Simultaneous Dosage Research Articles

Application of Multiplex Immunoassay in Aging Research: A Methodological Approach.

One of the characteristics of aging and age-related disorders is the formation and evolution of a chronic, low-grade, and hence subclinical, inflammatory state known as inflammaging. Although the progression of inflammaging is now recognized as one of the main driving forces of aging and one of the main risk factors for morbidity and mortality in older subjects, current knowledge on the causative agents of inflammaging itself and chronic, aging-related diseases is still incomplete. In this chapter, we offer a methodological approach for assessing inflammation associated with aging through the use of multiplex immunoassay, which enables the rapid, reproducible, and simultaneous dosage of several cytokines, chemokines, and inflammatory mediators with little biological sample usage.

Glycerol used for denitrification in full-scale wastewater treatment plants: nitrous oxide emissions, sludge acclimatization, and other insights.

Glycerol is commonly employed for denitrification purposes in full-scale wastewater treatment. In non-acclimatized biomass, the glycerol is very inefficient resulting in a high C/N ratio and low-standard denitrification rates. The acclimatization is driven by the microbial enrichment of Saccharimonadales and Propionibacteriales as found in different sampled municipal sludges flanking the dominant presence of Burkholderiales. The selective strategy is based on a very efficient process in terms of C/N ratios and standard denitrification rates, but it leads to nitrite accumulation. As a result, severe and unexpected nitrous oxide emissions were found in full-scale with emission factors up to 2.5% kgN2O (kgKJNremoved)-1. Simultaneous dosage of isobutirate in a full-scale experiment could counter the nitrous oxide emissions. As nitrous oxide emissions were found proportional to the dosed glycerol-based COD, the authors suggest that, in case of acclimatization of biomass to glycerol, an emission factor based on the dosed COD should substitute the general nitrous oxide emission factors based on incoming or removed nitrogen to the plant.

Prescription of the thyroid test results at the biochemistry laboratory in Antananarivo in 2018

According to National Authority for Health, the isolated dosage of TSH, in first-line, is a sufficient supply for the diagnosis and monitoring of thyroid dysfunction. The purpose of this study are to determine the prevalence of prescriptions of thyroid test, evaluate the practices on the prescription of thyroid tests compared to international recommendations. It is a descriptive retropective study within a period of 12 months. All the files with a request for TSH and / or thyroid hormone were included in this study. All files with a previous thyroid check-up or as part of a dysthyroidism follow-up assessment were excluded. Among the 72600 prescriptions for biochemical tests, 184 corresponded to the prescription of thyroid tests, it means 0.25% compared to other biochemical blood tests recorded. Among the 184 prescriptions requesting thyroid tests,117 files were retained. The mean age of the patients was 42.3 years, with a sex ratio of 0.18. One hundred sixteen files included a request of TSH dosage; 28,21% included only a TSH dosage and 70.94% included a request of simultaneous TSH dosage with one of two thyroid hormones. One prescription (0.85%) asked for a thyroid hormones dosage only without preliminary TSH dosage. TSH ranged from <0.05 to 93.97µUI/mL. It was normal in 68.96%, reduced in 16.39% and increased in 14.65% of the dosages. The number of thyroid hormone dosage in first-line in this study is important. Their prescription should be adapted to current recommendations in order to avoid the additional cost of unnecessary dosages for patients.

Genetic interaction screen for severe neurodevelopmental disorders reveals a functional link between Ube3a and Mef2 in Drosophila melanogaster

Neurodevelopmental disorders (NDDs) are clinically and genetically extremely heterogeneous with shared phenotypes often associated with genes from the same networks. Mutations in TCF4, MEF2C, UBE3A, ZEB2 or ATRX cause phenotypically overlapping, syndromic forms of NDDs with severe intellectual disability, epilepsy and microcephaly. To characterize potential functional links between these genes/proteins, we screened for genetic interactions in Drosophila melanogaster. We induced ubiquitous or tissue specific knockdown or overexpression of each single orthologous gene (Da, Mef2, Ube3a, Zfh1, XNP) and in pairwise combinations. Subsequently, we assessed parameters such as lethality, wing and eye morphology, neuromuscular junction morphology, bang sensitivity and climbing behaviour in comparison between single and pairwise dosage manipulations. We found most stringent evidence for genetic interaction between Ube3a and Mef2 as simultaneous dosage manipulation in different tissues including glia, wing and eye resulted in multiple phenotype modifications. We subsequently found evidence for physical interaction between UBE3A and MEF2C also in human cells. Systematic pairwise assessment of the Drosophila orthologues of five genes implicated in clinically overlapping, severe NDDs and subsequent confirmation in a human cell line revealed interactions between UBE3A/Ube3a and MEF2C/Mef2, thus contributing to the characterization of the underlying molecular commonalities.

