Analysis of zinc oxide nanoparticles as a potential mutagen of respiratory epithelia

Metallische Nanopartikel und ihre Oxide (z.B. ZnO NP, TiO2 NP und Fe2O3 NP) werden aufgrund ihrer chemischen und physikalischen Eigenschaften häufig als Additive in der Reifenproduktion, in Katalysatoren, Lebensmitteln, Arzneimitteln und Kosmetikprodukten verwendet. Künftig wird ein kontinuierlicher Anstieg der industriellen Anwendung (~ 1663 Tonnen im Jahr 2025) mit gesteigerter Freisetzung in die Umwelt erwartet, was zwangsläufig zu einer vermehrten Aufnahme über das respiratorische Epithel führt. Metalldampffieber ist als gesundheitsschädigender Effekt von Metalloxid-haltigen Aerosolen (z.B. ZnO) nach Inhalation bekannt. Immunreaktionen, wie beispielsweise Entzündungen, werden häufig mit der Entstehung von Sauerstoffradikalen (ROS) in Verbindung gebracht, die wiederum zu DNA-Schäden führen können. Drei mögliche Ursachen der Genotoxität werden angenommen: direkte Interaktion von Nanopartikeln mit intrazellulären Strukturen, Interaktion von Ionen dissoziierter Partikel mit intrazellulären Strukturen sowie die Entstehung von ROS initiiert durch Partikel oder Ionen.rnDie vorliegende Studie befasst sich mit den Mechanismen der Genotoxizität von ZnO Nanopartikeln (ZnO NP), als Beispiel für metallische Nanopartikel, im respiratorischen Epithel. In der Studie wurde gezielt die intrazelluläre Aufnahme und Verteilung von ZnO NP, deren Toxizität, deren DNA schädigendes Potential sowie die Aktivierung der DNA damage response (DDR) analysiert.rnEs konnten kaum internalisierte ZnO NP mittels TEM detektiert werden. Innerhalb der ersten Sekunden nach Behandlung mit ZnO NP wurde spektrofluorometrisch ein starker Anstieg der intrazellulären Zn2+ Konzentration gemessen. In unbehandelten Zellen war Zn2+ in granulären Strukturen lokalisiert. Die Behandlung mit ZnO NP führte zu einer Akkumulation von Zn2+ in diesen Strukturen. Im zeitlichen Verlauf verlagerten sich die Zn2+-Ionen in das Zytoplasma, sowie in Zellkerne und Mitochondrien. Es wurde keine Kolokalisation von Zn2+ mit den frühen Endosomen und dem endoplasmatischen Retikulum beobachtet. Die Vorbehandlung der Zellen mit Diethylen-triaminpentaessigsäure (DTPA), als extrazellulärem Komplexbildner, verhinderte den intrazellulären Anstieg von Zn2+ nach Behandlung mit den Partikeln.rnDie Behandlung mit ZnO NP resultierte in einer zeit- und dosisabhängigen Reduktion der zellulären Viabilität, während die intrazelluläre ROS-Konzentrationen in den ersten 30 min leicht und anschließend kontinuierlich bis zum Ende der Messung anstiegen. Außerdem verringerte sich das mitochondriale Membranpotential, während sich die Anzahl der frühapoptotischen Zellen in einer zeitabhängigen Weise erhöhte. rnDNA Doppelstrangbrüche (DNA DSB) wurden mittels Immunfluoreszenz-Färbung der γH2A.X foci sichtbar gemacht und konnten nach Behandlung mit ZnO NP detektiert werden. Die Vorbehandlung mit dem Radikalfänger N-Acetyl-L-Cytein (NAC) resultierte in stark reduzierten intrazellulären ROS-Konzentrationen sowie wenigen DNA DSB. Die DNA Schädigung wurde durch Vorbehandlung mit DTPA ganz verhindert.rnDie Aktivierung der DDR wurde durch die Analyse von ATM, ATR, Chk1, Chk2, p53 und p21 mittels Western Blot und ELISA nach Behandlung mit ZnO NP überprüft. Der ATR/Chk1 Signalweg wurde durch ZnO NP nicht aktiviert. Die Komplexierung von Zn2+ resultierte in einer verminderten ATM/Chk2 Signalwegaktivierung. Es zeigte sich, dass das Abfangen von ROS keinen Effekt auf die ATM/Chk2 Signalwegaktivierung hatte.rnZusammengefasst wurde festgestellt, dass die Exposition mit ZnO NP in der Entstehung von ROS, reduzierter Viabilität und vermindertem mitochondrialem Membranpotential resultiert, sowie zeitabhängig eine frühe Apoptose initiiert. ZnO NP dissoziierten extrazellulär und wurden schnell als Zn2+ über unbekannte Mechanismen internalisiert. Die Zn2+-Ionen wurden im Zytoplasma, sowie besonders in den Mitochondrien und dem Zellkern, akkumuliert. Die DDR Signalgebung wurde durch ZnO NP aktiviert, jedoch nicht durch NAC inhibiert. Es wurde gezeigt, dass DTPA die DDR Aktivierung komplett inhibierte. Die Behandlung mit ZnO NP induzierte DNA DSB. Die Inhibition von ROS reduzierte die DNA DSB und die Komplexierung der Zn2+ verhinderte die Entstehung von DNA DSB.rnDiese Daten sprechen für die Dissoziation der Partikel und die hierbei freigesetzten Zn2+ als Hauptmediator der Genotoxizität metallischer Nanopartikel. rn

