), acid detergent fiber (ADF), acid detergent lignin (ADL), crude protein (CP), and neutral detergent and acid detergent insoluble proteins, but higher starch and net energy of lactation (NEL) values. Results of the in situ incubation experiment indicated that compared to leafy corn brown midrib corn had greater ruminal DM (64 vs. 54%), CP (73 vs. 71%), and NDF (32 vs. 24%) degradabilities. Brown midrib corn silage also had greater DM ruminal (53 vs. 48%) and total tract (67 vs. 61%) digestibilities, as well as greater NDF ruminal (34 vs. 25%), intestinal (10 vs. 8%), and total tract (43 vs. 33%) digestibilities. Type of corn hybrid will thus greatly affect silage chemical composition and nutrient digestibility.     

Effects of vitreousness and particle size of maize grain on ruminal and intestinal in sacco degradation of dry matter,starch and nitrogen

We assessed the effects of vitreousness and particle size of maize grain on ruminal and intestinal in sacco degradation of dry matter, starch and nitrogen. Six maize grain (Zea mays) genotypes characterized by differing vitreousness (proportion of vitreous in total endosperm) were ground (3-mm screen; Gr, ground particles, mean particle size (MPS): 526 μm) and cracked with a roller mill using two gap width settings (CS, cracked small particles, MPS: 1360 μm; CL, cracked large particles, MPS: 2380 μm). The ruminal and intestinal in sacco degradation of dry matter, starch and nitrogen was measured on three dry Holstein cows, fitted with rumen, proximal duodenum and terminal ileum cannulas, fed maize silage ad libitum twice daily. The ruminal starch degradability and intestinal digestibility differed among genotypes (P<0.001) and decreased as particle size increased (P<0.001). For the same particle size, starch ruminal degradability decreased (P<0.05) and intestinal digestibility decreased (P<0.002) with vitreousness. Particle size and vitreousness of maize grain are efficient factors for manipulating the amount of starch escaping rumen degradation, but may be limiting for the amount of starch digested in the small intestine.     

Fattening Pelibuey lambs with sugar cane tops and corn complemented with or without slow intake urea supplement

One hundred and eighty Pelibuey lambs were fattened for 120 days with three treatments evaluating in situ DM disappearance, voluntary DM or OM intake, rumen degradation, rate of passage, NH₃ and VFA concentration, apparent digestibility, ruminal pH, total fermentable carbohydrates and weight gains. The first diet (T1), 60 lambs (25.7 ± 0.7 kg BW) plus two cannulated sheep were fed 100% sugar cane tops (SCT) per day; treatment 2 (T2), 60 lambs (25.4 ± 0.4 kg BW) plus two cannulated sheep diet was SCT supplemented with 200 g DM of slow intake urea supplement (SIUS) (a 4% urea mixture) per day. Treatment 3 (T3), 60 sheep (25.5 + 0.5 kg BW) plus two cannulated animals, fed SCT and corn crop chopped (C) mixture (40:60%) were supplemented with 200 g SIUS per day. Ruminal kinetics were determined thereafter in four adult ruminal cannulated sheep in metabolic cages. A 100% corn diet was added to the second trial (T4). At all times, fibrous forages were available and exceeded VDMI. Daily BW gain by SCT animals was 70 g/d compared to 135 g/d for SCT/SIUS and 218 g/d for SCT/C/SIUS diets (P < 0.05). Total forage intake was similar for T1 and T2 treatments and higher for T3 (P < 0.05). Kinetics observation showed a total DMI variation increasing from 474 g/d for T1; 597 g/d for T4 to 797 g/d for T2, and 917 g/d for T3, being different for the last two treatments compared with the first two (P < 0.001) and similar among SIUS diets. Rumen pH diminished from 6.8 to 6.7 at 2 h, when SIUS was offered, and stayed above 6.5 during the 12 h of sampling when SIUS was the only supplement and 6.3 when corn was added, while in SCT lambs, ruminal pH decreased to 6.2 by 6 h and rose again to 6.4 after 12 h favoring bacterial multiplication. Ammonia concentration and digestion of potential digestible and indigestible fractions were significantly augmented (P < 0.05) by SIUS. In vivo nitrogen digestibility was different (P < 0.05) among diets: T1, T2, T3, and T4 (36.5%, 74.1%, 76.5%, and 39.4%, respectively). In vivo digestibility of DM, OM, cellulose and hemicellulose was similar among T2 and T3 groups but different when T1 or T4 diets were offered (P < 0.05). NDF digestibility was higher for T3 (77.1%) and T2 (75.1%) but different from T1 (57.1%) and T4 (59.1%) (P < 0.01). In situ DM disappearance did not show differences among T2 and T3 diets at 9, 12, 24 and 48 h of incubation but differed in percentage for T1 and T4. Digestion rate constant (k_d) was similar among SIUS diets 0.038 and 0.039, but different from T1 and T4 diets 0.023; 0.021 (P < 0.05). True digestibility of NDF was higher (P < 0.05) in SCT/SIUS/C (32.5%) and SCT/SIUS (32.7%) compared with SCT (26.2%) and C (27.4%). Passage of NDF was different (P < 0.05) between T1 (0.059 h⁻¹) and T4 (0.061 h⁻¹) from 0.079 (T2) and 0.081 k_p/h (T3). The half-time (t_1/2) disappearance for hemicellulose was higher (P < 0.05) for SCT/SIUS (22.1 h) and SCT/SIUS/C (29.2 h) as compared to SCT (18.1 h). Kinetic performance among SCT and C diet did not have a statistical difference. Utilization of fibrous forages by lambs was higher (P < 0.05) for SIUS diets. VDMI and apparent digestibility showed better performance for DM and OM, lower acetate molar proportions and greater feed intake by both SIUS diets which was reflected in body weight gain.     