Development and validation of a liquid chromatography/tandem mass spectrometry method for the simultaneous quantification of serotonin and thromboxane B2 from activated platelets.

Availability of a rapid and reliable platelet activation assay avoiding limitations of current techniques would be valuable to diagnose heparin-induced thrombocytopenia and platelet secretion disorders. The first aim was to develop and validate an ultra-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method to quantify in a single run TxB2 synthesized and serotonin released from platelets. The second aim was to use our method in association with light transmission aggregometry (LTA) to select good platelet responders for the diagnosis of HIT. Electrospray ionization and chromatographic separation were optimized for the simultaneous dosage of serotonin and TxB2. The method was validated according to the European Medicines Agency (EMA) guideline for bioanalytical method validation. LTA was performed with monoclonal anti-CD9 (clone ALB6) as platelet activator to select good responders. Detection was performed using a tandem mass spectrometer with alternated positive and negative electrospray ionization. The total run time was 6minutes. The method was validated for calibration curves, precision, accuracy, lower limit of quantification, carry-over, selectivity, and matrix effect. Platelet response to ALB6 was highly variable among donors. We developed and validated a UHPLC-MS/MS method for the simultaneous quantification of TxB2 and serotonin.

Development of a novel ion-pairing HPLC-FL method for the separation and quantification of hydroxychloroquine and its metabolites in whole blood.

Hydroxychloroquine (HCQ) is an old antimalarial drug that has proven to be a safe and effective treatment for systemic lupus erythematosus (SLE) and other autoimmune diseases. Since hematic concentration of HCQ is closely related to the therapeutic response, monitoring the levels of the drug and its metabolites in the blood of HCQ-treated patients helps the clinician in the evaluation of partial or complete unresponsiveness to treatment. We developed and validated a novel ion-pairing HPLC-FL method for the simultaneous dosage of HCQ, and its major metabolites desethylhydroxychloroquine, desethylchloroquine and bisdesethylchloroquine, after extraction from whole blood. This methodological approach was used for the analysis of real samples obtained from patients affected by SLE and undergoing HCQ treatment. The same samples were also analyzed using a previously validated LC/MS/MS method and data obtained with the two approaches were in substantial agreement with each other. Results presented in this work indicate that this approach can be successfully used to monitor the level of HCQ and its metabolites in the blood of various categories of patients (i.e. low and high responders, or those not adhering to the therapy). Comparison of HPLC-FL and LC/MS/MS data confirmed the efficacy of the proposed method for routine clinical analyses.

The operation of reverse osmosis system with CO2 as a scale inhibitor: A study on operational behavior and membrane morphology

Fouling is one of the acute concerns during the application of reverse osmosis (RO) technology for wastewater reclamation. This study emphasizes the effectiveness of CO2 injection for scale inhibition on the surface of membranes during wastewater reclamation. The main objective of CO2 injection is to reduce pH of feed stream for avoiding inorganic scale precipitation. Two different experimental setups (lab- and pilot-scale) were installed and each was operated with four different RO modules including control (without any scale inhibitor), dosage of antiscalant, CO2 injection and simultaneous dosage of antiscalant and CO2 in an influent line. The pH of influent stream was reduced by CO2 purging at the level of 6 and 5. Data from operational setups, water analysis and morphological study of membrane from each module was used to establish comparative result. In comparison with other modules, the RO modules operated with CO2 were found successful to mitigate the inorganic scale formation on the surface of membrane. Furthermore, the RO modules operated with CO2 presented the successful restoration of system performance with clean-in-place.

Fluorometric Investigation on the Binding of Letrozole and Resveratrol with Serum Albumin.