Matrix metalloproteinases and angiogenic factors: predictors of survival after radical prostatectomy for clinically organ-confined prostate cancer?

The aim of the present study was to investigate whether biomarkers improve the prediction of recurrence-free, disease-specific, and overall survival in patients with clinically localized prostate cancer. A tissue microarray was constructed from prostate specimens of 278 patients who underwent open radical retropubic prostatectomy for clinically localized prostate cancer. For immunohistochemical studies, antibodies were used against matrix metalloproteinase (MMP)-2, MMP-3, MMP-7, MMP-9, MMP-13, and MMP-19, as well as against vascular endothelial growth factor, hypoxia-induced factor 1 , basic fibroblast growth factor, and cluster of differentiation 31. Univariate and multivariable analyses were performed to evaluate the potential predictors of overall, disease-specific, and recurrence-free survival. In univariate analysis of patients with clinically organ-confined prostate cancer, only higher expression levels of MMP-9 (hazard ratio [0.6], 95% CI 0.45-0.8) had a protective effect in terms of overall survival. This positive effect of high MMP-9 expression was also observed for recurrence-free (HR 0.88, 95% CI 0.78-0.99) and disease-specific survival (HR 0.5, 95% CI 0.36-0.73). In multivariable analysis, none of these potential markers was found to be an independent prognostic factor of survival. Of all MMPs and angiogenic factors tested, MMP-9 expression has the potential as a prognostic marker in patients undergoing radical prostatectomy for clinically organ-confined cases of prostate cancer.

Exploration of the Potential Energy Surfaces, Prediction of Atmospheric Concentrations, and Prediction of Vibrational Spectra for the HO2···(H2O)n (n = 1−2) Hydrogen Bonded Complexes

The hydroperoxy radical (HO2) plays a critical role in Earth's atmospheric chemistry as a component of many important reactions. The self-reaction of hydroperoxy radicals in the gas phase is strongly affected by the presence of water vapor. In this work, we explore the potential energy surfaces of hydroperoxy radicals hydrogen bonded to one or two water molecules, and predict atmospheric concentrations and vibrational spectra of these complexes. We predict that when the HO2 concentration is on the order of 108molecules·cm-3 at 298 K, that the number of HO2···H2O complexes is on the order of 107molecules·cm-3 and the number of HO2···(H2O)2 complexes is on the order of 106molecules·cm-3. Using the computed abundance of HO2···H2O, we predict that, at 298 K, the bimolecular rate constant for HO2···H2O + HO2 is about 10 times that for HO2 + HO2.