Conceptual model of seagrass die-off in Florida Bay: Links to biogeochemical processes

Seagrasses are recognized as important plant communities in coastal estuaries and lagoons across both tropical and temperate climes; thus, large-scale seagrass die-off events worldwide are of general concern. In Florida Bay, at the southern terminus of the Florida peninsula, seagrass die-off events up to 4000 ha have been reported and smaller scale mortality events are noted annually. In the present study, we examined several hypothesized causative factors (high temperature, hypersalinity, sulfide toxicity) of seagrass (Thalassia testudinum) mortality in Florida Bay. To test sulfide effects, in situ sulfide production was stimulated by applying a labile carbon source (glucose) to sulfate reducers in the sediment at five sites across the bay (northeastern, northcentral, and southwestern basins). During the one year study, high temperature (32–36 °C) and salinity (> 50 psu) were recorded in the bay associated with a regional drought. We also experienced major seagrass die-off events at two of our southwestern bay sites. These field conditions provided an excellent opportunity to closely examine cause–effect relationships among stressors and die-off events in the field, and verify results of our previous mesocosm experiments. Even though glucose amendments stimulated porewater sulfides in bay sediments (4–8 mmol L^− 1), no significant differences in biomass, short shoot density or final growth rates were found between control and glucose plots. In addition, the highest growth rates and shoot densities were concomitant with maximum water column salinity (> 50 psu) and temperature (32–36 °C), when porewater sulfides were also in the millimolar range. Large-scale seagrass mortality events, encompassing  50% of the entire meadow at one site, occurred at southwestern bay sites when plants were down regulating (slower growth and shoot density), probably in response to shorter day length and lower temperature (30–34 to 23–26 °C) from October, 2004 to January, 2005. Sulfate reduction rates (SRR) were also 2-fold higher in the southwestern (214–488 nmol cm^− 3 d^− 1) versus northcentral and northeastern (97–240 nmol cm^− 3 d^− 1) bay sites, possibly limited by labile carbon, which we found to stimulate SRR 3-fold in northeastern and northcentral bay sites (461–708 nmol cm^− 3 d^− 1) and 4-fold at southwestern bay sites (1211–2036 nmol cm^− 3 d^− 1). Based on a synthesis of the field data reported herein, our mesocosm experiments to date, and contributions by others, we present a conceptual model of seagrass die-off in Florida Bay outlining a cascade of stressors, stimulated by P enrichment, which leads to high O₂ consumption in the system triggering a seagrass die-off event.     