Breast cancer is the most common cancer in women worldwide. Combination of drugs during long-therm cancer therapy can increase free, biological active form of the drug and cause dangerous side effects. The 21<sup>st</sup> century is a period of searching for a progress in cancer chemotherapy. The simultaneous dosage of drugs and natural agents isolated from fruits and vegetables used in breast cancer treatment could be more effective and less toxic. The aim of the study was to determine the binding sites of both letrozole (LET) and polyphenol product, resveratrol (RES) in tertiary structure of human serum albumin (HSA) based on the fluorescence spectroscopy. The binding of LET and RES to HSA was studied by monitoring the changes in emission fluorescence spectra of albumin in the presence of ligands at 280 nm and 295 nm excitation wavelengths. To identify the binding sites of LET and RES on HSA, warfarin (WAR) and 5-(Dimethylamino)-1-naphthalenesulfonamide (DNSA) were used as site probes for binding site I, while dansyl-L-proline (DP) was studied as a marker for binding site II. The binding sites for LET and RES in HSA were found to be located in subdomain IIIA. Based on the Stern - Volmer and binding isotherm using non-linear regression methods, the formation of complexes of LET and RES with HSA and association constants were obtained. The binding analysis showed that the association constants indicated a stronger interaction of HSA with RES than LET and the presence of RES in the tertiary system alters the stability of LET-albumin complex. This conclusion points to the necessity of precaution and monitoring therapy when resveratrol as a natural compound and letrozole are used together.

Abstract 4832: Independent prognostic role of chromosome 12p deletion in prostate cancer

Abstract Deletion of chromosome 12p is a recurrent alteration in prostate cancer, but the prevalence and clinical significance of this alteration is unclear. More than 7,000 prostate cancers with clinical follow-up data in a tissue microarray (TMA) format were analyzed by dual labeling fluorescence in situ hybridization (FISH) using probes for 12p13 (CDKN1B; p27) and for centromere 12 as a reference. CDKN1B was selected as a probe because this gene is located in the center of the deletion, which spans >10 Mb and includes >50 genes in 80% of cancers with 12p deletion. In 13.7% of prostate cancers a deletion of 12p was found (13.5% hemizygeous; 0.2% homozygeous deletions). 12p deletions were linked to advanced tumor stage (p<0.0001), high Gleason grade (p<0.0001), lymph node metastasis (p = 0.0004), rapid tumor cell proliferation (p<0.0001), and early biochemical recurrence (BCR) (p = 0.0027). Multivariate analysis including pT stage (p<0.0001), Gleason grade (p<0.0001), pN status (p = 0.0001), preoperative PSA levels (p = 0.0001), and resection margin status (p = 0.0001) revealed an independent prognostic value of 12p deletion (p = 0.0014). There was no association with ERG fusion status. Deletion of 12p deletion was only minimally linked to p27 expression status, which by itself was unrelated to clinical outcome. This argues against p27 as the key target gene of 12p deletions. Since none of the involved 12p genes is recurrently mutated in next generation sequencing studies, simultaneous dosage reduction of multiple genes may exert a relevant biological effect in 12p deleted cancers. In summary, the results of our study demonstrate that 12p deletion is frequent and provides independent prognostic information in prostate cancer. Citation Format: Sarah Jane Pauline Minner, Ramin Ahray, Martina Kluth, Thorsten Schlomm, Ronald Simon, Guido Sauter, Maria Christina Tsourlakis. Independent prognostic role of chromosome 12p deletion in prostate cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4832. doi:10.1158/1538-7445.AM2015-4832

