Photosynthesis in response to salinity and submergence in two Rhizophoraceae mangroves adapted to different tidal elevations.

Mangrove ecosystems are vulnerable to rising sea levels. When the sea level rises, the plants are exposed to increased salinity and tidal submergence. In Taiwan, the mangrove species Kandelia obovata and Rhizophora stylosa grow in different habitats and at different elevations. To understand the response of photosynthesis to salinity and submergence in mangroves adapted to different tidal elevations, gas exchange and chlorophyll fluorescence parameters were measured in K. obovata and R. stylosa under different salinity (20 and 40‰) and submergence treatments. The period of light induction of photosynthesis for the two mangrove species was >60 min. In the induction process, the increase in photosystem efficiency was faster than the increase in stomatal opening, but CO2 fixation efficiency was restricted by stomatal conductance. The constraint of stomatal opening speed is related to the conservative water-use strategy developed in response to mangrove environments. Submergence increased the photosynthetic rate of K. obovata, but not that of R. stylosa. Although R. stylosa was more salt tolerant than K. obovata, R. stylosa was not submergence tolerant in a high-salinity environment, which may be the reason for the higher intertidal elevations observed for R. stylosa in comparison with K. obovata. The photosynthetic rate and energy-dependent quenching (qE) of the two mangroves presented a negative relationship with photoinhibition, and high-salt treatment simultaneously reduced photosynthetic rate and qE. A decrease in the photosynthetic rate increased excess energy, whereas a decrease in qE decreased photoprotection; both increased photoinhibition. As the degree of photoinhibition can be easily measured in the field, it is a useful ecological monitoring index that provides a suitable reference for mangrove restoration, habitat construction and ecological monitoring.

Dynamic real-time monitoring of chloroform in an indoor swimming pool air using open-path Fourier transform infrared spectroscopy.

This study used open-path Fourier transform infrared (OP-FTIR) spectroscopy to continuously assess the variation in chloroform concentrations in the air of an indoor swimming pool. Variables affecting the concentrations of chloroform in air were also monitored. The results showed that chloroform concentrations in air varied significantly during the time of operation of the swimming pool and that there were two peaks in chloroform concentration during the time of operation of the pool. The highest concentration was at 17:30, which is coincident with the time with the highest number of swimmers in the pool in a day. The swimmer load was one of the most important factors influencing the chloroform concentration in the air. When the number of swimmers surpassed 40, the concentrations of chloroform were on average 4.4 times higher than the concentration measured without swimmers in the pool. According to the results of this study, we suggest that those who swim regularly should avoid times with highest number of swimmers, in order to decrease the risk of exposure to high concentrations of chloroform. It is also recommended that an automatic mechanical ventilation system is installed to increase the ventilation rate during times of high swimmer load.

First and second generation total synthesis of the teicoplanin aglycon.

Full details of studies leading to the total synthesis of the teicoplanin aglycon are provided. Key elements of the first generation approach (26 steps from constituent amino acids, 1% overall) include the coupling of an EFG tripeptide precursor to the common vancomycin/teicoplanin ABCD ring system and sequential DE macrocyclization of the 16-membered ring with formation of the diaryl ether via a phenoxide nucleophilic aromatic substitution of an o-fluoronitroaromatic (80%, 3:1 atropisomer diastereoselection) followed by 14-membered FG ring closure by macrolactamization (66%). Subsequent studies have provided a second generation total synthesis which is shorter, more convergent, and highly diastereoselective (22 steps, 2% overall). This was accomplished by altering the order of ring closures such that FG macrolactamization (95%) preceded coupling of the EFG tripeptide to the ABCD ring system and subsequent DE ring closure. Notably, DE macrocyclization via diaryl ether formation on substrate 57, the key intermediate in the latter approach incorporating the intact FG ring system, occurred with exceptional diastereoselection for formation of the natural atropisomer (>10:1, 76%) without problematic C(2)(3) epimerization provided the basicity of the reaction is minimized.

Novel CCK-B receptor agonists: diketopiperazine analogues derived for CCK4 bioactive conformation.