Falls are associated with decreased renal function and insufficient calcitriol production by the kidney

In a double blind placebo controlled 3-year osteoporosis study in elderly women, we collected prospective data on falls. The study population comprised 489 normal elderly women aged 65–77 years randomized to four groups: placebo, calcitriol 0.25 mcg b.i.d., conjugated equine estrogens (0.625 mg/day) and calcitriol + estrogen. Falls occurred in 57% of all women.

Using a Poisson regression model, the placebo group with low GFR-creatinine clearance (CrCl < 60 ml/min) had 60% more falls compared to the group with CrCl ≥ 60 ml/min. Further sub group analyses showed that there is no increased risk of falls with CrCl 60–70, 70–80 and >90 ml/min. Calcitriol treatment significantly reduced the number of falls by 50% (OR = 0.5, 95% CI: 0.4–0.9, p = 0.010) compared to placebo in the low CrCl group.

The group with lower CrCl had lower calcium absorption (p < 0.001), lower serum 1,25-dihydroxyvitamin D (1,25(OH)₂D) (p < 0.001) and normal serum 25OHD suggesting that there is decreased conversion of 25OHD to 1,25(OH)₂D by the aging kidney. It is postulated that the decrease in falls on calcitriol therapy is related to an increase in serum 1,25(OH)₂D, upregulation of VDR and improvement in muscle strength although one cannot exclude an effect on the central nervous system.     

Critical review of the methods used to measure the apparent dissociation constant and ligand purity in Ca2+ and Mg2+ buffer solutions

Using simulated Ca²⁺ and Mg²⁺ buffers, methods proposed to measure both ligand purity and the apparent dissociation constant (K_app) were investigated regarding (1) predicted accuracy of both parameters and (2) generality of the solution.

The Bers’ Ca²⁺ macroelectrode method [Bers, D. M., 1982 A simple method for the determination of free [Ca] in Ca-EGTA solutions Am. J. Physiol. 242, C404–C408] cannot be used with Mg²⁺-macroelectrodes and is partly arbitrary since the linear part of the Scatchard plot is judged subjectively. Iterative methods have therefore been introduced. Iteration based on the Bers’ method or the lumped interference in the Nicolsky–Eisenman equation also failed with Mg²⁺ macroelectrodes. The Oiki et al., method [Oiki, S., Yomamoto, T., Okada, Y., 1994. Apparent stability constants and purity of Ca-chelating agents evaluated using Ca-sensitive electrodes by the double-log optimization method Cell Calcium 15, 209–46.] cannot be applied to Mg²⁺ macroelectrodes. The pH titration method of Moisescu and Pusch (Pflügers, Arch., 355, R122, 1975) predicted EGTA purity and Ca²⁺ contamination, but K_app values for EGTA were approximate. It cannot be applied to Mg²⁺ binding. The partition method [Godt, R.E., 1974. Calcium-activated tension of skinned muscle fibres of the frog. Dependence on magnesium adenosine triphosphate concentration J. Gen. Physiol. 63, 722–739.] only approximately estimated the K_app. Calibration, maintaining contaminating [Ca²⁺]/[Mg²⁺] at <1 μmol l⁻¹, and setting standards by dilution, is the ultimate check of calculated ionised concentrations, although technically difficult. The macroelectrode method of Lüthi et al. [1997. Calibration of Mg²⁺-selective macromolecules down to 1 μmol l⁻¹ in intracellular and Ca⁺- containing extracellular solutions. Exp. Physiol. 82, 453–467] accurately predicted purity and K_app at pK_app values >4 and was independent of electrode characteristics. It is considered the method of choice.

Macroelectrode primary calibration should be carried out in solutions varying from 0.5 to 10 mmol l⁻¹ combined with either Ca–EGTA or Mg–EDTA buffers; the [Ca²⁺] and [Mg²⁺] in other buffer ligands can be measured in a secondary calibration.     