MP55-08 GENOMIC DELETION OF CHROMOSOME 12P IS AN INDEPENDENT PROGNOSTIC MARKER IN PROSTATE CANCER

You have accessJournal of UrologyProstate Cancer: Basic Research III1 Apr 2015MP55-08 GENOMIC DELETION OF CHROMOSOME 12P IS AN INDEPENDENT PROGNOSTIC MARKER IN PROSTATE CANCER Raisa Pompe, Martina Kluth, Sarah Minner, Philipp Gild, Ronald Simon, Pierre Tennstedt, Markus Graefen, Guido Sauter, and Thorsten Schlomm Raisa PompeRaisa Pompe More articles by this author , Martina KluthMartina Kluth More articles by this author , Sarah MinnerSarah Minner More articles by this author , Philipp GildPhilipp Gild More articles by this author , Ronald SimonRonald Simon More articles by this author , Pierre TennstedtPierre Tennstedt More articles by this author , Markus GraefenMarkus Graefen More articles by this author , Guido SauterGuido Sauter More articles by this author , and Thorsten SchlommThorsten Schlomm More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2015.02.2051AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Deletion of 12p is a recurrent alteration in prostate cancer, but the prevalence and clinical consequences of this alteration have not been studied in detail. METHODS Dual labeling fluorescence in situ hybridization (FISH) using probes for 12p13 (CDKN1B; p27) and for centromere 12 as a reference was used to analyze more than 7,000 prostate cancers with clinical follow-up data assembled in a tissue microarray (TMA) format. CDKN1B was selected as a probe because it is located in the center of the deletion, which, however, spans >10 Mb and includes >50 genes in 80% of cancers with 12p deletion. Results were compared to clinically follow-up data, ERG status and p27 protein expression in univariate und multivariate analyses. RESULTS Deletion of 12p was found in 13.7% of cancers and included 13.5% hemizygeous and 0.2% homozygeous deletions. 12p deletion were linked to advanced tumor stage (p<0.0001), high Gleason grade (p<0.0001), lymph node metastasis (p=0.0004), rapid tumor cell proliferation (p<0.0001), and early biochemical recurrence (BCR) (p=0.0027). Multivariate cox regression analysis including pT stage (p<0.0001), Gleason grade (p<0.0001), pN status (p=0.0001), preoperative PSA levels (p=0.0001), and resection margin status (p=0.0001) revealed an independent prognostic value of 12p deletion (p=0.0014). Deletion of 12p was unrelated to the ERG fusion status. Deletion of 12p was only marginally linked to reduced p27 expression, which by itself was unrelated to clinical outcome. This argues against p27 as the key target gene of 12p deletions. As none of the involved 12p genes is recurrently mutated in next generation sequencing studies, simultaneous dosage reduction of multiple genes may exert a relevant biological effect in 12p deleted cancers. CONCLUSIONS The results of our study demonstrate that 12p deletion is frequent in prostate cancer and provides independent prognostic information. 12p deletion analysis alone, or in combination with other prognostic parameters may thus have clinical utility. © 2015 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 193Issue 4SApril 2015Page: e675-e676 Advertisement Copyright & Permissions© 2015 by American Urological Association Education and Research, Inc.MetricsAuthor Information Raisa Pompe More articles by this author Martina Kluth More articles by this author Sarah Minner More articles by this author Philipp Gild More articles by this author Ronald Simon More articles by this author Pierre Tennstedt More articles by this author Markus Graefen More articles by this author Guido Sauter More articles by this author Thorsten Schlomm More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Synergistic interactions between Drosophila orthologues of genes spanned by de novo human CNVs support multiple-hit models of autism.

Autism spectrum disorders (ASDs) are highly heritable and characterised by deficits in social interaction and communication, as well as restricted and repetitive behaviours. Although a number of highly penetrant ASD gene variants have been identified, there is growing evidence to support a causal role for combinatorial effects arising from the contributions of multiple loci. By examining synaptic and circadian neurological phenotypes resulting from the dosage variants of unique human:fly orthologues in Drosophila, we observe numerous synergistic interactions between pairs of informatically-identified candidate genes whose orthologues are jointly affected by large de novo copy number variants (CNVs). These CNVs were found in the genomes of individuals with autism, including a patient carrying a 22q11.2 deletion. We first demonstrate that dosage alterations of the unique Drosophila orthologues of candidate genes from de novo CNVs that harbour only a single candidate gene display neurological defects similar to those previously reported in Drosophila models of ASD-associated variants. We then considered pairwise dosage changes within the set of orthologues of candidate genes that were affected by the same single human de novo CNV. For three of four CNVs with complete orthologous relationships, we observed significant synergistic effects following the simultaneous dosage change of gene pairs drawn from a single CNV. The phenotypic variation observed at the Drosophila synapse that results from these interacting genetic variants supports a concordant phenotypic outcome across all interacting gene pairs following the direction of human gene copy number change. We observe both specificity and transitivity between interactors, both within and between CNV candidate gene sets, supporting shared and distinct genetic aetiologies. We then show that different interactions affect divergent synaptic processes, demonstrating distinct molecular aetiologies. Our study illustrates mechanisms through which synergistic effects resulting from large structural variation can contribute to human disease.

Ascorbic Acid may Exacerbate Aspirin-Induced Increase in Intestinal Permeability.