Systematic Review and Cumulative Analysis of Oncologic and Functional Outcomes After Robot-assisted Radical Cystectomy

CONTEXT Although open radical cystectomy (ORC) is still the standard approach, laparoscopic radical cystectomy (LRC) and robot-assisted radical cystectomy (RARC) are increasingly performed. OBJECTIVE To report on a systematic literature review and cumulative analysis of pathologic, oncologic, and functional outcomes of RARC in comparison with ORC and LRC. EVIDENCE ACQUISITION Medline, Scopus, and Web of Science databases were searched using a free-text protocol including the terms robot-assisted radical cystectomy or da Vinci radical cystectomy or robot* radical cystectomy. RARC case series and studies comparing RARC with either ORC or LRC were collected. A cumulative analysis was conducted. EVIDENCE SYNTHESIS The searches retrieved 105 papers, 87 of which reported on pathologic, oncologic, or functional outcomes. Most series were retrospective and had small case numbers, short follow-up, and potential patient selection bias. The lymph node yield during lymph node dissection was 19 (range: 3-55), with half of the series following an extended template (yield range: 11-55). The lymph node-positive rate was 22%. The performance of lymphadenectomy was correlated with surgeon and institutional volume. Cumulative analyses showed no significant difference in lymph node yield between RARC and ORC. Positive surgical margin (PSM) rates were 5.6% (1-1.5% in pT2 disease and 0-25% in pT3 and higher disease). PSM rates did not appear to decrease with sequential case numbers. Cumulative analyses showed no significant difference in rates of surgical margins between RARC and ORC or RARC and LRC. Neoadjuvant chemotherapy use ranged from 0% to 31%, with adjuvant chemotherapy used in 4-29% of patients. Only six series reported a mean follow-up of >36 mo. Three-year disease-free survival (DFS), cancer-specific survival (CSS), and overall survival (OS) rates were 67-76%, 68-83%, and 61-80%, respectively. The 5-yr DFS, CSS, and OS rates were 53-74%, 66-80%, and 39-66%, respectively. Similar to ORC, disease of higher pathologic stage or evidence of lymph node involvement was associated with worse survival. Very limited data were available with respect to functional outcomes. The 12-mo continence rates with continent diversion were 83-100% in men for daytime continence and 66-76% for nighttime continence. In one series, potency was recovered in 63% of patients who were evaluable at 12 mo. CONCLUSIONS Oncologic and functional data from RARC remain immature, and longer-term prospective studies are needed. Cumulative analyses demonstrated that lymph node yields and PSM rates were similar between RARC and ORC. Conclusive long-term survival outcomes for RARC were limited, although oncologic outcomes up to 5 yr were similar to those reported for ORC. PATIENT SUMMARY Although open radical cystectomy (RC) is still regarded as the standard treatment for muscle-invasive bladder cancer, laparoscopic and robot-assisted RCs are becoming more popular. Templates of lymph node dissection, lymph node yields, and positive surgical margin rates are acceptable with robot-assisted RC. Although definitive comparisons with open RC with respect to oncologic or functional outcomes are lacking, early results appear comparable.

Immediate versus deferred chemotherapy after radical cystectomy in patients with pT3-pT4 or N+ M0 urothelial carcinoma of the bladder (EORTC 30994): an intergroup, open-label, randomised phase 3 trial

