Substantial rate enhancements of the esterification reaction of phthalic anhydride with methanol at high pressure and using supercritical CO2 as a co-solvent in a glass microreactor.

The esterification reaction of phthalic anhydride with methanol was performed at different temperatures in a continuous flow glass microreactor at pressures up to 110 bar and using supercritical CO(2) as a co-solvent. The design is such that supercritical CO(2) can be generated inside the microreactor. Substantial rate enhancements were obtained, viz. a 53-fold increase was obtained at 110 bar and 60 degrees C. Supercritical CO(2) as a co-solvent gave rise to a 5400-fold increase (both with respect to batch experiments at 1 bar at the same temperature).

Differences in the response of a striped dolphin (Stenella coeruleoalba) and a harbour porpoise (Phocoena phocoena) to an acoustic alarm.

Small cetacean bycatch in gillnet fisheries may be reduced by deterring odontocetes from nets acoustically. However, different odontocete species may respond differently to acoustic signals from alarms. Therefore, in this study a striped dolphin and a harbour porpoise were subjected simultaneously to sounds produced by the XP-10 experimental acoustic alarm. The alarm produced 0.3s tonal signals randomly selected from a set of 16 with fundamental frequencies between 9 and 15kHz, with a constant pulse interval of 4.0s (duty cycle 8%) and a Source Level range of 133-163dB re 1muPa (rms). The effect of the alarm was judged by comparing the animals' respiration rate and position relative to the alarm during test periods with those during baseline periods. As in a previous study on two porpoises with the same alarm, the porpoise in the present study reacted strongly to the alarm by swimming away from it and increasing his respiration rate. The striped dolphin, however, showed no reaction to the active alarm. Based on harbour porpoise audiograms and the specific audiogram of the striped dolphin in the present study, and the low background noise levels during the experiment, both animals must have heard the alarm signals clearly. This study indicates that cetacean species are not equally sensitive to human-made noise disturbance. Therefore, source levels of acoustic alarms should be adapted to the species they are supposed to deter. In addition, alarms should be tested on each odontocete species for which they are intended to reduce bycatch.

Multi-podant diglycolamides and room temperature ionic liquid impregnated resins: An excellent combination for extraction chromatography of actinides.

Extraction chromatography resins, prepared by impregnating two multi-podant diglycolamide ligands, viz. diglycolamide-functionalized calix[4]arene (C4DGA) and tripodal diglycolamide (T-DGA) dissolved in the room temperature ionic liquid 1-butyl-3-methylimidazolium bis(trifluoromethanesulfonyl)amide (RTIL: C4mimTf2N) on Chromosorb-W (an inert solid support), gave excellent results for the removal of trivalent actinides from acidic waste solutions. Distribution coefficient measurements on several metal ions showed selective sorption of Am(III) over hexavalent uranyl ions and other fission product elements such as strontium and cesium. The sorbed metal ions could be efficiently desorbed with a complexing solution containing guanidine carbonate and EDTA buffer. The sorption of Am(III) on both resins followed pseudo-second order rate kinetics with rate constants of 1.37×10(-6) and 6.88×10(-7)g/cpmmin for T-DGA and C4DGA resins, respectively. The metal sorption on both resins indicated the Langmuir monolayer chemisorption phenomenon with Eu(III) sorption capacities of 4.83±0.21 and 0.52±0.05mg per g of T-DGA and C4DGA resins, respectively. The results of column studies show that these resins are of interest for a possible application for the recovery of hazardous trivalent actinides from dilute aqueous solutions.

The influence of acoustic emissions for underwater data transmission on the behaviour of harbour porpoises (Phocoena phocoena) in a floating pen.