Recently, we proposed a CCK-B agonist bioactive conformation characterized by an 'S' shape of the peptidic backbone which was derived from structure-activity relationships and conformational analysis of CCK4 (Trp-Met-Asp-Phe-NH2) analogues. Using this template, we report here the synthesis of cyclic CCK4 analogues which contain, in place of the Trp-Met dipeptide, a diketopiperazine moiety resulting from a cyclization between Nle and N-substituted (D)Trp residues and coupled with a small linker to Asp-Phe-NH2. Some of these compounds displayed good affinities and selectivities for the CCK-B receptor. The results are discussed in terms of size, hydrophobicity and spatial orientation of the side-chains on the diketopiperazine ring. The most potent ligand exhibited potent and full CCK-B receptor agonist properties in promoting the hydrolysis of inositol phosphates (EC50 = 8 nM) in CHO cells, stably transfected with the rat brain CCK-B receptor. This compound was also shown to be a potent selective CCK-B/gastrin receptor agonist since, it increased gastric acid secretion measured in anesthetized rats on i.v. administration. These compounds provide a rigid template for the design of non-peptide CCK-B agonists, by modification of the remaining peptide moiety.

Comparative conformational analysis of CCK-B agonist Boc-Trp-Phg-Asp-(1-Nal)-NH2 and CCK1-B antagonist Boc-Trp-Phg-Asp-(1-Nal)-N(Me)2 using 1HNMR spectroscopy and restrained molecular dynamics.

The tetrapeptide Boc-Trp-Phg-Asp-(1-Nal)-NH2 is a potent CCK-B agonist. Interestingly, bis-methylation of the C-terminal carboxamide group of this compound leads to Boc-Trp-Phg-Asp-(1-Nal)-N(Me)2 which behaves as a CCK-B antagonist in electrophysiological studies on hippocampal neurones (Corringer et al., 1993). In order to ascertain whether bismethylation of the terminal carboxamide group has an influence on the conformational preferences of the peptide, we have undertaken a comparative conformational analysis of the two tetrapeptides by the combined use of 2D NMR spectroscopy and restrained molecular dynamics. The solution conformation of the two peptides were examined by 1H NMR in a d6-DMSO/H2O (80:20) mixture. 1H-1H distance constraints, derived from 2D NOESY and ROESY experiments, were used as inputs for subsequent restrained molecular dynamics simulations. Comparison of the NMR and molecular modeling data indicates different conformational preferences for these two peptides. Interestingly, the aromatic side chains of the CCK-B antagonist Boc-Trp-Phg-Asp-(1-Nal)-N(Me)2 in its preferential conformation, overlap their corresponding moieties in the two non peptide CCK-B antagonists L-362,260 and LY-288,513. The differences in conformational behaviour of the studied tetrapeptides could, at least in part, account for their opposite agonist/antagonist profile, a findings which could serve for the design of new conformationally restricted CCK-B analogs.

Role of N- and C-terminal substituents on the CCK-B agonist-antagonist pharmacological profile of Boc-Trp-Phg-Asp-Nal-NH2 derivatives.

Among the CCK derivatives, the tetrapeptide Boc-Trp-Phg-Asp-Nal-NH2 (1) behaves as a short potent CCK-B agonist which led to the development of an efficient peptidase-resistant CCK-B antagonist by bismethylation of its terminal CONH2 group. Further modifications of the N- and C-terminal moieties of 1 have been performed and are described in this paper, together with the pharmacological profile of the novel synthetized compounds. Introduction of more bulky substituents than NalNH2 on the C-terminal part decreased the CCK-B receptor binding affinity. In the series of N-protected tetrapeptides X30-Phg31-Asp32-Nal33-N(CH3)2, the Boc-substituent was shown to be optimal among the N-protecting groups Boc, 2Adoc, propionyl or acetyl when X = Trp. On the other hand, when X = alpha MeTrp, its optimal N-protecting group was 2Adoc and its configuration was preferentially D. In the newly synthesized compounds, 13: 2Adoc-D-alpha MeTrp-Phg-Asp-NalN(CH3)2 and 16: 2Adoc-D-alpha MeTrp-Phg-Asp-NalNH2 had the best CCK-B receptor affinities (KI = 3.5 and 3.4 nM, respectively) and were selected for further biological evaluation. Interestingly, when tested for their capacity to influence inositol phosphate formation, induced by CCK8 in CHO cells transfected with the rat CCK-B receptor, compound 13 behaved as a full CCK-B antagonist with an IC50 value of 18 +/- 1 nM, being as potent as the antagonist L-365,260 and PD-134,308 (IC50 values respectively, 39 +/- 17 and 30 +/- 2 nM), whereas compound 16 was found to behave as a partial CCK-B agonist. Indeed 16 behaved as an antagonist on the firing rate of rat CA1 hippocampal neurons and acted as an agonist in the pentagastrin stimulated gastric acid secretion (EC50 = 12 nmol/kg) in anesthetized rats. Compound 13 in contrast, was found to inhibit the pentagastrin action at a dose (ID50 = 0.56 mumol/kg) similar to the potent antagonist PD-134,308 (ID50 = 0.4 mumol/kg). The antagonist/agonist properties of compounds 13 and 16 show that both N- and C-terminal substituents modulate the pharmacological properties in the Boc-CCK4 derivatives presented here.

