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Towards the electrospray ionization mass spectrometry ionization efficiency scale of organic compounds.

An approach that allows setting up under predefined ionization conditions a rugged self-consistent quantitative experimental scale of electrospray ionization (ESI) efficiencies of organic compounds is presented. By ESI ionization efficiency (IE) we mean the efficiency of generating gas-phase ions from analyte molecules or ions in the ESI source. The approach is based on measurement of relative ionization efficiency (RIE) of two compounds (B1 and B2) by infusing a solution containing both compounds at known concentrations (C1 and C2) and measuring the mass-spectrometric responses of the protonated forms of the compounds (R1 and R2). The RIE of B1 and B2 is expressed as logRIE(B1, B2) = log[(R1 . C2)/(C1 . R2)]. The relative way of measurement leads to cancellation of many of the factors affecting IE (ESI source design, voltages in the source and ion transport system, solvent composition, flow rates and temperatures of the nebulizing and drying gases). Using this approach an ESI IE scale containing ten compounds (esters and aromatic amines) and spanning over 4 logRIE units has been compiled. The consistency of the scale (the consistency standard deviation of the scale is s = 0.16 logRIE units) was assured by making measurements using different concentration ratios (at least 6-fold concentration ratio range) of the compounds and by making circular validation measurements (the logRIE of any two compounds was checked by measuring both against a third compound).

Do Only Small Uremic Toxins, Chromophores, Contribute to the Online Dialysis Dose Monitoring by UV Absorbance?

The aim of this work was to evaluate the contributions of the main chromophores to the total UV absorbance of the spent dialysate and to assess removal dynamics of these solutes during optical on-line dialysis dose monitoring. High performance chromatography was used to separate and quantify UV-absorbing solutes in the spent dialysate sampled at the start and at the end of dialysis sessions. Chromatograms were monitored at 210, 254 and 280 nm routinely and full absorption spectra were registered between 200 and 400 nm. Nearly 95% of UV absorbance originates from solutes with high removal ratio, such as uric acid. The contributions of different solute groups vary at different wavelengths and there are dynamical changes in contributions during the single dialysis session. However, large standard deviation of the average contribution values within a series of sessions indicates remarkable differences between individual treatments. A noteworthy contribution of Paracetamol and its metabolites to the total UV absorbance was determined at all three wavelengths. Contribution of slowly dialyzed uremic solutes, such as indoxyl sulfate, was negligible.

HPLC study of uremic fluids related to optical dialysis adequacy monitoring

PURPOSE The aim of this study was to investigate uremia-related high-performance liquid chromatography (HPLC) ultraviolet (UV) absorbance profiles of serum and spent dialysate and to study the removal of uremic retention solutes in connection with optical dialysis adequacy monitoring. METHODS 10 uremic patients were investigated using online spectrophotometry at a wavelength of 280 nm over the course of 30 hemodialysis treatments. The dialysate and blood samples were taken and analyzed simultaneously using standard biochemical methods and reversed-phase HPLC. Filters with cutoff at 3 kDa and 70 kDa were used for the pre-treatment of the serum. The chromatographic peaks were detected by a UV detector at wavelengths of 254 and 280 nm. RESULTS This study indicated that the main solute responsible for UV absorbance in the spent dialysate is a low-molecular-weight, water-soluble, non-protein-bound compound uric acid (UA). Three additional uremic retention solutes - creatinine (CR), indoxyl sulphate (IS) and hippuric acid (HA) - were identified from the HPLC profiles. The number of detected HPLC peaks was not significantly different for a serum filtered through the 3 kDa or 70 kDa cutoff filters, and was lower for the spent dialysate, indicating that the molecular weight (MW) of the main UV chromophores in the uremic fluids did not exceed 3 kDa. The reduction ratio (RR) estimated by the total area of HPLC peaks at 254 nm and 280 nm in the serum and by the online UV absorbance at 280 nm was best related to the removal of small water-soluble non-protein bound solutes like urea (UR), CR and UA. CONCLUSIONS The present study contributes new information on the removal of uremic retention solutes during hemodialysis and on the origin of the optical dialysis adequacy monitoring signal.

Optical dialysis adequacy sensor: contribution of chromophores to the ultra violet absorbance in the spent dialysate

Several on-line methods have been developed to standardize the assessment of dialysis adequacy. Earlier studies have demonstrated that on-line monitoring of total ultra violet (UV) absorbance in spent dialysate can be utilized to follow continuously a single hemodialysis session. The aim of this study was to investigate the contribution of different compounds, acting as chromophores, to the UV-absorbance in the spent dialysate in order to explain origin of the cumulative and integrated UV-absorbance measured by the optical dialysis adequacy sensor. Four uremic patients, during 12 hemodialysis treatments, were followed by the optical dialysis adequacy sensor using the wavelength of 280 mn. The dialysate samples were taken and analyzed using reversed phase high performance liquid chromatography (HPLC). The total number of detected peaks from the HPLC gradient separation profiles measured at the wavelength 280 nm for the samples collected 10 mm after the start of hemodialysis (MeanplusmnSD) was 38plusmn6. The relative contribution from the area of 10 main peaks to the total area of all detected peaks in percentage was 91.01plusmn2.52 %. The optical dialysis adequacy sensor provides continuous, on-line hemodialysis measurements and may immediately identify and alert to any deviations in the dialysis. Our study indicates that there exists a number of prevalent compounds that are the main cause of the cumulative and integrated UV- absorbance

Quantification of Indoxyl Sulphate in the Spent Dialysate Using Fluorescence Spectra

The aim of this study was to investigate the possibility to determine the amount of Indoxyl Sulphate (IS) in the spent dialysate using fluorescence spectra.