To prevent grounding of ships and collisions between ships in shallow coastal waters, an underwater data collection and communication network is currently under development: Acoustic Communication network for Monitoring of underwater Environment in coastal areas (ACME). Marine mammals might be affected by ACME sounds since they use sounds of similar frequencies (around 12 kHz) for communication, orientation, and prey location. If marine mammals tend to avoid the vicinity of the transmitters, they may be kept away from ecologically important areas by ACME sounds. One marine mammal species that may be affected in the North Sea is the harbour porpoise. Therefore, as part of an environmental impact assessment program, two captive harbour porpoises were subjected to four sounds, three of which may be used in the underwater acoustic data communication network. The effect of each sound was judged by comparing the animals' positions and respiration rates during a test period with those during a baseline period. Each of the four sounds could be made a deterrent by increasing the amplitude of the sound. The porpoises reacted by swimming away from the sounds and by slightly, but significantly, increasing their respiration rate. From the sound pressure level distribution in the pen, and the distribution of the animals during test sessions, discomfort sound level thresholds were determined for each sound. In combination with information on sound propagation in the areas where the communication system may be deployed, the extent of the 'discomfort zone' can be estimated for several source levels (SLs). The discomfort zone is defined as the area around a sound source that harbour porpoises are expected to avoid. Based on these results, SLs can be selected that have an acceptable effect on harbour porpoises in particular areas. The discomfort zone of a communication sound depends on the selected sound, the selected SL, and the propagation characteristics of the area in which the sound system is operational. In shallow, winding coastal water courses, with sandbanks, etc., the type of habitat in which the ACME sounds will be produced, propagation loss cannot be accurately estimated by using a simple propagation model, but should be measured on site. The SL of the communication system should be adapted to each area (taking into account bounding conditions created by narrow channels, sound propagation variability due to environmental factors, and the importance of an area to the affected species). The discomfort zone should not prevent harbour porpoises from spending sufficient time in ecologically important areas (for instance feeding areas), or routes towards these areas.

Redox cycling of potential antitumor aziridinyl quinones.

The formation of reactive oxygen intermediates (ROI) during redox cycling of newly synthesized potential antitumor 2,5-bis (1-aziridinyl)-1,4-benzoquinone (BABQ) derivatives has been studied by assaying the production of ROI (superoxide, hydroxyl radical, and hydrogen peroxide) by xanthine oxidase in the presence of BABQ derivatives. At low concentrations (< 10 microM) some BABQ derivatives turned out to inhibit the production of superoxide and hydroxyl radicals by xanthine oxidase, while the effect on the xanthine-oxidase-induced production of hydrogen peroxide was much less pronounced. Induction of DNA strand breaks by reactive oxygen species generated by xanthine oxidase was also inhibited by BABQ derivatives. The DNA damage was comparable to the amount of hydroxyl radicals produced. The inhibiting effect on hydroxyl radical production can be explained as a consequence of the lowered level of superoxide, which disrupts the Haber-Weiss reaction sequence. The inhibitory effect of BABQ derivatives on superoxide formation correlated with their one-electron reduction potentials: BABQ derivatives with a high reduction potential scavenge superoxide anion radicals produced by xanthine oxidase, leading to reduced BABQ species and production of hydrogen peroxide from reoxidation of reduced BABQ. This study, using a unique series of BABQ derivatives with an extended range of reduction potentials, demonstrates that the formation of superoxide and hydroxyl radicals by bioreductively activated antitumor quinones can in principle be uncoupled from alkylating activity.

Synthesis, mechanism of action, and biological evaluation of mitosenes.

Mitosenes of both the pyrrolo- and pyrido[1,2-a]indole type have been prepared via modification of these heterotricyclic compounds. Several mitosenes have been studied for their reactions with nucleophiles under reductive conditions. The results of these experiments show that the biological activity of mitosenes is based on the mechanism of bioreductive activation. When both leaving groups at C-1 and C-10 in the mitosene are the same, the nucleophile preferably adds to C-10 under reductive conditions. All mitosenes were studied for their biological activities in vitro against L1210, WiDr, and A204. On the basis of these results a selection of three mitosenes was made for a more detailed biological evaluation. Several tumor model systems were used, viz. P388, human tumor xenografts, MAC 13, and MAC 16. The results of these studies show that mitosenes have a more limited range of activities than mitomycin C. Surprisingly, the in vivo activities of mitosene diol 8b and mitosene diacetate 10b against the gastric human tumor xenograft GXF 97 were very high and comparable with that of mitomycin C.

Reductive activation of potential antitumor mitosene compounds.

The reductive activation of mitosene compounds was studied with cyclic voltammetry and HPLC analysis. Reduction of mitosenes, possessing good leaving groups at C-1 and C-10, was shown to result in loss of these groups at pH 7.0 and pH 6.0. The loss of leaving groups from mitosenes occurred faster at lower pH. Mitosenes without good leaving groups were found to be stable upon reduction. In the presence of acetoxy groups at C-1 and C-10, the C-10 site is the most reactive site upon reductive activation. This is opposite to the case of mitomycin C, where the C-1 site is the first to react upon reduction. At pH 6.0 without reduction, acid degradation also caused the loss of leaving groups of mitosenes, although at a very slow rate. In contrast to reductive activation, upon acid degradation of a diacetoxymitosene the C-1 group appeared to be lost faster. Electrochemical as well as dithionite reduction of a bifunctional (diacetoxy) mitosene compound in the presence of calf thymus DNA at pH 5.5 resulted in the formation of DNA interstrand cross-links. Depending on activation method, this diacetoxymitosene was at least as efficient in DNA cross-linking as mitomycin C under comparable conditions.