Ascorbic acid in combination with aspirin has been used to prevent aspirin-induced oxidative GI damage. We aimed to determine whether ascorbic acid reduces or prevents aspirin-induced changes in intestinal permeability over a 6-hr period using saccharidic probes mannitol and lactulose. The effects of administration of 600mg aspirin alone, 500mg ascorbic acid alone and simultaneous dosage of both agents were compared in a cross-over study in 28 healthy female volunteers. These effects were also compared with that of a placebo. The ability of ascorbic acid to mitigate the effects of aspirin when administered either half an hour before or after dosage with aspirin was also assessed in 19 healthy female volunteers. The excretion of lactulose over the 6-hr period was augmented after consumption of either aspirin or ascorbic acid compared with that after consumption of placebo. Dosage with ascorbic acid alone augmented the excretion of lactulose more than did aspirin alone. Simultaneous dosage with both agents augmented the excretion of lactulose in an additive manner. The timing of dosage with ascorbic acid in relation to that with aspirin had no significant effect on the excretion of the two sugars. These findings indicate that ascorbic acid does not prevent aspirin-induced increase in gut permeability rather that both agents augment it to a similar extent. The additive effect on simultaneous dosage with both agents in augmenting the absorption of lactulose suggests that each influences paracellular permeability by different pathways.

Removal of phosphorus and residual aluminium with the simultaneous use of chitosan and alum on the effluent of an MBBR biological system during start-up

Physico–chemical treatment with aluminium salts is a common practice to remove total phosphorus (TP) from wastewaters. However, the use of alum can increase the residual aluminium concentration both in the effluent and biosolids. Chitosan, an alternative coagulant, does not allow for the removal of TP below the requirement level when lower than the soluble phosphorus fraction of the water. Hence, simultaneous dosage of alum and chitosan solutions was evaluated on the effluent of a newly installed MBBR (moving bed biofilm reactor) system for residual TP and aluminium removal. At alum optimal dosage, the most effective chitosan solution generated: 1) an optimal dosage zone for which residual TP and aluminium concentrations were minimal; 2) maximal abatements for TP and aluminium that reached 35 and 85%, respectively, the concentrations observed with alum only. Also, the fraction of residual aluminium in the biosolids was increased, particularly from 84 to 98% at optimal chitosan dosage.

Parallelism and paradoxes on viability and the life span of two loss-of-function mutations: Heat shock protein transcriptional regulator hsf 1 and l(2)gl tumor suppressor in Drosophila melanogaster

In this study we analyzed how a dosage decrease in mono- and diheterozygotes on both lethal alleles of the lgl-gene and hsfheat shock regulator influences viability and life span at optimal and high temperature 29 degrees C conditions. We found that hsf(1) (1 dosage of active hsf-factor) heterozygote animals had significantly increased viability (up to 30-39%) in case of its development from egg to imago under the stress of 29 degrees C. However, this stress-protective effect of a decreased dosage of HSF1 was suppressed in diheterozygotes, while the dosage of tumor suppressor lgl was simultaneously decreased. Under stress temperature conditions, a decrease in dosage of one of the alleles also increased the average life span and delayed aging, especially in the case of maternal inheritance of each of the loss-of-function mutations. In diheterozygotes the average life span had intermediate meanings. However, in diheterozygote males under stress conditions the positive longevity effect of hsf was suppressed in the presence of the lgl-mutation. Paradoxically, that decrease of expression of each of the studied vital genes provided a positive effect on both viability and life span under stress conditions. However, a simultaneous dosage decrease of two loss-of-function alleles in diheterozygotes resulted in disbalanced effects on the organism level. The received data indicate interaction between HSF1 and LGL gene products during ontogenesis and stress-defending processes.

Optimization of intermittent, simultaneous dosage of nitrite and hydrochloric acid to control sulfide and methane productions in sewers

Free nitrous acid (FNA) was previously demonstrated to be biocidal to anaerobic sewer biofilms. The intermittent dosing of FNA as a measure for controlling sulfide and methane productions in sewers is investigated. The impact of three key operational parameters namely the dosing concentration, dosing duration and dosing interval on the suppression and subsequent recovery of sulfide and methane production was examined experimentally using lab-scale sewer reactors. FNA as low as 0.26 mg-N/L was able to suppress sulfide production after an exposure of 12 h. In comparison, 0.09 mg-N/L of FNA with 6-h exposure was adequate to restrain methanogenesis effectively. The recovery of sulfide production was well described by an exponential recovery equation. Model-based analysis revealed that 12-h dosage at an FNA concentration of 0.26 mg-N/L every 5 days can reduce the average sulfide production by >80%. Economic analysis showed that intermittent FNA dosage is potentially a cost-effective strategy for sulfide and methane control in sewers.