Concentration of uric acid removed during dialysis. estimated by multi wavelength and processed ultra violet absorbance spectra

The aim of this study was to estimate the concentration of uric acid (UA) optically by using original and processed ultra violet (UV) absorbance spectras of the spent dialysate. Also the effect of using several wavelengths for estimation was examined.

Beta2-microglobulin Measurements in the Spent Dialysate Using Fluorescence Spectra

The aim of this study was to measure the concentration of beta2-microglobulin (B2M) in the spent dialysate using optical method utilizing fluorescence spectra of the spent dialysate.

A multicentre study of an enhanced optical method for measuring concentration of uric acid removed during dialysis

The aim of this study was to compare concentration measurements of uric acid (UA) removed during dialysis by two algorithms based on UV-absorbance and the 1st derivate of UV absorbance. Ten uremic patients from Tallinn and ten from Linköping, during 30+40 haemodialysis treatments, were followed at the Departments of Dialysis and Nephrology at North-Estonian Medical Centre and at Linköping University Hospital. The dialysate samples were taken and analyzed by means of UA concentration at the chemical laboratory and with a double-beam spectrophotometer. UV absorbance and derivate of UV absorbance was transformed into UA concentration in the spent dialysate using the regression models from the calibration set of material, noted as UV-absorbance (UV_A) and the 1st derivate of UV absorbance (UV_D) method. These models were tested on validation set of material and concentrations of UA from the two methods were compared regarding mean values and SD. Mean concentration of UA were 52.7 ± 25.0 micromol/l measured at the chemical laboratory (UA_Lab), 54.9 ± 23.8 micromol/l determined by UV_A and 52.9 ± 23.0 micromol/l determined by UV_D. The results of mean concentrations were not significantly different (p ≥ 0.54). The systematic errors were -7.8 % and -3.3% and random errors were 15.8 % and 10.4 % using UV_A and UV_D respectively. The systematic and random errors were significantly different (p < 0.05) indicating that the new algorithm enables more accurate UA estimation.

Is Fluorescence Valid to Monitor Removal of Protein Bound Uremic Solutes in Dialysis

The aim of this study was to evaluate the contribution and removal dynamics of the main fluorophores during dialysis by analyzing the spent dialysate samples to prove the hypothesis whether the fluorescence of spent dialysate can be utilized for monitoring removal of any of the protein bound uremic solute. A high performance liquid chromatography system was used to separate and quantify fluorophoric solutes in the spent dialysate sampled at the start and the end of 99 dialysis sessions, including 57 hemodialysis and 42 hemodiafiltration treatments. Fluorescence was acquired at excitation 280 nm and emission 360 nm. The main fluorophores found in samples were identified as indole derivatives: tryptophan, indoxyl glucuronide, indoxyl sulfate, 5-hydroxy-indoleacetic acid, indoleacetyl glutamine, and indoleacetic acid. The highest contribution (35 ± 11%) was found to arise from indoxyl sulfate. Strong correlation between contribution values at the start and end of dialysis (R2 = 0.90) indicated to the stable contribution during the course of the dialysis. The reduction ratio of indoxyl sulfate was very close to the decrease of the total fluorescence signal of the spent dialysate (49 ± 14% vs 51 ± 13% respectively, P = 0.30, N = 99) and there was strong correlation between these reduction ratio values (R2 = 0.86). On-line fluorescence measurements were carried out to illustrate the technological possibility for real-time dialysis fluorescence monitoring reflecting the removal of the main fluorophores from blood into spent dialysate. In summary, since a predominant part of the fluorescence signal at excitation 280 nm and emission 360 nm in the spent dialysate originates from protein bound derivatives of indoles, metabolites of tryptophan and indole, the fluorescence signal at this wavelength region has high potential to be utilized for monitoring the removal of slowly dialyzed uremic toxin indoxyl sulfate.

Optical dialysis adequacy sensor: wavelength dependence of the ultra violet absorbance in the spent dialysate to the removed solutes

A need for dialysate-based, on-line, continuous monitoring systems for the control of dialysis efficiency and the prevention of dialysis-associated complications is arisen due to increasing number of dialysis patients and related treatment quality requirements. The aim of this study was to investigate the wavelength dependence between the the ultraviolet (UV) absorbance in the spent dialysate and the retained solutes removed during the hemodialysis in order to explain possibilities to estimate removal of the solutes by the optical dialysis adequacy sensor. Ten uremic patients, during 30 hemodialysis treatments, were followed at the Department of Dialysis and Nephrology, North-Estonian Regional Hospital. The dialysate samples were taken and analyzed with spectrophotometer to get absorbance spectra. The results confirm previous studies considering similarity for the UV-spectrum on the spent dialysate samples during a single dialysis session indicating presence of the same type of chromophores in the spent dialysate removed from the patients blood for different patients groups. At the same time the highest correlation in the spent dialysate for urea, creatinine, potassium, and phosphate was obtained at the wavelength 237 nm that is a new finding compared to earlier results. The highest correlation between the UV-absorbance and uric acid in the spent dialysate was obtained at the wavelength 294 nm. Presence of at least two different wavelength ranges may add selectivity for monitoring several compounds. Our study indicates that the technique has a potential to estimate the removal of retained substances.