In vivo activity and hydrophobicity of cytostatic aziridinyl quinones.

For a series of 3,6-disubstituted bisaziridinylbenzoquinones the in vivo and in vitro activities against murine tumors, as well as the in vivo toxicity, are analyzed. Properties describing biochemical and physicochemical reactions are also incorporated in the analyses. The important 1-octanol/water partition coefficients were determined, using a fast variation of the shake flask method. New pi'-values were calculated for the substituents in this series. These quinone pi'-values deviate strongly from the standard pi-values, especially for hydrogen-bonding substituents. To discriminate between the toxic and therapeutic activity of the compounds, principal components and partial least squares analyses were applied. Evidence is presented for selective antitumor action of the investigated compounds. The L1210 clonogenic assay only seems to relate to the general cytotoxicity and has no predictive value for in vivo activity for these compounds. The activity is correlated to the hydrophobicity of the quinones. The toxicity correlates with the ease of reduction, contrary to the hypothesis of bioreductive activation as a mechanism for selectivity.

Microsomal superoxide anion production and NADPH-oxidation in a series of 22 aziridinylbenzoquinones.

Several 2,5-bis(1-aziridinyl)-1,4-benzoquinones (BABQs) can be activated to alkylating species by reduction of the quinone moiety. On the other hand, cytotoxicity of these compounds can be induced by redox cycling. A series of BABQs and their methylated analogues (BMABQs) with different substituents at the 3- and 6-position was synthesized in order to investigate the influence of the substituents on the reduction of the quinone moiety and on the generation of superoxide anion radicals with rat liver microsomes. Superoxide anion production (SAP) ranged from 3.7 +/- 0.1 to 742 +/- 74 nmoles/min/mg protein with quinone concentrations of 10 nmoles/ml. NADPH-oxidation was measured under the same conditions and it correlated well (r = 0.88, P less than 0.001) with SAP. It ranged from 1.4 +/- 0.2 to 494 +/- 60 nmoles/min/mg protein. SAP for 22 B(M) ABQs showed a good correlation with the summated electronic substituent constant sigma para.total (r = 0.86, P less than 0.001). It can be concluded that superoxide anion production by 22 B(M)ABQs in rat liver microsomes can be predicted from structural features of the compounds.

Potential antitumour mitosenes: relationship between in vitro DNA interstrand cross-link formation and DNA damage in Escherichia coli K-12 strains.

This investigation was aimed at determining the possible relationship between DNA interstrand cross-linking and the cytotoxic activity of potential antitumour mitosene compounds. Mitosenes, possessing two good leaving groups at C-1 and C-10, were found to be able to cross-link calf thymus DNA under hypoxic conditions following sodium dithionite (Na2S2O4) reduction at pH 7.0 and pH 5.5. DNA interstrand cross-linking was pH dependent for most of the mitosenes used, with a higher amount of cross-links formed at pH 5.5 compared to pH 7.0. Without reduction or under aerobic conditions no cross-link formation was detected. The importance of DNA damage for the toxic effect of these mitosenes was assayed by comparing the survival in a DNA repair deficient and a DNA repair proficient E. coli K-12 strain. A correlation between the number of cross-links formed in calf thymus DNA in vitro and the IC50 values in the DNA repair deficient E. coli strain was found. The effect of hypoxia on toxicity of mitosenes was studied in Chinese hamster V79 cells. In these cells, mitosenes appeared to be very active. Under severe hypoxic conditions toxicity of these mitosenes increased, most likely due to the increased lifetime of the activated mitosene species as compared to aerobic conditions. The results suggest that DNA cross-linking following reductive activation is important for the eventual activity of mitosenes in a bacterial system. Increased activity of mitosenes under hypoxic conditions in the V79 cells indicates that these mitosenes may be more active in hypoxic parts of tumours.

Reduction of antitumour mitosenes in non-aqueous and aqueous environment. An electron spin resonance and cyclic voltammetry study.

