Recovery of small bioparticles by interfacial partitioning.

In this article, a qualitative study of the recovery of small bioparticles by interfacial partitioning in liquid-liquid biphasic systems is presented. A range of crystallised biomolecules with varying polarities have been chosen such as glycine, phenylglycine and ampicillin. Liquid-liquid biphasic systems in a range of polarity differences were selected such as an aqueous two-phase system (ATPS), water-butanol and water-hexanol. The results indicate that interfacial partitioning of crystals occurs even when their density exceeds that of the individual liquid phases. Yet, not all crystals partition to the same extent to the interface to form a stable and thick interphase layer. This indicates some degree of selectivity. From the analysis of these results in relation to the physicochemical properties of the crystals and the liquid phases, a hypothetical mechanism for the interfacial partitioning is deduced. Overall these results support the potential of interfacial partitioning as a large scale separation technology.

Caries experience of 15-year-old children in The Netherlands after discontinuation of water fluoridation.

In 1973 the fluoridation of drinking water in the Dutch town of Tiel was discontinued. In order to monitor the effect of this measure, the caries experience in 15-year-old children was investigated annually from 1979 to 1988, both in Tiel and in Culemborg. In the latter town the drinking water had never been fluoridated. The caries data of 15-year-old children examined between 1968 and 1969 in Tiel (children having used fluoridated water from birth) and Culemborg were used as historical controls. In Tiel the mean number of DMFS increased between 1968/69 and 1979/80 from 10.8 to 12.7 (+18%) and decreased to 9.6 (-26%) in the following years; in 1987/88 the mean DMFS was 11% lower than in 1968/69. In Culemborg the mean DMFS score decreased between 1968/69 and 1987/88 from 27.7 to 7.7 (-72%). In 1968/69 the mean DMFS score in Tiel was 61% lower and in 1987/88 17% higher than in Culemborg. The question as to whether water fluoridation would have had an additional effect if it had been continued (presuming the application of existing preventive measures) cannot be answered, as there are no remaining communities with fluoridated water in The Netherlands.

[Cessation of fluoridation of drinking water; results of caries research in Tiel and Culemborg in the period of 1968-1988]. / Stopzetting van drinkwaterfluoridering. Resultaten van het cariësonderzoek in Tiel en Culemborg in de periode 1968-1988.

In December 1973 water fluoridation was discontinued in Tiel, the Netherlands. Six years later an epidemiological study was started, with the aim to investigate the consequences. In the period from 1979 to 1988 a yearly caries investigation was carried out in 15 year old children born and living in Tiel and children living in the control town of Culemborg, where the drinking water had never been fluoridated. The 15 year old children in 1979-1980 in Tiel had a 18% higher caries experience than the same age group in 1968-1969, to whom fluoridated water had been available from birth onward. In the same period a 28% decrease of the caries was found in Culemborg. In 1979-1980 the DMFS-index in Tiel was still 36% lower than in Culemborg. Between 1979-1980 and 1987-1988 the caries experience decreased in both towns.

Reliability of the determination of whole-blood oxygen affinity by means of blood-gas analyzers and multi-wavelength oximeters.

Determination of the oxygen affinity of human whole blood with the help of blood-gas analyzers and multi-wavelength oximeters is compared with an accurate method for recording hemoglobin oxygen dissociation curves (Clin Chem 1982;28:1287-92). P50 (oxygen tension at half saturation; So2 = 50%) and Hill's n (delta log [So2/(1-So2)]/delta log Po2) were determined in blood of 24 healthy donors. Three slightly different procedures were used for determination of P50 on the basis of Po2, pH, Pco2, and So2 measured with each of four different blood-gas analyzer/oximeter combinations. These methods were not able to discriminate between high and low values for P50 within the normal reference interval, but never failed to detect the high oxygen affinity of blood stored for 12 days, reflected in low values of P50. The methods thus proved suitable for detecting clinically significant deviations in oxygen affinity such as occur in patients with, e.g., abnormal hemoglobins, anemias, or carbon monoxide poisoning. Determination of Hill's n by these methods did not produce useful results.

Human whole-blood O2 affinity: effect of CO2.