BACKGROUND Patients with muscle-invasive urothelial carcinoma of the bladder have poor survival after cystectomy. The EORTC 30994 trial aimed to compare immediate versus deferred cisplatin-based combination chemotherapy after radical cystectomy in patients with pT3-pT4 or N+ M0 urothelial carcinoma of the bladder. METHODS This intergroup, open-label, randomised, phase 3 trial recruited patients from hospitals across Europe and Canada. Eligible patients had histologically proven urothelial carcinoma of the bladder, pT3-pT4 disease or node positive (pN1-3) M0 disease after radical cystectomy and bilateral lymphadenectomy, with no evidence of any microscopic residual disease. Within 90 days of cystectomy, patients were centrally randomly assigned (1:1) by minimisation to either immediate adjuvant chemotherapy (four cycles of gemcitabine plus cisplatin, high-dose methotrexate, vinblastine, doxorubicin, and cisplatin [high-dose MVAC], or MVAC) or six cycles of deferred chemotherapy at relapse, with stratification for institution, pT category, and lymph node status according to the number of nodes dissected. Neither patients nor investigators were masked. Overall survival was the primary endpoint; all analyses were by intention to treat. The trial was closed after recruitment of 284 of the planned 660 patients. This trial is registered with ClinicalTrials.gov, number NCT00028756. FINDINGS From April 29, 2002, to Aug 14, 2008, 284 patients were randomly assigned (141 to immediate treatment and 143 to deferred treatment), and followed up until the data cutoff of Aug 21, 2013. After a median follow-up of 7·0 years (IQR 5·2-8·7), 66 (47%) of 141 patients in the immediate treatment group had died compared with 82 (57%) of 143 in the deferred treatment group. No significant improvement in overall survival was noted with immediate treatment when compared with deferred treatment (adjusted HR 0·78, 95% CI 0·56-1·08; p=0·13). Immediate treatment significantly prolonged progression-free survival compared with deferred treatment (HR 0·54, 95% CI 0·4-0·73, p<0·0001), with 5-year progression-free survival of 47·6% (95% CI 38·8-55·9) in the immediate treatment group and 31·8% (24·2-39·6) in the deferred treatment group. Grade 3-4 myelosuppression was reported in 33 (26%) of 128 patients who received treatment in the immediate chemotherapy group versus 24 (35%) of 68 patients who received treatment in the deferred chemotherapy group, neutropenia occurred in 49 (38%) versus 36 (53%) patients, respectively, and thrombocytopenia in 36 (28%) versus 26 (38%). Two patients died due to toxicity, one in each group. INTERPRETATION Our data did not show a significant improvement in overall survival with immediate versus deferred chemotherapy after radical cystectomy and bilateral lymphadenectomy for patients with muscle-invasive urothelial carcinoma. However, the trial is limited in power, and it is possible that some subgroups of patients might still benefit from immediate chemotherapy. An updated individual patient data meta-analysis and biomarker research are needed to further elucidate the potential for survival benefit in subgroups of patients. FUNDING Lilly, Canadian Cancer Society Research.

How often parametrial involvement leads to post-operative adjuvant treatment in locally advanced cervical cancer after neoadjuvant chemotherapy and type C radical hysterectomy?

OBJECTIVE Parametrial involvement (PMI) is one of the most important factors influencing prognosis in locally advanced stage cervical cancer (LACC) patients. We aimed to evaluate PMI rate among LACC patients undergoing neoadjuvant chemotherapy (NACT), thus evaluating the utility of parametrectomy in tailor adjuvant treatments. METHODS Retrospective evaluation of consecutive 275 patients affected by LACC (IB2-IIB), undergoing NACT followed by type C/class III radical hysterectomy. Basic descriptive statistics, univariate and multivariate analyses were applied in order to identify factors predicting PMI. Survival outcomes were assessed using Kaplan-Meier and Cox models. RESULTS PMI was detected in 37 (13%) patients: it was associated with vaginal involvement, lymph node positivity and both in 10 (4%), 5 (2%) and 12 (4%) patients, respectively; while PMI alone was observed in only 10 (4%) patients. Among this latter group, adjuvant treatment was delivered in 3 (1%) patients on the basis of pure PMI; while the remaining patients had other characteristics driving adjuvant treatment. Considering factors predicting PMI we observed that only suboptimal pathological responses (OR: 1.11; 95% CI: 1.01, 1.22) and vaginal involvement (OR: 1.29 (95%) CI: 1.17, 1.44) were independently associated with PMI. PMI did not correlate with survival (HR: 2.0; 95% CI: 0.82, 4.89); while clinical response to NACT (HR: 3.35; 95% CI: 1.59, 7.04), vaginal involvement (HR: 2.38; 95% CI: 1.12, 5.02) and lymph nodes positivity (HR: 3.47; 95% CI: 1.62, 7.41), independently correlated with worse survival outcomes. CONCLUSIONS Our data suggest that PMI had a limited role on the choice to administer adjuvant treatment, thus supporting the potential embrace of less radical surgery in LACC patients undergoing NACT. Further prospective studies are warranted.