Chemical reduction of mitosenes under aerobic conditions in DMSO showed characteristic ESR signals of the mitosene derived semiquinone free radicals. However, these signals diminished strongly upon addition of water to the reaction mixture, indicating a short lifetime of the mitosene semiquinone free radicals under aqueous conditions. In addition, enzymatic one-electron reduction of these mitosenes with either xanthine oxidase or purified NADPH cytochrome P450 reductase under anaerobic conditions showed no signals of the mitosene semiquinone free radicals. Subsequent cyclic voltammetry measurements demonstrated facilitation of the further one-electron reduction of the mitosene semiquinone free radicals in the presence of water in comparison with non-aqueous conditions. The present results strongly suggest that in the presence of water relatively stable hydroquinones are formed upon reduction of mitosenes. Consequently, the steady state concentrations of mitosene semiquinone free radicals will be lowered substantially in aqueous environment. Thus under physiological conditions, two-electron reduction and formation of the mitosene hydroquinone might be important in processes leading to DNA alkylation by these mitosenes.

Electrochemistry of potential bioreductive alkylating quinones: its use in the development of new aziridinylquinones.

The concept of bioreductive alkylation as a mechanism of action of quinone-containing anticancer agents was investigated, using electrochemical techniques. According to this concept, an electrochemical step (reduction of the quinone ring) is followed by one or more chemical steps, leading to formation of the actual alkylating species. The proper use of electrochemical analysis of potential bioreductive alkylating quinones in the design of new analogs is limited. Up to now, the only electrochemical parameter frequently used in structure-activity relationship studies, is the half-wave potential of the quinone reduction. However, reliable information can only be obtained from the found value of this parameter when the reduction mechanism has been elucidated. Furthermore, it only gives information about the first step of the model. More detailed electrochemical analysis of potential bioreductive alkylating quinones, in combination with a biological evaluation, is required to gain more insight in their mechanism of action and to yield quantitative information about substituent effects on both the electrochemical and the chemical step(s) of the model. Results of such studies of a series of aziridinylquinones indicate, that the biological activity in vitro is correlated with the ease of protonation of the aziridines after quinone reduction, which is in accordance with the concept of bioreductive activation. No correlation with the ease of protonation of the aziridines prior to quinone reduction or with the quinone reduction step itself can be found.

DNA alkylation and formation of DNA interstrand cross-links by potential antitumour 2,5-bis(1-aziridinyl)-1,4-benzoquinones.

A series of 3,6-substituted 2,5-bis(1-aziridinyl)-1,4-benzoquinone derivatives was shown to alkylate calf thymus DNA and to form DNA interstrand cross-links. Alkylation and cross-link formation were enhanced after electrochemical reduction of the compounds and increased with lower pH in the pH range from 4.5 to 8.0. Reduction especially shifts the pH at which cross-linking and alkylation occurs to higher values, which are more physiologically relevant. This shift is probably caused by the increase in pKa value of the aziridine ring after reduction of the quinone moiety. The inactivation of single-stranded bacteriophage M13mp19 DNA to form phages in an E. coli host, by the 3,6-unsubstituted parent compound 2,5-bis(1-aziridinyl)-1,4-benzoquinone (TW13) was dependent upon reduction and pH in a similar way as was alkylation. The compound in our series with the least bulky, 3,6-substituents, TW13, caused a high amount of cross-link formation. Compounds with methyl-substituted aziridine rings showed low cross-linking ability. Our results support the concept that the protonated reduced compound is the reactive species that alkylates DNA, and that steric factors play an important role in the reactivity towards DNA. A correlation is observed between the ability to induce DNA interstrand cross-links and inactivation of M13mp19 bacteriophage DNA. Cross-link formation was also demonstrated in E. coli K12 cells, where the compounds are reduced endogenously by bacterial reductases.

Covalent binding studies on the 14C-labeled antitumor compound 2,5-bis(1-aziridinyl)-1,4-benzoquinone. Involvement of semiquinone radical in binding to DNA, and binding to proteins and bacterial macromolecules in situ.