Full‐scale sludge liquor treatment for ammonium reduction with low carbon dosage

The separate treatment of sludge liquor, produced by dewatering anaerobic digested sludge at wastewater treatment plants, gives rise to extreme environments for nitrogen removal. A full‐scale sequencing batch reactor was operated with the aim of introducing and studying denitritation as a supplement to nitritation in order to reduce operating costs. Since the main plant only has problems with ammonium reduction capacity, the initial strategy was to have sufficient ammonium reduction with optimal alkalinity production by denitrifiers, i.e. low carbon dosage and minimum alkalinity and residual oxidized ammonium in the effluent. This strategy led to an unbalanced and sensitive process because the denitrifiers were often inhibited. High dissolved oxygen (DO) readings and no decrease in oxidation–reduction potential (ORP) during anoxic phases with simultaneous ethanol dosage indicated inhibition of denitrifiers, probably by the intermediate product NO, which may have interfered with the DO sensor. Nitric oxide production was believed to be favoured in the beginning of the anoxic phase as a result of low pH and high nitrite concentration. A stable nitritation–denitritation process could be achieved when the aerobic hydraulic retention time (HRT) was decreased to the same length as the anoxic HRT, which resulted in increased unused alkalinity.

Quinine impairs quinidine clearance in rat perfused liver.

We have examined the disposition of the cinchona alkaloids quinine and quinidine in the rat recirculating isolated perfused liver preparation. When administered as separate 1 mg doses, the hepatic clearances of quinine and quinidine were similar to the hepatic perfusate rate of 10 mL min-1. When 1 mg of each was administered simultaneously, mean hepatic clearance of quinine was unchanged (9.00 +/- 2.20 mL min-1 separate dosage, n = 7; 6.87 +/- 1.77 mL min-1 simultaneous dosage, n = 7; P > 0.05). By contrast, mean hepatic clearance of quinidine was reduced significantly by concomitant quinine (10.6 +/- 1.72 mL min-1 separate dosage, n = 7; 4.82 +/- 1.25 mL min-1 simultaneous dosage, n = 7; P < 0.05). There was no significant difference in volumes of distribution when each alkaloid was administered separately (131 +/- 46 mL quinine, 129 +/- 21 mL quinidine; P > 0.05) but concomitant quinine administration increased quinidine volume of distribution to 169 +/- 30 mL (P < 0.05). Four further experiments with simultaneous dosages of 0.5 mg of each alkaloid produced similar findings, indicating that the interactions did not derive from nonlinear drug disposition.

Abstract B47: Genomic screen of colorectal tumors and adenomas: Disproportionate frequency of 1p35-1pter deletion in synchronous advanced adenomas

Abstract B47 The development of somatic colorectal cancer (CRC) has been characterized by the sequential acquisition of non-random gene mutations that over time lead to widespread genomic instability. Long-oligonucleotide microarrays for comparative genomic hybridization (oligo-aCGH) provide simultaneous gene dosage measures at high genomic resolution (average resolution 6 kb). Utilizing specialized protocols we routinely co-hybridize FFPE sample DNA with sex-mismatched genomic reference DNA to generate sensitive gene dosage measures at the individual gene level. Standard analyses by built-in bioinformatics allow uniform data analysis at the gene, chromosome and genome levels. Stringent quality control measures provide strong confidence of our experimental techniques in profiling &amp;gt;100 colorectal archival samples stored in the paraffin block 4 to 11 years. FFPE samples included non-advanced adenomas to 3 mm diameter (n = 21), advanced adenomas (n = 33), invasive CRCs (n = 42) and normal margin (n = 10). We have previously reported on the most frequent recurrent alteration of low level gain of the HOXA genes at 7p15 in all 40 - 65% of adenoma subtypes as well as invasive CRC. Subsequent functional studies have shown crucial biologic significance of HOXA9 based on cell line and shRNAi experiments. Here we report on the disproportionate frequency of 1p terminal deletions in 6 of 13(45%) of the synchronous advanced adenoma (SAA) subset. In the advanced adenoma (AA) subset, the 1p35-1pter deletion was found exclusively in the SAA. The same 1p deletion was detected in 2 / 42 (6%) of the invasive CRCs and was not detected in non-advanced adenomas. Historically, the 1p region is commonly altered in solid tumors, including CRCs, and reports link the loss of terminal 1p to development of anueploidy and increased genomic instability. For example, SAA patient #1 presented with multiple advanced adenomas but without known family history CRC. Analysis of 3 separate SAAs showed a common 1p terminal deletions with hemizygous loss starting at position 29.92 MB to 35.4 MB. These SAA showed variable losses or gains of whole chromosomes (gain of 7, loss of 10q, loss of 18, and gain of 20q) to suggest increased genomic instability. The 1p deletion, however, was the only large scale alteration in common. Numerous past studies have examined allelic imbalances at 1p loci in colorectal adenomas and CRC; however, isolating a putative tumor suppressor remains elusive. Our strategy includes further analyses of the enriched subset of SAA with more detailed analysis of the high resolution (6 kb average probe coverage) oligo-aCGH data to detect homozygous loss within the 1p region of hemizygous loss. Candidate genes identified in our screens partially confirm those previously described in the literature and include p73, MAD2L2, AK2. In addition, we will describe several novel tumor suppressors identified from the commonly deleted 1p region. Citation Information: Cancer Prev Res 2008;1(7 Suppl):B47.