Characterization of the manganese-resistant mutants derived from Vibrio parahaemolyticus.

The virulence and some related factors of Vibrio parahaemolyticus are regulated by the level of iron. In this study, five Mn-resistant mutants were selected after N-methyl-N'-nitrosoguanidine treatment and two transfers in medium containing high levels of manganese chloride. Production of siderophores and the 77-kDa iron-regulated outer-membrane protein and the bacterial growth in these Mn-resistants were deregulated, as compared with the wild-type strain. In addition, the regulation of these phenomena was partially or completely restored by the introduction of Escherichia coli fur gene. Also, the total cellular protein profiles of the wild-type and mutants showed that production of some proteins were positively or negatively regulated by iron, and expression of some of these proteins remained unaffected in these mutants. These results suggested the presence of a complicated iron regulation system, similar to the Fur system of E. coli, in this pathogen.

Conformational analysis of CCK-B agonists using 1H-NMR and restrained molecular dynamics: comparison of biologically active Boc-Trp-(N-Me) Nle-Asp-Phe-NH2 and inactive Boc-Trp-(N-Me)Phe-Asp-Phe-NH2.

The tetrapeptide Boc-Trp-(N-Me)Nle-Asp-Phe-NH2 is a potent CCK-B agonist. Replacement in this analogue of the norleucine residue by a phenylalanine, to yield Boc-Trp-(N-Me) Phe-Asp-Phe-NH2, led to a 740-fold decrease in affinity whereas the same decrease in affinity was not observed in their nonmethylated counterparts. In order to ascertain the conformational preferences of these two N-methylated tetrapeptides, a study by two-dimensional (2D) nmr spectroscopy and molecular modeling was undertaken. The solution conformation of the two peptides was examined by 1H-nmr in a d6-DMSO/H2O (80:20) mixture. A cis-trans equilibrium, induced by N-methylation, was observed for both analogues, and the proton spectra of the two rotamers were fully characterized in each case. 1H-1H distance constraints, derived from 2D nuclear Overhauser effect spectroscopy and rotating frame nuclear Overhauser effect spectroscopy experiments, were used as inputs for subsequent restrained molecular dynamics simulations. Comparisons of the nmr and molecular modeling data point toward distinct conformational preferences for these two peptides with an opposite spatial orientation of the Trp residue, and could explain the large difference in their biological activities. Furthermore, the tridimensional structure of Boc-Trp-(N-Me)Nle-Asp-Phe-NH2 could serve as a model for the design of nonpeptide CCK-B agonists.

Cholecystokinin peptidomimetics as selective CCK-B antagonists: design, synthesis, and in vitro and in vivo biochemical properties.

Antagonists of cholecystokinin-B (CCK-B) receptors have been shown to alleviate CCK4-induced panic attacks in humans and to potentiate opioid effects in animals. The clinical use of these compounds is critically dependent on their ability to cross the blood-brain barrier. In order to improve this property, new, peptoid-derived CCK-B antagonists, endowed with high affinity, selectivity, and increased lipophilicity have been developed. The affinity and selectivity of these compounds have been characterized in vitro and in vivo using guinea pig, rat, and mouse. Most of these compounds proved to be selective for the CCK-B receptor, the most potent analog, N-[N-[(2-adamantyloxy)carbonyl]-D-alpha- methyltryptophanyl]-N-[2-(4-chlorophenyl)ethyl]glycine (26A), having a Ki value of 6.1 nM for guinea pig cortex membranes in vitro and a good selectivity ratio (Ki CCK-A/Ki CCK-B = 174). Furthermore, the in vivo affinity of 26A for mouse brain CCK-B receptors, following intracerebroventricular injection at different concentrations, was found to be 10 nmol. Using competition experiments with the specific CCK-B ligand [3H]pBC 264, compound 26A was shown to cross the blood-brain barrier (0.2%) after intraperitoneal administration in mice. This compound is therefore an interesting pharmacological tool to further elucidate the physiopathological role of endogenous CCK.