2,5-Bis(1-aziridinyl)-1,4-benzoquinone (BABQ) is a compound from which several antitumour drugs are derived, such as Trenimone, Carboquone and Diaziquone (AZQ). The mechanism of DNA binding of BABQ was studied using 14C-labeled BABQ and is in agreement with reduction of the quinone moiety and protonation of the aziridine ring, followed by ring opening and alkylation. The one-electron reduced (semiquinone) form of BABQ alkylates DNA more efficiently than two-electron reduced or non reduced BABQ. Covalent binding to polynucleotides did not unambiguously reveal preference for binding to specific DNA bases. Attempts to elucidate further the molecular structure of DNA adducts by isolation of modified nucleosides from enzymatic digests of reacted DNA failed because of instability of the DNA adducts. The mechanism of covalent binding to protein (bovine serum albumin, BSA) appeared to be completely different from that of covalent binding to DNA. Binding of BABQ to BSA was not enhanced by reduction of the compound and was pH dependent in a way that is opposite to that of DNA alkylation. Glutathione inhibits binding of BABQ to BSA and forms adducts with BABQ in a similar pH dependence as the protein binding. The aziridine group therefore does not seem to be involved in the alkylation of BSA. Incubation of intact E. coli cells, which endogenously reduce BABQ, resulted in binding to both DNA and RNA, but also appreciable protein binding was observed.

Evaluation of several multiple diglycolamide-functionalized calix[4]arene ligands for the isolation of carrier free 90Y from 90Sr.

Several diglycolamide-functionalized calix[4]arenes containing four and eight diglycolamide (DGA) moieties were evaluated for their relative extraction efficiencies towards Y(III) and Sr(II). Ligands containing four DGA units with n-propyl, iso-pentyl, and n-octyl groups at the amidic N atom adjacent to the calix[4]arene skeleton showed efficient extraction of Y(III) from 3M HNO3. The extraction of Sr(II) was poor in all cases in the entire acidity range (0.1-6M HNO3) studied. The ligands with a hydrogen atom and an n-propyl group at the concerning amidic N atom showed a very high separation efficiency as reflected in separation factor (S.F.=DY/DSr) values in the range of 10(5)-10(6). A method was developed for the separation of carrier-free (90)Y from a (90)Y-(90)Sr mixture involving consecutive extraction-stripping cycles. The product purity was checked using half-life measurements. Two consecutive cycles of extraction and stripping were found to be sufficient for obtaining pure (90)Y. The results obtained in the present studies were compared with those obtained previously using analogous ligands such as TODGA (N,N,N',N'-tetraoctyl diglycolamide), T2EHDGA (N,N,N',N'-tetra-2-ethylhexyl diglycolamide), and PC-88A (bis(2-ethylhexyl) phosphonic acid).

Cavitand zn(II)-porphyrin capsules with high affinities for pyridines and N-methylimidazole.

Covalent cavitand Zn(II)-porphyrins 17-20 were prepared via multistep syntheses. These host molecules show moderate to excellent binding affinities to N-methylimidazole and pyridine guests. The complexing behavior strongly depends on the spacer's length, number, and rigidity, in addition to the guest size. Cavitand capping and strapping of porphyrins strongly influence the complex formation and result in a 10-700-fold enhancement of the binding strength compared to tetraphenyl Zn(II)-porphyrin.

Synthesis of 18F-labelled 2-fluoro-1,4-quinones using acetylhypofluorite.

The fluorination of 1,4-benzo- and naphthoquinones using [18F]acetylhypofluorite is described. For compounds with electron-donating substituents fair to good radiochemical yields have been reached.

Quantitative structure activity relationship for the acute cytotoxicity of 13 (bis)aziridinyl-benzoquinones: relation to cellular ATP depletion.

This study was performed to establish relationships between the structure of 2,5-bis(1-aziridinyl)-1,4-benzoquinones (BABQs) bearing different substituents at the 3- and 6-position and their acute toxic effects in rat hepatocytes. The cell viability, loss of cellular glutathione (GSH+GSSG) and loss of ATP were followed during 4 h of incubation of freshly isolated hepatocytes. The toxicity of these compounds (100 microM) was predicted better by their reactivity with GSH than by their redox cycling in rat liver microsomes. The time of 50% loss of viability (LT50) correlated very well with the time of 50% depletion of ATP (AT50). LT50 could be adequately predicted by using the electronic field parameter (Ftotal) describing the electron withdrawing or donating properties for all the substituents on the quinone-nucleus. 7-(Di)halogen-substituted BABQs that all very rapidly depleted cellular glutathione showed significant differences in AT50 as well as in LT50. This suggests that alterations in ATP levels are important for explaining the differences in cytotoxicity of these compounds.