Tissue Depletion of Amoxicillin and Its Major Metabolites in Pigs: Influence of the Administration Route and the Simultaneous Dosage of Clavulanic Acid

A residue depletion study of amoxicillin (AMO) and its major metabolites, amoxicilloic acid (AMA) and amoxicillin diketopiperazine-2',5'-dione, was performed after a single oral (p.o.) and intravenous (i.v.) administration of amoxicillin (20 mg kg (-1)) and amoxicillin/clavulanic acid (20 and 5 mg kg (-1)) to pigs. Animals were slaughtered 12, 36, 48, 60, 72, and 84 h after dosing. Tissue samples were analyzed using liquid chromatography-tandem mass spectrometry. Kidney samples contained high concentrations of amoxicilloic acid metabolite, which depleted much slower from tissues than amoxicillin, both after p.o. (t1/2AMO = 4.5 h vs t1/2AMA = 8 h) and i.v. (t1/2AMO = 4 h vs t1/2AMA = 8 h) administration. Moreover, after oral administration, significantly higher amoxicilloic acid concentrations were measured in liver and kidney than after i.v. administration. The coadministration of amoxicillin with clavulanic acid provoked no significant differences in amoxicilloic acid tissue concentrations as compared to an amoxicillin dosing. The prolonged presence of residues of amoxicilloic acid in edible tissues can play an important role in food safety, because the compound could give rise to a possible health risk, although it is not included in the maximum residue limit legislation.

Polyoxyethylene-Δ 9,11-didehydrostearate and glycerol-polyoxyethylene-Δ 9,11-didehydrostearate: two new components of the non-ionic emulsifier Cremophor® EL

The polyethoxylated heterogeneous components of the so far poorly characterised non-ionic emulsifier Cremophor® EL (polyoxyl 35 castor oil) (CrEL) were fractionated by cyclodextrin modified micellar electrokinetic capillary chromatography (CD-MEKC) combined with indirect UV detection. The resulting peaks were assigned to the corresponding components by delayed extraction matrix-assisted laser desorption/ionization time of flight mass spectrometry (DE–MALDI–TOF–MS) as detection device. In order to combine CE and MS the fractionating robot Probot was employed which enables both the online fractionation of the CE eluate on a MALDI target during the electrophoretic separation and the simultaneous dosage of the MALDI matrix. The obtained mass spectra were evaluated by comparing the residue masses of the homologue peak series of the polyethoxylates with the calculated residue masses of potential CrEL-components. The overlapping of homologue peak series with isobaric residue masses was detected by using the residue mass plot, the newly developed evaluation method. Combining theses techniques, both the first detailed structure elucidation and a semiquantitative analysis of the polyethoxylated CrEL-components was achieved. Together with the polyethoxylate series of yet elucidated structures two additional series were observed the corresponding components of which could not be identified at the beginning. Systematic investigations showed that the elimination of water from ricinoleic acid during the synthesis of the emulsifier leads to the polyethoxylates glycerol POE-Δ 9,11-didehydrostearate and POE-Δ 9,11-didehydrostearate so far unknown in CrEL.