The proton Bohr factor (phi H = alpha log PO2/alpha pH), the carbamate Bohr factor (phi C = alpha log PO2/alpha log PCO2), the total Bohr factor (phi HC = d log PO2/dpH[base excess) and the CO2 buffer factor (d log PCO2/dpH) were determined in the blood of 12 healthy donors over the whole O2 saturation (SO2) range. All three Bohr factors proved to be dependent on SO2, although to a lesser extent than reported in some of the recent literature. At SO2 = 50% and 37 degrees C, we found phi H = -0.428 +/- 0.010 (SE), phi C = 0.054 +/- 0.006, and phi HC = -0.488 +/- 0.007. The values obtained for phi H, phi C, and d log PCO2/dpH were used to calculate phi HC. Calculated and measured values of phi HC proved to be in good agreement. In an additional series of 12 specimens of human blood we determined the influence of PCO2 on phi H and the influence of pH on phi C. At SO2 = 50%, phi H varied from -0.49 +/- 0.009 at PCO2 = 15 Torr to -0.31 +/- 0.010 at PCO2 = 105 Torr and phi C from 0.157 +/- 0.015 at pH = 7.80 to 0.006 +/- 0.009 at pH = 7.00. When on the basis of these data a second-order term is taken into account, a still slightly better agreement between measured and calculated values of phi HC can be attained.

Calibration of a practical haemoglobinometer.

A practical instrument for routine haemoglobinometry (HemoCue) was compared with the reference haemiglobincyanide (methaemoglobincyanide (HiCN] method. The precision of the routine method was adequate, but a systematic error of about -3.5% was found. As this resulted from errors in the procedure for coupling the haemoglobinometer read-out to the reference HiCN method, we drew up an improved calibration procedure for practical haemoglobinometers to ensure comparability of results with the reference method. After recalibration according to these rules, the HemoCue system proved to be accurate to within +/- 1.5%.

Human whole-blood oxygen affinity: effect of temperature.

phe effect of temperature changes on human whole-blood O2 affinity was measured in the blood of six healthy donors over almost the entire O2 saturation (SO2) range (1-99%). The results showed that temperature has no influence on the shape of the O2 dissociation curve, implying that the temperature coefficient (delta log PO2/delta T) is independent of SO2. Simultaneous measurements of the total (proton) Haldane factor (delta[HbH]/[delta HbO2]) at the five temperatures under study (22, 27, 32, 37, and 42 degrees C) revealed that this factor depends on temperature. The liberation of protons from hemoglobin appeared to be linear with respect to changes in SO2. We therefore conclude that the (proton) Bohr factor (H+ factor) is dependent on temperature over the entire SO2 range in the same way as previously described for SO2 = 50%. The exothermic oxygenation reaction in whole blood was accompanied by a heat evolution (delta HO2) of 42.7 kJ/mol (monomeric) hemoglobin.

Human whole-blood oxygen affinity: effect of carbon monoxide.

Oxygen dissociation curves (ODC) were recorded in the presence of carboxyhemoglobin fractions (FHbCO) up to 60%. The gradual shift to the left of the ODC at increasing amounts of HbCO was reflected in a gradual fall in the half-saturation pressure of the remaining Hb and was accompanied by a gradual change in the shape of the ODC to a hyperbolic one. The H+ factor (delta log PO2/delta pH) was determined over the entire oxygen saturation (SO2) range at three different FHbCO levels (14, 30, and 52%). At FHbCO = 14 and 30% and for the SO2 range 20-90%, the H+ factor vs. SO2 curve was not significantly different from that in the absence of HbCO. At FHbCO = 52%, however, the value found for the H+ factor (-0.55) was appreciably more negative than in the case of blood containing less than 1% HbCO (-0.44), and there was no dependence on SO2. Comparison of measured and calculated ODCs at varying HbCO fractions showed, for FHbCO less than or equal to 50%, that measured and calculated ODCs coincide over the greater part of the SO2 range. For FHbCO greater than 50%, the measured ODC was situated to the left of the calculated one over the entire SO2 range. We conclude that the heme-heme interaction for CO is appreciably larger than for O2 only for FHbCO greater than 50%, whereas for FHbCO less than 50% there is virtually no difference.

Oxygen dissociation curves for whole blood, recorded with an instrument that continuously measures pO2 and sO2 independently at constant t, pCO2, and pH.

We describe a method for recording oxygen dissociation curves for whole-blood specimens. The blood sample is placed in a thermostated measuring chamber, and pO2 and SO2 are measured continuously by polarography and by reflectometry, respectively. During the recording of an oxygen dissociation curve, the pO2 and SO2 signals are stored in a data-acquisition system, while pH, pCO2, and temperature are kept constant. Determination of precision and error discussion indicated that the coefficient of variation (CV) of the determination of the oxygen dissociation curve is mainly determined by the error in the measurement of SO2. The overall CV of pO2 values belonging to the lower, mid-, and upper parts of the SO2 range is estimated to be about 2.6, 3.1, and 2.1%, respectively. In practice the measurements are about 30% more precise than estimated. With our method, the fixed-acid-induced Bohr effect (H+ factor) can be determined over the entire SO2 range with much greater precision than hitherto.

Determination of interdependent ligand effects on human red cell oxygen affinity.

