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The acid dissociation constant (pKa) is among the most frequently used physicochemical parameters, and its determination is of interest to a wide range of research fields. We present a brief introduction on the conceptual development of pKa as a physical parameter and its relationship to the concept of the pH of a solution. This is followed by a general summary of the historical development and current state of the techniques of pKa determination and an attempt to develop insight into future developments. Fourteen methods of determining the acid dissociation constant are placed in context and are critically evaluated to make a fair comparison and to determine their applications in modern chemistry. Additionally, we have studied these techniques in light of present trends in science and technology and attempt to determine how these trends might affect future developments in the field.

A fluorosurfactant, the anionic N-ethyl-N-[(heptadecafluorooctyl)sulfonyl]glycine potassium salt, trade name FC-129 [CAS 2991-51-7] was investigated for possible application in micellar electrokinetic capillary chromatography (MEKC). The surfactant was characterized with conductometric titration and test sample mixtures were investigated in MEKC systems, and compared with sodium dodecylsulphate. An increased efficiency and interesting selectivity differences were observed.

Background: Low-grade ischemia-reperfusion in claudicants leads to damage of local tissues and remote organs. Since this damage is partly caused by oxygen-derived free radicals (ODFR), scavenging these ODFR could reduce the local and remote injury., Methods: Using a new method by which a free radical reaction product (ortho-APOH) of the exogenous marker antipyrine is measured to quantify the oxidative stress, 16 stable claudicants performed a standard walking test before and after administration of vitamin E (200 mg) and vitamin C (500 mg) daily for 4 weeks., Findings: Ortho-APOH was significantly increased during the reperfusion period (P = 0.026) before administration of the vitamins. After 4 weeks of vitamin supplementation no rise was found in the reperfusion period. Malondialdehyde showed no changes in either group., Interpretation: These findings indicate that administering extra antioxidants to claudicants reduces oxidative stress in these patients. This may also have an effect on the remote ischemia-reperfusion damage and reduce cardiovascular morbidity in this group., (Copyright 2001 Academic Press.)

The performance of a biotrickling filter for treatment of concentrated waste gases was investigated. The macrokinetics of methylmethacrylate degradation in the biotrickling filter is studied by measuring the degradation product methacrylic acid in the drain of the filter. The drain was analysed using isotachophoresis (ITP) and capillary zone electrophoresis (CZE). The CZE analyses were carried out in an I.D. 75 microm capillary at 20 kV (negative inlet polarity) using a 0.01 M Tris-acetate buffer of pH 4.45. The electroosmotic flow (EOF) was suppressed by addition of CTA and PVA to the buffer. Detection was at 214 nm. After filtration through a 0.45-microm filter, samples were directly injected. The calibration graph was linear between 10 and 800 mg/l methacrylic acid, with an analysis time under 2 min.

Discontinuities of pH at zone boundaries of strong ion sample zones in isotachophoretic systems with poorly buffering leading electrolytes have been discovered by dynamic computer simulations of isotachophoretic separations with uncommon electrolyte systems. When using a salt solution containing a strong cation (anion) as leading ion and a weak anion (cation) as counterion for the isotachophoretic separation of strong cations (anions), severe pH discontinuities are present at zone boundaries of these cations (anions). The magnitude of these pH discontinuities has been investigated as a function of several parameters, namely counter ion/leading ion concentration ratio of the leading electrolyte, pK values, and mobilities of sample ions and counterion.

Chiral interaction in capillary electrophoresis can be modeled using pK values, mobilities of analytes, and their formation constants with the chiral selector. An existing steady-state simulation program for CE (HPCESIM) was recently extended with a chiral submenu involving the chiral parameters listed above. These were experimentally determined in both our laboratories for mandelic acid and terbutaline using hydroxypropylated beta-cyclodextrin as chiral selector. A comparison was made between both sets of parameters and between experimental electropherograms and those obtained from simulation. Error analysis of the results indicate the sensitivity of the obtained results.

Solutions of histamine and methacholine bromide in different matrices for diagnostic purposes were analyzed for stability and quality control using capillary electrophoresis. Histamine (2,[4-imidazolyl]ethylamine) [CAS No. 51-45-6] was determined using a 0.1 M Tris-borate buffer of pH 8.3 with 5.10(-5) M cetyltrimethylammonium bromide (CTAB) and 0.005% poly(vinyl alcohol) (PVA) and detected at 214 nm using clenbuterol [4-amino-alpha-(tert.-butylaminomethyl)-3,5-dichlorobenzyl alcohol] [37148-27-9] as an internal standard. Metacholine bromide (acetyl-beta-methacholine bromide) [333-31-3] was determined with a 0.01 M creatinine-chloride buffer of pH 4.85 and detected with indirect UV at 230 nm using potassium as an internal standard. Histamine solutions were stable for a prolonged period of time, whereas under enforced degradation conditions methacholine was hydrolyzed, yielding acetic acid and (tentatively) beta-methylcholine as reaction products.