Spatial compartmentalization of free radical formation and mitochondrial heterogeneity in bivalve gills revealed by live-imaging techniques, supplementary material

Live-imaging techniques (LIT) utilize target-specific fluorescent dyes to visualize biochemical processes using confocal and multiphoton scanning microscopy, which are increasingly employed as non-invasive approach to physiological in-vivo and ex-vivo studies. Here we report application of LIT to bivalve gills for ex-vivo analysis of gill physiology and mapping of reactive oxygen (ROS) and nitrogen (RNS) species formation in the living tissue. Our results indicate that H2O2, HOO. and ONOO- radicals (assessed through C-H2DFFDA staining) are mainly formed within the blood sinus of the filaments and are likely to be produced by hemocytes as defense against invading pathogens. The oxidative damage in these areas is controlled by enhanced CAT (catalase) activities recorded within the filaments. The outermost areas of the ciliated epithelial cells composing the filaments, concentrated the highest mitochondrial densities (MTK Deep Red 633 staining) and the most acidic pH values (as observed with ageladine-a). These mitochondria have low (depolarized) membrane potentials (D psi m) (JC-1 staining), suggesting that the high amounts of ATP required for ciliary beating may be in part produced by non-mitochondrial mechanisms, such as the enzymatic activity of an ATP-regenerating kinase. Nitric oxide (NO, DAF-2DA staining) produced in the region of the peripheral mitochondria may have an effect on mitochondrial electron transport and possibly cause the low membrane potential. High DAF-2DA staining was moreover observed in the muscle cells composing the wall of the blood vessels where NO may be involved in regulating blood vessel diameter. On the ventral bend of the gills, subepithelial mucus glands (SMG) contain large mucous vacuoles showing higher fluorescence intensities for O2.- (DHE staining) than the rest of the tissue. Given the antimicrobial properties of superoxide, release of O2.- into the mucus may help to avoid the development of microbial biofilms on the gill surface. However, cells of the ventral bends are paying a price for this antimicrobial protection, since they show significantly higher oxidative damage, according to the antioxidant enzyme activities and the carbonyl levels, than the rest of the gill tissue. This study provides the first evidence that one single epithelial cell may contain mitochondria with significantly different membrane potentials. Furthermore, we provide new insight into ROS and RNS formation in ex-vivo gill tissues which opens new perspectives for unraveling the different ecophysiological roles of ROS and RNS in multifunctional organs such as gills.

Relationship between the oxidation potential and electron spin density of the primary electron donor in reaction centers from Rhodobacter sphaeroides

The primary electron donor in bacterial reaction centers is a dimer of bacteriochlorophyll a molecules, labeled L or M based on their proximity to the symmetry-related protein subunits. The electronic structure of the bacteriochlorophyll dimer was probed by introducing small systematic variations in the bacteriochlorophyll–protein interactions by a series of site-directed mutations that replaced residue Leu M160 with histidine, tyrosine, glutamic acid, glutamine, aspartic acid, asparagine, lysine, and serine. The midpoint potentials for oxidation of the dimer in the mutants showed an almost continuous increase up to ≈60 mV compared with wild type. The spin density distribution of the unpaired electron in the cation radical state of the dimer was determined by electron–nuclear–nuclear triple resonance spectroscopy in solution. The ratio of the spin density on the L side of the dimer to the M side varied from ≈2:1 to ≈5:1 in the mutants compared with ≈2:1 for wild type. The correlation between the midpoint potential and spin density distribution was described using a simple molecular orbital model, in which the major effect of the mutations is assumed to be a change in the energy of the M half of the dimer, providing estimates for the coupling and energy levels of the orbitals in the dimer. These results demonstrate that the midpoint potential can be fine-tuned by electrostatic interactions with amino acids near the dimer and show that the properties of the electronic structure of a donor or acceptor in a protein complex can be directly related to functional properties such as the oxidation–reduction midpoint potential.

beta-Amyloid toxicity in organotypic hippocampal cultures: protection by EUK-8, a synthetic catalytic free radical scavenger.