Physicochemical properties of endosperm and pericarp starches during maize development

Endosperm starch and pericarp starch were isolated from maize (B73) kernels at different developmental stages. Starch granules, with small size (2–4 μm diameter), were first observed in the endosperm on 5 days after pollination (DAP). The size of endosperm-starch granules remained similar until 12DAP, but the number increased extensively. A substantial increase in granule size was observed from 14DAP (diameter 4–7 μm) to 30DAP (diameter10–23 μm). The size of starch granules on 30DAP is similar to that of the mature and dried endosperm-starch granules harvested on 45DAP. The starch content of the endosperm was little before 12DAP (less than 2%) and increased rapidly from 10.7% on 14DAP to 88.9% on 30DAP. The amylose content of the endosperm starch increased from 9.2% on 14DAP to 24.2% on 30DAP and 24.4% on 45DAP (mature and dried). The average amylopectin branch chain-length of the endosperm amylopectin increased from DP23.6 on 10DAP to DP26.9 on14DAP and then decreased to DP25.4 on 30DAP and DP24.9 on 45DAP. The onset gelatinization temperature of the endosperm starch increased from 61.3 °C on 8DAP to 69.0 °C on 14DAP and then decreased to 62.8 °C on 45DAP. The results indicated that the structure of endosperm starch was not synthesized consistently through the maturation of kernel. The pericarp starch, however, showed similar granule size, starch content, amylose content, amylopectin structure and thermal properties at different developmental stages of the kernel.     

Effect of the proportions of neutral detergent fibre and starch, and their degradation rates, on in vitro ruminal fermentation

Effects of proportions of neutral detergent fibre (aNDFom) and starch, as well as their degradation rates, on rumen fermentation were tested using an in vitro rumen simulation system (SIMCO). The in vitro system was designed to simulate selective particle retention and had an average fluid volume of 1150 ml with a liquid dilution rate of approximately 0.07 h⁻¹. Two types of hay (aNDFom sources) and two types of starch were each included at two different levels in the diet and were examined in an experiment following a 2×2×2 factorial arrangement of treatments (eight diet combinations). The hay was either late-cut timothy (Phleum pretense L.) or early cut meadow grass (Poa pratensis L.), with ruminal in situ aNDFom digestion rates of 0.03–0.04 and 0.07–0.08 h⁻¹, respectively. The two starch types were raw (R) and cooked (C) potato starch with previously determined in vitro ruminal digestion rates of 0.04 and 0.20 h⁻¹, respectively. The starch levels were 300 and 600 g/kg diet dry matter (DM) with the remaining being hay (282–682 g/kg DM) and peptone (14–111 g/kg DM). The aNDFom level varied among the diets with different starch levels and hay types. The peptone acted as a source of peptides and, together with ammonia salts from buffer, was used to balance the N contents of the diets. The feeding level for each of the eight vessels was 28 g DM/d. Two 10-day simulations were made with the system. The average pH was higher (P<0.05) for all treatments with raw potato starch (6.19) versus cooked starch (6.07). Protozoa scores, on a qualitative scale, declined faster at the higher starch level. The aNDFom digestibility was, as expected, higher (P<0.001) for meadow hay (0.57) than timothy (0.32), and was also higher (P<0.001) at the lower starch level (0.54) versus the higher (0.35). Microbial protein production efficiency (mg microbial N/g organic matter truly digested) was higher for the faster degrading aNDFom (P<0.01) and starch (P<0.05) sources, but was not affected by starch level. Cooked starch resulted in a lower acetate proportion (449 mmol/mol versus 591 mmol/mol VFA; P<0.001) but higher proportions of propionate (297 mmol/mol versus 236 mmol/mol VFA; P<0.001), and butyrate (169 mmol/mol versus 127 mmol/mol VFA; P<0.01). Butyrate increased with starch level (127 mmol/mol versus 169 mmol/mol VFA; P<0.01), and was also higher for meadow hay versus timothy (168 mmol/mol versus 128 mmol/mol VFA; P<0.01). Interactions between the treatments demonstrate that the response in VFA pattern to starch level is dependent on starch and aNDFom sources. Substrates such as starch and aNDFom are fermented differently depending on their rates of ruminal degradation.     