Computer programs, simulating electrophoretic separations, were evaluated and discussed with respect to their suitability for training purposes. Quite a number of them, mainly those dealing with steady-state phenomena, are sufficiently fast and user-friendly for the purpose of visualization of electrophoretic principles. Transient-state or dynamic models, however, are more suitable for the advanced user, mainly because of their inherent complexity and long calculation times.

The suitability of isotachophoresis for the analysis of metals in, e.g., environmental samples was studied. In a cationic operational system the heavy metals Fe, Cu, Ni, Cd, Co, Zn, Pb and Mn were simultaneously determined. The separation was achieved through complex formation with one of the counter ions, hydroxyisobutyric acid. The other counter ion was acetic acid, the leading ion was 0.02 M potassium or sodium (pH 4.1) and the terminator was H+. The analysis time was 15 min at 60 microA in a 0.2 mm I.D. capillary. Aqueous samples containing ppm and ppb amounts were enriched on a cation exchanger with an extremely low affinity for sodium (Chelex 100). Good recovery, linearity, precision and accuracy were obtained even down to the ppb range. Although the sensitivity of the method is not greater than that of some of the more established methods for the individual metals, a great advantage of isotachophoresis is the simultaneous determination of the metals, with equal response factors. An example is given of the determination of metals, including aluminium, in serum.

This study investigated possible differences between hypertensive men and hypertensive women concerning the hemodynamic effects of incremental doses of exogenous epinephrine. The study population comprised 38 men (37 +/- 10 years) (standard deviation) and 25 women (33 +/- 9 years) with mild essential hypertension (mean blood pressure 147/90 and 147/93 mm Hg, respectively). Body mass index was slightly higher in men (25 +/- 3 kg/m2) than in women (23 +/- 2 kg/m2). Both groups received an intravenous infusion with epinephrine of 15 and 30 ng/kg/min for 8 minutes each. Despite the similar doses of epinephrine infused in both groups, the increase of venous plasma epinephrine in men was nearly twice that in women (1.04 +/- 0.09 vs 0.67 +/- 0.09 nmol/liter, p less than 0.01), suggesting that women cleared the infused epinephrine more efficiently than men. At the highest infusion dose, the increase of systolic blood pressure was larger in men than in women (5.3 +/- 1.2 vs 1.7 +/- 1.1 mm Hg, p less than 0.05). Conversely, the decrease of diastolic blood pressure was also larger in men than in women (-8.8 +/- 1.0 vs -5.8 +/- 1.0 mm Hg, p less than 0.05). The heart rate increased to the same extent in both groups (11.5 +/- 0.8 and 13.7 +/- 1.2 beats/min). If the blood pressure responses were corrected for the increase of plasma epinephrine, the difference between men and women disappeared.(ABSTRACT TRUNCATED AT 250 WORDS)

A method for the determination of conjugated bile acids in human bile using isotachophoresis in 95% methanol is described. The leading ion is 0.01 M chloride, the counter ion is hydroxylamine at its pK value and the terminating ion is N-2-hydroxyethylpiperazine-N'-2-ethanesulphonic acid (HEPES). The sample preparation consists of C18-silica cartridge adsorption. Microlitre amounts of the methanol eluate are injected and analysed within 20 min in a 0.2 mm I.D. PTFE capillary. The sensitivity of the method is better than 50 ng of each of the conjugated bile acids using a.c. conductivity detection.

One aspect of the isotachophoretic determination of protein patterns in biological samples of interest is the characterization of allergens. This group of (glyco) proteins, causing allergic reactions, is used both for diagnosis and in the treatment of allergy. The aim of this investigation was to obtain a maximum amount of information, within one run, on the (glyco)protein composition of a number of allergenic extracts (e.g., from pollen or house dust mites). Commercially available extracts were dialysed prior to analysis to remove disturbing buffer constituents. A high-pH system was chosen in order to obtain a maximum amount of information from the samples (1-2 microliter). The leading electrolyte was 0.01 M C1-, buffered with Tris (pH 8.2), containing 0.2% w/v hydroxyethylcellulose, and the terminating electrolyte was beta-alanine, buffered to pH 10 with Ba(OH)2. The total analysis time was 15-20 min using a PTFE capillary (0.2 mm I.D.). The pre-separation current was 30 microA and the current during detection was 15 microA. UV absorption was measured at 280 nm. For optimal discrimination of the compounds of interest, an ampholyte mixture was used for spacing. The analytical procedure yielded highly reproducible UV patterns. Significant differences between various allergenic extracts were observed. It was concluded that isotachophoresis is a powerful method for the physico-chemical characterization of individual allergenic extracts, e.g., with respect to manufacturing and quality control.