CCK-B agonist or antagonist activities of structurally hindered and peptidase-resistant Boc-CCK4 derivatives.

Replacement of Met31 by (N-Me)Nle in CCK8 or CCK4 has been shown to improve the affinity and selectivity for CCK-B receptors. In order to obtain molecules with enhanced bioavailability, two novel series of protected tetrapeptides of the general formula Boc-Trp30-X-Asp-Y33 have been developed. Introduction of (N-Me)Nle and the bulky, aromatic naphthylalaninamide (Nal-NH2) in positions X and Y, respectively, does not greatly modify the affinity for guinea pig brain CCK-B receptors. In contrast, incorporation of hindering N-methyl amino acids such as (N-Me)Phe, (N-Me)Phg, or (N-Me)Chg, but not their non-methylated counterparts, in position X induced a large decrease in affinity for the CCK-B binding sites. Among the various peptides synthesized, Boc-[(N-Me)Nle31,1Nal-NH2(33)]CCK4 (2) (KI = 2.8 nM), Boc-[Phg31,1Nal-NH2(33)]CCK4 (15) (KI = 14 nM), and Boc-[Phg31,1Nal-N(CH3)2(33)]CCK4 (17) (KI = 39 nM) displayed good affinities for brain CCK-B receptors and had good selectivity ratios. These pseudopeptides, in which the presence of unnatural and hydrophobic residues is expected to improve their penetration of the central nervous system, were shown to be very resistant to brain peptidases. Interestingly, whereas compounds 2 and 15 proved to be full agonists for rat hippocampal CCK-B receptors when measured in an electrophysiological assay, compound 17 behaved as a potent antagonist in the same test and displayed a good affinity in rat brain KI(CCK-B) = 51 nM as compared to the Merck antagonist L365,260,KI(CCK-B) = 12 nM. This illustrates a simple means to obtain CCK-B antagonists and suggests that the free, CONH2 group plays a critical role in the recognition of the agonist state of brain CCK-B receptors.

[Studies on structure-activity relationships and receptor binding feature for 3-methylfentanyl derivatives].

Ohmefentanyl is not only a potent analgesic agent, but also a selective ligand for mu opioid receptor. In order to search for more potent analgesics, more selective ligands and long duration of analgesic action in 3-methylfentanyl derivatives, we made modifications of the 1-phenethyl and 4-N-propionyl groups in 3-methylfentanyl and synthesized 15 new compounds. The analgesic activity, duration of analgesic action, receptor binding affinity and opioid sub-receptor selectivity of some of these compounds were measured. Primary pharmacological results showed that most of the compounds in this series possessed morphine-like effects. The analgesic activities of them were about 2-180 times more potent than that of morphine. The duration of analgesic action of the tested compounds was 6-10 times longer than that of fentanyl. The receptor binding affinities (IC50) of compounds 1-4 were 10(-7)-10(-8) mol. Compound 13 displayed the best binding selectivity on mu opioid receptor site with ratio mu/delta greater than 700 in rat brain membrane and mu/delta = 1000 in mouse brain membrane. The new compound may be proposed as a useful tool in studying the opioid receptor. According to the result of equations in reference 11, compounds 7-14 were designed and prepared. The observed log 1/c values of them were much closer to the calculated values.

Differences between Indica and Japonica rice varieties in CO2 exchange rates in response to leaf nitrogen and temperature.

Four Indica and five Japonica varieties of rice (Oryza sativa L.) were examined to elucidate their differences in photosynthetic activity and dark respiratory rate as influenced by leaf nitrogen levels and temperatures. The photosynthetic rates of single leaf showed correlations with total nitrogen and soluble protein contents in the leaves. Respiratory rate was also positively correlated with the leaf nitrogen content. When compared at the same level of leaf nitrogen or soluble protein content, the four Indica varieties and one of Japonica varieties, Tainung 67, which have some Indica genes derived from one of its parents, showed higher photosynthetic rates than the remaining four Japonica varieties. At the same photosynthetic rate, the Indica varieties showed lower respiratory rate than Japonica varieties. When the leaf temperature rose from 20°C to 30°C, the photosynthetic rate increased by 18 to 41%, whereas the respiratory rate increased by 100 to 150%. These increasing rates in response to temperature were higher in the Japonica than in the Indica varieties. In this respect, Tainung 67 showed the same behavior as of the other four Japonica varieties.