Effect of monensin, fish oil or their combination on in vitro fermentation and conjugated linoleic acid (CLA) production by ruminal bacteria

An in vitro study was conducted to examine the effect of adding monensin, fish oil, or their combination on rumen fermentation and conjugated linoleic acid (CLA) production by mixed ruminal bacteria when incubated with safflower oil. Concentrate (1 g/100 ml) with safflower oil (0.2 g/100 ml) was added to a mixed solution (600 ml) of strained rumen fluid and buffer (control). Monensin (10 ppm), fish oil (0.02 g/100 ml), or monensin plus fish oil was also added into control mixture. All the culture solutions prepared were incubated anaerobically at 39 °C for 12 h. A higher pH and ammonia concentration were observed from the culture solution containing monensin at 12 h of incubation than those from the control or the culture containing fish oil. Monensin increased (P < 0.007) the C₃ content over all the collection times of culture solution while reducing the C₄ content at 6 h (P < 0.018) and 12 h (P < 0.001) of incubations. Supplementation of monensin, fish oil or their combination changed the content of C₁₈-fatty acids of ruminal culture. Monensin alone reduced (P < 0.021) the content of cis-9, trans-11 CLA compared to fish oil at all sampling times, but increased (P < 0.041) the trans-10, cis-12 CLA production compared to fish oil addition and the control which were similar at incubation for 12 h. The combination of monensin and fish oil increased the content of cis-9, trans-11 CLA (P < 0.023) and transvaccenic acid (TVA, P < 0.018) significantly compared to the control or monensin alone at incubation for 12 h.     

Binding activities by propiverine and its N-oxide metabolites of L-type calcium channel antagonist receptors in the rat bladder and brain

The present study was undertaken to characterize the binding activities of propiverine and its N-oxide metabolites (1-methyl-4-piperidyl diphenylpropoxyacetate N-oxide: P-4(N → O), 1-methyl-4-piperidyl benzilate N-oxide: DPr-P-4(N → O)) toward L-type calcium channel antagonist receptors in the rat bladder and brain. Propiverine and P-4(N → O) inhibited specific (+)-[³H]PN 200–110 binding in the rat bladder in a concentration-dependent manner. Compared with that for propiverine, the K_i value for P-4(N → O) in the bladder was significantly greater. Scatchard analysis has revealed that propiverine increased significantly K_d values for bladder (+)-[³H]PN 200–110 binding. DPr-P-4(N → O) had little inhibitory effects on the bladder (+)-[³H]PN 200–110 binding. Oxybutynin and N-desethyl-oxybutynin (DEOB) also inhibited specific (+)-[³H]PN 200–110 binding in the rat bladder. Propiverine, oxybutynin and their metabolites inhibited specific [N-methyl-³H]scopolamine methyl chloride ([³H]NMS) binding in the rat bladder. The ratios of K_i values for (+)-[³H]PN 200–110 to [³H]NMS were markedly smaller for propiverine and P-4(N → O) than oxybutynin and DEOB. Propiverine and P-4(N → O) inhibited specific binding of (+)-[³H]PN 200–110, [³H]diltiazem and [³H]verapamil in the rat cerebral cortex in a concentration-dependent manner. The K_i values of propiverine and P-4(N → O) for [³H]diltiazem were significantly smaller than those for (+)-[³H]PN 200–110 and [³H]verapamil. Further, their K_i values for [³H]verapamil were significantly smaller than those for (+)-[³H]PN 200–110. The K_i values of propiverine for each radioligand in the cerebral cortex were significantly (P < 0.05) smaller than those of P-4(N → O). In conclusion, the present study has shown that propiverine and P-4(N → O) exert a significant binding activity of L-type calcium channel antagonist receptors in the bladder and these effects may be pharmacologically relevant in the treatment of overactive bladder after oral administration of propiverine.     