Isotachophoresis seems a likely candidate for a reference method, as it is more accurate and precise than common routine methods. The response is highly linear and depends on a well defined transport number of the leading ion, stability of the driving current, mobility of the separand, which is well controlled by the leading electrolyte and the use of a high-resolution detector. The suitability of isotachophoresis as a reference method was investigated for the determination of sodium in human serum. The operational conditions were 0.01 M K+/citrate (leading electrolyte) at pH 5.5 and 0.01 M creatinine X HCl (terminating electrolyte). Both n-butylamine and ammediol could be used as internal standards. The calibration graph constructed from standard solutions, diluted by weight with the internal standard, yielded a correlation coefficient of 0.99994 (n = 50) in the working range. The method seems especially useful for the determination of any ionic solute in, e.g., clinical samples (lithium, calcium, creatinine or drugs).

Free theophylline was isolated from human serum by ultrafiltration and analysed in a leading electrolyte of 7.5 mM morpholinoethanesulphonic acid with ammediol as a counter ion at pH 8.90 and alpha-alanine as a terminator. The UV (280 nm) absorbance of the theophylline spike between serine and bicine as spacers was integrated. Binding percentages to human pool serum, human albumin and alpha 1-acid glycoprotein (orosomucoid) were determined at physiological concentrations, and found to be 55, 44 and 12%, respectively. The calibration lines were straight from 0 to 30 mg/l, with a standard deviation of 0.2 mg/l. The detection limit was 1 mg/l. The time of analysis was 12 min at 40 microA in a 0.2 mm I.D. capillary.

Twenty-six drugs in solutions for intravenous injection were determined by capillary isotachophoresis with only one calibration point for each component. The maximum deviation of the labelled concentration was 2%. A new calibration constant is introduced, viz., the response factor (RF, dimensionless), which is independent of the diameter of the capillary, the construction of the universal detector and the driving current used during detection. It is shown that the RF can be used on different equipment, using different currents during detection. It appears that the RF is usable for routine analysis when a deviation of 5% is acceptable. Daily one-point recalibration, however, improves this value to 1%.

The spectrum of low molecular weight compounds, in particular of ribonucleotides, within first cleavage stage embryos of the polar lobe-forming mollusc Nassarius reticulatus and the distribution of the compounds within the embryo at the trefoil stage of first cleavage are analysed by means of capillary isotachophoresis after 0.5 M PCA extraction. The compounds which are found in the whole trefoil embryo (T), the lobeless part (LL), and the polar lobe (PL) respectively, and the mean quantities (nmol. microliter-1; n = 6) are: UTP (11.5, 4.8, 5.6), ITP (8.5, 3.6, 5.0), GTP (10.3, 3.0, 9.0), ATP (29.8, 13.4, 18.8), UDP (11.8, 3.4, 8.7), CTP (8.0, 3.1, 4.5), GDP (5.3, 2.6, 3.4), ADP (16.5, 6.1, 11.6), CDP (4.0, 1.4, 2.6), GMP (4.7, 2.7, 4.3), glucose-6-phosphate (G6P) (53.5, 38.8, 13.0). These compounds appear to be localized in the non-yolk cytoplasmic pool. As the volume ratio of PL/LL for total volume and for non-yolk cytoplasmic volume is about 0.74 and 0.60 respectively, the concentration of all nucleotides in PL as compared to LL is significantly higher (HO, p less than 0.001), both relative to the total volume and to the non-yolk cytoplasmic volume. The G6P concentration is considerably higher in the lobeless part. The morphogenetic role of the vegetal pole compartment of the egg apparently is correlated with a relatively high level of its nucleotide contents.

The suitability of isotachophoresis for the determination of quinine in different samples was investigated. The operational conditions were 0.01 M potassium-morpholinoethanesulphonic acid (MES) (pH 6.0) with 0.05% Mowiol as the leading electrolyte and ca. 0.005 M creatinine-MES as the terminating electrolyte. The analyses were carried out at 25 microA in a 0.2 mm I.D. PTFE capillary with UV and conductivity detection. Quinine-containing beverages were degassed by sonification and directly injected. The limit of detection was 5 mg/l with a 4 microliter injection volume. The allowed concentrations could be determined with sufficient accuracy. Analgesic preparations were dissolved in a solution of 5 X 10(-3) M MES with sonification. The quinine levels found agreed well with the declared values. The other constituents of the pharmaceuticals did not interfere with the analysis. Urine samples from volunteers were analysed after consumption of tonic. The samples were extracted with dichloromethane-isopropanol (95:5), vortexed, centrifuged, evaporated to dryness, the residue dissolved in 5 X 10(-3) M MES and analysed. At a concentration factor of 33, the limit of detection was ca. 60 micrograms in 48-h urine: 2-15% of the quinine consumed was excreted as the parent compound in the first 48 h after consumption. The combination of the extraction procedure and the operational system makes the method suitable for the determination of a number of other alkaloids in physiological samples.