Determination of time delay between blood and interstitial adipose tissue glucose concentration change by microdialysis in healthy volunteers.

For the development and use of subcutaneous glucose sensors it is important to know the time lag between changes in blood glucose and subcutaneous interstitial glucose concentration. To determine the time lag we inserted a microdialysis probe into the abdominal subcutaneous adipose tissue of healthy volunteers (n = 19) and performed oral glucose tolerance tests (n = 39) over a 7-day period. After correction for the microdialysis system time lag, we compared the change in dialysate glucose concentration with the capillary blood glucose concentration. We found no significant delay time between a change in capillary blood glucose concentration and subcutaneous interstitial fluid glucose concentration using the Mann-Whitney test. The substantial interindividual variation of glucose recovery and the changing recovery in time makes it difficult to draw unambiguous conclusions about the exact physiological time lag. Based on the present experimental findings and theoretical calculations of glucose transport in adipose tissue, the physiological lag time is short and negligible compared to the system delay time of a glucose sensor.

Microdialysis measurement of glucose in subcutaneous adipose tissue up to three weeks in type 1 diabetic patients.

BACKGROUND: Microdialysis of subcutaneous adipose tissue may provide an opportunity to monitor glucose continuously, when the device is connected to an extracorporal glucose sensor. We assessed whether our microdialysis probes are capable of measuring adipose tissue glucose over a prolonged period in Type 1 diabetic patients. Furthermore, the relationship between abdominal skinfold thickness and glucose recovery and the effect of spontaneous glucose excursions on its recovery were evaluated. METHODS: Microdialysis probes were pairwise inserted subcutaneously into the abdominal fat and remained in situ for 3 weeks in eight Type 1 diabetic patients. At days 1, 3, 4, 8, 11, 16, and 18 of probe retention, glucose, as measured by microdialysis, was compared to capillary blood glucose concentrations during a 4 h period. The recovery of glucose obtained by microdialysis was expressed as a percentage of the capillary blood glucose concentration. RESULTS: Eleven of the 16 inserted probes (69%) were evaluable during the complete study. Recovery of glucose was lower at day 1 and 3 (51+/-23% and 56+/-18%, respectively, mean+/-S.D.) compared to values found afterwards (67+/-19%, 72+/-13%, 76+/-14%, 71+/-16%, and 76+/-18%, for day 4, 8, 11, 16, and 18, respectively, for all P<0.05 vs. day 1 and 3). Skinfold thickness was inversely related to the overall 3 week glucose recovery (r=-0.76; P<0.03). Recovery was similar over a wide range of capillary blood glucose concentrations. CONCLUSIONS: Prolonged in vivo retention of microdialysis probes improves the recovery and lowers the variability of adipose tissue-sampled glucose in Type 1 diabetic patients. These findings show that microdialysis-based glucose measurements offer an opportunity for prolonged glucose monitoring.

Effects of glucose and insulin levels on adipose tissue glucose measurement by microdialysis probes retained for three weeks in Type 1 diabetic patients.

BACKGROUND: To evaluate the effects of acute hyperglycaemia and hyperinsulinaemia on adipose tissue glucose measurements by microdialysis probes inserted for a 3-week period. METHODS: Microdialysis probes were implanted pairwise in abdominal adipose tissue in seven Type 1 diabetic patients and remained in situ during the complete study. Stepped hyperglycaemic hyperinsulinaemic clamps were performed at weekly intervals at which the probes were prepared for microdialysis. Adipose tissue glucose by microdialysis was compared to venous and capillary blood glucose concentrations. RESULTS: The mean time after which the acute rise in blood glucose was first detected was 11.3 min, which corresponds to the system delay of the microdialysis probe. The increase of the glucose concentration in dialysate was completed during the following 16 min. Hyperglycaemia and hyperinsulinaemia did not influence recovery compared to venous blood glucose concentrations, while recovery values compared to capillary blood glucose levels increased slightly under hyperinsulinaemic conditions (P<0.01). CONCLUSIONS: In Type 1 diabetic patients, recovery of glucose in adipose tissue compared to venous and capillary blood does not decrease during acute hyperglycaemia and hyperinsulinaemia. Although there is still a relevant time-delay to monitor a rise in blood glucose, these results show that microdialysis may offer an opportunity for future glucose monitoring over a prolonged time-period.

Microdialysis of glucose in subcutaneous adipose tissue up to 3 weeks in healthy volunteers.