Covalent immobilisation of manganese peroxidases (MnP) from Phanerochaete chrysosporium and Bjerkandera sp. BOS55

Manganese peroxidases (MnP) from Phanerochaete chrysosporium and Bjerkandera sp. BOS55 were immobilised in glutaraldehyde–agarose gels. Four different strategies were considered concerning the activation of the support (low or high density) and the ionic strength (low or high). In terms of immobilisation rate and yield, better results were obtained when low ionic strength conditions and high density activated support (75 μEq/ml) were used. Immobilisation proceeds initially with an ionic adsorption which facilitates the further covalent attachment of the enzyme to the support. An almost complete immobilisation has been attained in a very short period (0.5–2 h). Immobilisation maintained a high percentage of MnP activity for long periods of time (activity levels of 50–60% after more than 1 year at room temperature storage). Other desirable effects such as increased thermostability at 50–60 °C for MnP from Bjerkandera and higher resistance to high H₂O₂ concentrations for MnP for P. chrysosporium were also obtained. This latter is quite an interesting feature because it avoids the inactivation of the enzyme in the presence of an unbalanced concentration of H₂O₂. The improved characteristics of the immobilised MnP make its application in several fields such as the enzymatic oxidation of hardly degradable compounds more feasible.     

Determination of zinc in vegetal tissue microsamples by platinum-wire loop in flame atomization atomic absorption spectrometry

The zinc content of 3 μL of vegetal samples (tree leaves, lichens and grape sap) atomized from a Pt-wire in the methane–air flame has been determined by atomic absorption spectrometry. The effect of gas flow rates and the atomization height in the flame on the absorption of zinc was evaluated at 213.9 nm. The best results were obtained at a height of 5 mm and gas flow rates of 200 L/h air and 26 L/h methane, respectively. The effect of Na, K, Ca, Mg, SO₄²⁻, and PO₄³⁻ on the absorption of zinc was studied too. The detection limit of 0.40 ± 0.21 ng was obtained at a significance level of 0.05, using the two-step Neyman–Pearson criterion. The zinc content of the samples has been determined with continuous nebulization and by atomization from the Pt-wire, using both the standard calibration curve and the standard addition method. The results of the two procedures agree within the determination errors.     

Estrone sulfate (E1S), a prognosis marker for tumor aggressiveness in prostate cancer (PCa)

Seeking insight into the possible role of estrogens in prostate cancer (PCa) evolution, we assayed serum E₂, estrone (E₁), and estrone sulfate (E₁S) in 349 PCa and 100 benign prostatic hyperplasia (BPH) patients, and in 208 control subjects in the same age range (50–74 years).

E₁ (pmol/L ± S.D.) and E₁S (nmol/L ± S.D.) in the PCa and BPH patients (respectively 126.1 ± 66.1 and 2.82 ± 1.78, and 127.8 ± 56.4 and 2.78 ± 2.12) were significantly higher than in the controls (113.8 ± 47.6 and 2.11 ± 0.96). E₂ was not significantly different among the PCa, BPH, and control groups. These assays were also carried out in PCa patients after partition by prognosis (PSA, Gleason score (GS), histological stage, and surgical margins (SM)). Significantly higher E₁S levels were found in PCa with: PSA > 10 ng/L (3.05 ± 1.92) versus PSA ≤ 10 ng/mL (2.60 ± 1.55), stage pT3-T4 (2.99 ± 1.80) versus pT2 (2.58 ± 1.58), and positive (3.26 ± 1.95) versus negative margins (2.52 ± 1.48). E₁ was higher in poor- than in better-prognosis PCa. E₂ was significantly higher in PCa with GS ≥ 4 + 3 (109.5 ± 43.8) versus GS ≤ 3 + 4 (100.6 ± 36.5) and increased significantly when GS increased from 3 + 3 to 4 + 4. Estrogens, especially E₁S appeared to be possible markers of PCa progression.

Attempting to identify potential sources of E₂ in PCa according to prognosis, as well as in BPH, we found a significant correlation coefficient between E₁S and E₂ (0.266–0.347) in poor-prognosis PCa and no correlation in BPH (0.026) and better-prognosis PCa (0.013–0.104).

It is as though during progression of PCa from good to poor prognosis there were a shift in the E₁ to E₂ metabolic pathway from predominantly oxidative to predominantly reductive.     