Oxygen free radicals have been proposed to mediate amyloid peptide (beta-AP)-induced neurotoxicity. To test this hypothesis, we evaluated the effects of EUK-8, a synthetic catalytic superoxide and hydrogen peroxide scavenger, on neuronal injury produced by beta-AP in organotypic hippocampal slice cultures. Cultures of equivalent postnatal day 35 (defined as mature) and 14 (defined as immature) were exposed to various concentrations of beta-AP (1-42 or 1-40) in the absence or presence of 25 microM EUK-8 for up to 72 hours. Neuronal injury was assessed by lactate dehydrogenase release and semiquantitative analysis of propidium iodide uptake at various times after the initiation of beta-AP exposure. Free radical production was inferred from the relative increase in dichlorofluorescein fluorescence, and the degree of lipid peroxidation was determined by assaying thiobarbituric acid-reactive substances. Treatment of mature cultures with beta-AP (50-250 microg/ml) in serum-free conditions resulted in a reproducible pattern of damage, causing a time-dependent increase in neuronal injury accompanied with formation of reactive oxygen species. However, immature cultures were entirely resistant to beta-AP-induced neurotoxicity and also demonstrated no dichlorofluorescein fluorescence or increased lipid peroxidation after beta-AP treatment. Moreover, mature slices exposed to beta-AP in the presence of 25 microM EUK-8 were significantly protected from beta-AP-induced neurotoxicity. EUK-8 also completely blocked beta-AP-induced free radical accumulation and lipid peroxidation. These results not only support a role for oxygen free radicals in beta-AP toxicity but also highlight the therapeutic potential of synthetic radical scavengers in Alzheimer disease.

Novel chromone and xanthone derivatives: Synthesis and ROS/RNS scavenging activities

Chromones and xanthones are oxygen-containing heterocyclic compounds acknowledged by their antioxidant properties. In an effort to develop novel agents with improved activity, a series of compounds belonging to these chemical classes were prepared. Their syntheses involve the condensation of appropriate 2-methyl-4H-chromen-4-ones, obtained via Baker-Venkataraman rearrangement, with (E)-3-(3,4-dimethoxyphenyl)acrylaldehyde to provide the corresponding 2-[(1E,3E)-4-(3,4-dimethoxyphenyl)buta-1,3-dien-1-yl]-4H-chromen-4-ones. Subsequent electrocyclization and oxidation of these compounds led to the synthesis of 1-aryl-9H-xanthen-9-ones. After cleavage of the protecting groups, hydroxylated chromones and xanthones were assessed as scavenging agents against both reactive oxygen species (ROS) [superoxide radical (O2(•-)), hydrogen peroxide (H2O2), hypochlorous acid (HOCl), singlet oxygen ((1)O2), and peroxyl radical (ROO(•))] and reactive nitrogen species (RNS) [nitric oxide ((•)NO) and peroxynitrite anion (ONOO(-))]. Generally, all the tested new hydroxylated chromones and xanthones exhibited scavenger effects dependent on the concentration, with IC50 values found in the micromolar range. Some of them were shown to have improved scavenging activity when compared with previously reported analogues, allowing the inference of preliminary conclusions on the structure-activity relationship.

Radical Language, Radical Identity: Korean Writers in Japanese Spaces and the Burden to "Represent"

Thesis (Ph.D.)--University of Washington, 2016-06

From radical to banal evil: Hannah Arendt against the justification of the unjustifiable

Two central strands in Arendt's thought are the reflection on the evil of Auschwitz and the rethinking in terms of politics of Heidegger's critique of metaphysics. Given Heidegger's taciturnity regarding Auschwitz and Arendt's own taciturnity regarding the philosophical implications of Heidegget's political engagement in 1933, to set out how these strands interrelate is to examine the coherence of Arendt's thought and its potential for a critique of Heidegger. By refusing to countenance a theological conception of the evil of Auschwitz, Arendt consolidates the break with theology that Heidegger attempts through his analysis of the essential finitude of Dasein. In the light of Arendt's account of evil, it is possible to see the theological vestiges in Heidegger's ontology. Heidegger's resumption of the question concerning the categorical interconnections of the ways of Being entails an abandonment of finitude: he accommodates and tacitly justifies that which can have no human justification.