Human whole blood oxygen affinity was determined as P50, i.e. PO2 for oxygen saturation (SO2) = 0.5, in a new system which allows the measurement of SO2, PO2 and pH continuously and independently, with control of PCO2 and temperature. The influence of pH on P50 expressed as the H+ factor (delta log P50/delta pH)PCO2 was measured under conditions of varying PCO2, temperature and concentration of 2,3-diphosphoglycerate (2,3-DPG), resulting in a set of data expressing second-order inter-ligand interactions. The H+ factor appeared to be only slightly dependent on PCO2. Similarly, the CO2 factor (delta log P50/delta log PCO2)pH shows only a minor dependence on pH. The H+ factor in linearly related to the temperature: at 17 degrees C and 42 degrees C the H+ factor is about -0.53 and -0.36, respectively. Likewise, the temperature factor (delta log P50/delta T)PCO2,pH is linearly related to pH. A pilot study on the effect of varying intra-erythrocytic 2,3-DPG concentrations on the oxygen affinity showed that a very low 2,3-DPH/Hb4 ratio apparently does not influence the H+ factor. A high ratio, however, seems to lower the H+ factor considerably.

Influence of variations in pH and PCO2 on scalp tissue oxygen tension and carotid arterial oxygen tension in the fetal lamb.

A description is given of the effect of hypercapnic acidaemia and hypocapnic alkalaemia on scalp tissue PO2 as measured with a subcutaneous needle-electrode and a transcutaneous electrode in 6 fetal lambs. The experiments were carried out under general anaesthesia with the fetus kept in utero. Hypocapnia was induced by hyperventilating the ewe and hypercapnia was achieved by administering extra CO2 to the ewe. Fetal carotid arterial, subcutaneous and transcutaneous PO2 were continuously recorded, and fetal and maternal arterial pH and arterial PCO2 were determined from blood samples taken at short intervals. In each experiment the H+ Bohr factor of fetal and maternal blood was measured. During hypocapnic alkalaemia, there was a fall in all fetal PO2 levels, whereas a marked rise was observed during hypercapnic acidaemia. The variations in fetal PO2 observed in vivo even exceeded the variations due to the H+ Bohr effect (measured in vitro). This was due to small variations in fetal carotid arterial oxygen saturation, which tended to fall during hypocapnic alkalaemia and to rise during hypercapnic acidaemia. The results of these findings strongly suggest that tissue PO2, as measured with the subcutaneous and transcutaneous electrodes, is dependent on the H+ Bohr effect. This adds to the uncertainty as to the value of subcutaneous and transcutaneous PO2 monitoring during labour as an early warning system for impending fetal asphyxia.

Continuous intra-arterial pH measurement.

A flexible glass electrode catheter with a diameter of 3 mm has been developed for pH measurement in the arterial system of dogs. In combination with a galvanically isolated amplifier, an undisturbed pH signal could be obtained from the aorta. The system was fast enough to truly record pH changes synchronous with respiration and was shown to be insensitive to variations in blood flow velocity. Good agreement was found between pH catheter readings nd pH values of simultaneously taken arterial samples as measured with a conventional capillary glass electrode.

Continuous measurement of scalp tissue oxygen tension and carotid arterial oxygen tension in the fetal lamb.

Scalp tissue PO2, carotid arterial PO2 fetal heart rate were continuously measured in the anaesthetized fetal lamb in utero while variations in oxygen supply were brought about. In some experiments the transcutaneously measured fetal scalp PO2 was recorded in addition. Scalp tissue PO2 was measured using specially designed miniature needle-type oxygen electrode, incorporated in an easily applicable spiral scalp electrode as commonly used for fetal heart rate monitoring. The measurements showed that fetal carotid arterial hypoxaemia is always nearly immediately followed by fetal scalp tissue hypoxia, and that the recovery of scalp tissue PO2 after a hypoxic period has a remarkably varying time course. Fetal heart rate usually decreased during hypoxia, but in some instances it did not change or even increased, demonstrating that heart rate is not always a reliable indicator of fetal hypoxia. PO2 values obtained with the transcutaneous method were higher than those with the needle electrode, because of the effect of the heating system of the transcutaneous electrode on tissue blood flow and haemoglobin oxygen affinity. It would seem that during hypoxaemia the decrease in scalp tissue PO2 is poossibly the combined result of the fall in arterial PO2 and a concomitant decrease in blood flow through the skin.

Spectrophotometric determination of oxygen saturation of blood independent of the presence of indocyanine green.

The strong absorbance of indocyanine green in a broad band around lambda = 800 nm invalidates the usual spectrophotometric two-wavelength methods for measuring oxygen saturation operating in the red and near infrared region. By proper wavelength selection, however, the effect of the dye can be eliminated. With the two-wavelength method utilising lambda = 660 and 860 nm oxygen saturation is measured virtually independent of the presence of indocyanine green.