Hydrogen and polyhydroxybutyrate producing abilities of microbes from diverse habitats by dark fermentative process

Thirty five bacterial isolates from diverse environmental sources such as contaminated food, nitrogen rich soil, activated sludges from pesticide and oil refineries effluent treatment plants were found to belong to Bacillus, Bordetella, Enterobacter, Proteus, and Pseudomonas sp. on the basis of 16S rRNA gene sequence analysis. Under dark fermentative conditions, maximum hydrogen (H₂) yields (mol/mol of glucose added) were recorded to be 0.68 with Enterobacter aerogenes EGU16 followed by 0.63 with Bacillus cereus EGU43 and Bacillus thuringiensis EGU45. H₂ constituted 63–69% of the total biogas evolved. Out of these 35 microbes, 18 isolates had the ability to produce polyhydroxybutyrate (PHB), which varied up to 500 mg/l of medium, equivalent to a yield of 66.6%. The highest PHB yield was recorded with B. cereus strain EGU3. Nine strains had high hydrolytic activities (zone of hydrolysis): lipase (34–38 mm) – Bacillus sphaericus strains EGU385, EGU399 and EGU542; protease (56–62 mm) – Bacillus sp. strains EGU444, EGU447 and EGU445; amylase (23 mm) – B. thuringiensis EGU378, marine bacterium strain EGU409 and Pseudomonas sp. strain EGU448. These strains with high hydrolytic activities had relatively low H₂ producing abilities in the range of 0.26–0.42 mol/mol of glucose added and only B. thuringiensis strain EGU378 had the ability to produce PHB. This is the first report among the non-photosynthetic microbes, where the same organism(s) – B. cereus strain EGU43 and B. thuringiensis strain EGU45, have been shown to produce H₂ – 0.63 mol/mol of glucose added and PHB – 420–435 mg/l medium.     

Metal–salt co-tolerance and metal removal by indigenous cyanobacterial strains

Chromium and salt tolerance in five indigenous cyanobacterial strains isolated from contaminated sites was investigated along with their metal bioaccumulative potential. All the five species showed significantly better growth when the medium was spiked with salt or chromium. As compared to single metal or salt treatment, the binary metal–salt (MS) treatments had more favorable effect on cyanobacterial growth as indicated by significantly higher concentration of the primary photosynthetic pigment chlorophyll at M₂₀S₂₀₀₀ (9.9–25.3 μg/mL) as compared to that at M₀S₀ (4.0–12.3 μg/mL). Similarly biomass was much higher at M₂₀S₁₀₀₀ and M₂₀S₂₀₀₀ (41.8–86.2 mg/10 mL) as compared to that at control, M₀S₀ (21.5–36.3 mg/10 mL). Accessory pigments like carotenoids and phycobilinproteins too tended to increase significantly in response to both metal and salts in the two species of Lyngbya (L. putealis and L. ceylanica var. constricta) and Gloeocapsa. These species also showed greater potential of chromium bioaccumulation, which increased further as both salt and metal concentration increased. In the two species of Nostoc however, bioaccumulative potential improve at higher metal concentration, but not affected significantly by salt concentration.     

New series of potent delta-opioid antagonists containing the H-Dmt-Tic-NH-hexyl-NH-R motif

Heterodimeric compounds H-Dmt-Tic-NH-hexyl-NH-R (R = Dmt, Tic, and Phe) exhibited high affinity to δ- (K_iδ = 0.13–0.89 nM) and μ-opioid receptors (K_iμ = 0.38–2.81 nM) with extraordinary potent δ antagonism (pA₂ = 10.2–10.4). These compounds represent the prototype for a new class of structural homologues lacking μ-opioid receptor-associated agonism (IC₅₀ = 1.6–5.8 μM) based on the framework of bis-[H-Dmt-NH]-alkyl (Okada, Y.; Tsuda, Y.; Fujita, Y.; Yokoi, T.; Sasaki, Y.; Ambo, A.; Konishi, R.; Nagata, M.; Salvadori, S.; Jinsmaa, Y.; Bryant, S. D.; Lazarus, L. H. J. Med. Chem. 2003, 46, 3201), which exhibited both high μ affinity and bioactivity.