Mass spectrometric analysis of HOCl- and free-radical-induced damage to lipids and proteins

In inflammatory diseases, release of oxidants leads to oxidative damage to biomolecules. HOCl (hypochlorous acid), released by the myeloperoxidase/H2O2/Cl- system, can cause formation of phospholipid chlorohydrins, or alpha-chloro-fatty aldehydes from plasmalogens. It can attack several amino acid residues in proteins, causing post-translational oxidative modifications of proteins, but the formation of 3-chlorotyrosine is one of the most stable markers of HOCl-induced damage. Soft-ionization MS has proved invaluable for detecting the occurrence of oxidative modifications to both phospholipids and proteins, and characterizing the products generated by HOCl-induced attack. For both phospholipids and proteins, the application of advanced mass spectrometric methods such as product or precursor ion scanning and neutral loss analysis can yield information both about the specific nature of the oxidative modification and the biomolecule modified. The ideal is to be able to apply these methods to complex biological or clinical samples, to determine the site-specific modifications of particular cellular components. This is important for understanding disease mechanisms and offers potential for development of novel biomarkers of inflammatory diseases. In the present paper, we review some of the progress that has been made towards this goal.

A spectroscope study of some semiquinone radical ions

The effect of substituents on the value of the oxidation potential of quinones is reviewed and attempts to prepare substituted diphenoquinones with high oxidation potentials are reported. Attempts to characterise the mechanism of addition and substitution in diphenoquinones by identifying the products of the Thiele acetylation of diphenoquinone are reported. The reaction proved most efficient when the incoming acetylinium ion is directed by substituents in the diphenoquinone. A 1,8-addition to diphenoquinone is reported and characterised by isolating the products of the reaction between acetyl chloride and diphenoquinone, with perchloric acid as catalyst. The alternating linewidth effects observed in e.s.r.spectra are discussed and applied to account for such effects observed in the e.s.r.spectra of diphenosemiquinone anion and cation radicals. The spectra are analysed and the intramolecular processes producing these effects are discussed. A dianion diradical where intramolecular rotation about the 1 - 1' bond is restricted is produced by the oxidation of 2,2' ,4,4' -tetra hydroxybiphenyl. Previous studies of diphenosemiquinone anions are reviewed and alkylated diphenosemiquinone anion are produced by the reduction of the parent quinone with potassium hydroxide solution, the resulting radical being stabilised by the presence of pyridine. A qualitative interpretation of the solvent-ion effect in alkylated diphenosemiquinone anions is given. Diphanosemiquinone cation radicals are reviewed and previous studies are re-examined.

Free radicals as potential antitumour agents

The aim of this work was to use extremely low concentrations of free radical generating compounds as a 'catalyst' to trigger endogenous free radical chain reactions in the host and to selectively eliminate neoplastic cells in the host. To test the hypothesis, a number of free radical generating compounds were screened on several malignant cell lines in vitro to select model compounds that were used against tumour models in vivo. 2,2-Diphenyl-1-picrylhydrazyl (DPPH) and its derivatives were selected at the model compounds for in vivo experiments in view of their high cytotoxic potency against several malignant cell lines in vitro. The water soluble derivative, 2,2-diphenyl-1-(2', 4'-dinitro-6'-sulphophenyl) hydrazyl (DDSH) given by subcutaneous injections demonstrated significant antitumour activities against the MAC 16 murine colon adenocarcinoma implanted subcutaneously in male NMRI mice at nanomolar concentration range. 40-60% of long term survival of over 60 days was achieved (compared with control survival of 20 days) with total tumour elimination. This compound was also active against both P388 leukaemia in male BDF1 mice and TLX5 lymphoid tumour in male CBA/CA mice at a similar concentration range. However, some of these animals died suddenly after treatment with no evidence of disease present at post mortem. The cause of death was unknown but thought to be related to the treatment. There was significant increase in serum level of malondialdehyde (MDA) following treatment, but did not correlate to the antitumour activities of these compounds. Induction of supcroxide dismutase (SOD), and glutathione peroxidase (GPx) occurred around day 8 after the administration of DDSH. Histological sections of MAC16 tumours showed areas of extensive massive haemorrhagic necrosis and vascular collapse associated with perivascular cell death following the administration of nanomolar concentration of DDSH which was probably compatible with the effects of free radicals. It was concluded that the antitumour activities of these compounds may be related to free radical and cytokine production.