OBJECTIVE: To measure possible changes in dialysate glucose concentrations over time, to validate the diffusional model for glucose transport from tissue to the probe, and to evaluate the actual glucose concentration in adipose tissue. RESEARCH DESIGN AND METHODS: Glucose concentrations in the subcutaneous adipose tissue of five healthy subjects (age 25 +/- 2.7 years, BMI 23.2 +/- 2.3 kg/m2 [mean +/- SD]) were measured by the microdialysis technique and compared with blood glucose. We applied microdialysis probes with hollow fibers of various membrane length (10-35 mm), used eight perfusion flow rates (0.5-20 microl/min), and perfused four glucose solutions (0.0, 2.8, 8.3, 11.1 mmol/l). RESULTS: After implantation, a substantial decrease in glucose recovery to the lowest value of 26 +/- 10% of the final plateau value was noted during the first few hours (n = 4). Recovery increased and stabilized after 5-9 days at 84.0 +/- 7.4% of capillary blood glucose when a flow rate of 0.5 microl/min was applied. According to the zero net-flux method, the glucose concentration in equilibrium, Cequi, with the surrounding tissue can be obtained. This concentration also decreases; however, 1 h after recovery, Cequi increases again over 1 or 2 days to a stable value that is not significantly different from the measured capillary blood glucose (P < 0.05). Using various perfusion flow rates and probes (membrane length 10-35 mm), it is shown that diffusion is the rate-limiting process for glucose transport through tissue. CONCLUSIONS: Insertion of the microdialysis probes causes damage to the adipose cells and the vascular bed around the probe. Glucose recovery decreases because of a lower blood supply. In 5-9 days, glucose recovery increases; apparently, this time is needed to repair the microstructure of tissue around the probe. After stabilization of the recovery, no loss of probe permeability, which is due to biocompatibility problems, was seen. The change during the 2 days in equilibrium concentration is probably caused by an inflammation reaction that consumes glucose around the probe. The individual increase in recovery during the 1st days after probe insertion until a stable plateau value is reached (flow rate >0 microl/min) is complicated for short-term clinical glucose measurements in adipose tissue. After stabilization, the mean equilibrium concentration of all subjects was equal to the mean capillary blood glucose concentration. Therefore, we conclude that capillary blood glucose concentration probably is the driving force for diffusion through the capillary wall into the probe and is not some interstitial concentration.

Glucose concentration in subcutaneous extracellular space.

OBJECTIVE: To compare the subcutaneous glucose sensor measurements with two reference methods. Previous studies provide conflicting findings about the real glucose concentrations in subcutaneous tissue. Some suggest substantially lower concentration, whereas others measure proportionally higher glucose concentrations compared with the blood compartment. Before these results can be taken seriously as an expression of the real glucose concentration in the extracellular space, the measurements must be validated by an independent method. RESEARCH DESIGN AND METHODS: We applied a microdialysis-based enzyme sensor to measure glucose concentration in subcutaneous tissue. We also developed two reference methods: subcutaneous filtrate collection and an equilibration method using ultrafiltration membranes to support the earlier findings. We provided an anatomical model to explain the results. RESULTS: The mean overall intercellular filtrate glucose concentration, sampled with the filtrate collector and taken after a 6-h stabilization time, including the values during the glucose clamp period, was 46 +/- 9%. The mean subcutaneous glucose concentration measured with the glucose sensor, calibrated in vitro, was 44 +/- 8% of the mean venous blood glucose concentration. Mean overall intercellular equilibrate glucose concentration, i.e., the mean glucose concentration in the subcutaneous extracellular space, taken after a 4-h stabilization time, was 46 +/- 15% of the mean venous blood glucose concentration. CONCLUSIONS: The close agreement between the mean values of subcutaneous glucose concentrations, obtained with three independent methods--filtration, equilibration, and dialysis (sensor)--shows the real glucose concentration in subcutaneous interstitial fluid is approximately 50% the blood glucose value in normal humans. Our results clarify some of the conflicting evidence presented in previous studies.

Calibration of a wearable glucose sensor.

Calibration of glucose sensors proved difficult for electrodes with immobilized glucose-oxidase. The correlation between the sensitivity of the electrodes in vitro and in vivo appeared to be poor. We developed a new type of glucose sensor, based on a microdialysis system, in which an oxygen electrode is used as detector outside the body and the enzyme glucose-oxidase dissolved in water is used as a dynamic selector. The enzyme solution is pumped through a hollow fiber placed subcutaneously, before the fluid passes the detector. The glucose sensor was tested in the subcutaneous abdominal tissue of 12 healthy volunteers and 12 type I diabetic patients. Blood glucose was clamped at two levels to permit a two-point calibration of the sensor in vivo. These values correlated well with the in vitro calibration factors (r = 0.949). In subcutaneous tissue the sensor measures 43 +/- 9% of the blood glucose value, using the in vitro calibration factor. No differences were detected between healthy volunteers and diabetic patients.

Development of a wearable glucose sensor; studies in healthy volunteers and in diabetic patients.

A glucose sensor with a subcutaneous dialysis system was tested in six healthy volunteers during an oral glucose tolerance test and in ten diabetic patients with hyperglycemia during rapid decline of blood glucose levels. There was a good correlation between sensor and blood glucose values. During oral glucose tolerance tests in the volunteers, there was a mean delay of 4.4 minutes in the rise of the value registered subcutaneously and of 8.2 minutes in the fall of the curves. In the diabetic patients the maximum delay was 22 minutes. Nine days after insertion of the dialysis system it was still functioning well.

The influence of pH on rectal absorption of sodium benzoate studied in man by rectal lumen perfusion.

The influence of pH on rectal absorption of sodium benzoate in man was studied by means of a rectal lumen perfusion method and compared with in vitro measurements on diffusional transport of sodium benzoate across an octanol/water interface. For nonbuffered solutions of benzoate in vitro, it was shown that mass flux across an octanol/water interface occurs in agreement with the pH-partition model. In vivo however, mass flux increases less with decreasing pH of unbuffered perfusate than is anticipated on the basis of the pH-partition model. Probably an alkaline flow across the rectal mucosa into the lumen is present as a physiological neutralization mechanism. In contrast, buffered solutions of benzoate show a linear relationship between mass flux and decreasing pH in vitro as well as in vivo. The effect of buffer on the concentration profile of benzoic acid is qualitatively explained. It is shown that an alkaline flow across the rectal mucosa only slightly influences absorption of benzoic acid from strongly buffered solutions in the rectal lumen. It is concluded that the use of strong buffers in rectal solutions induces a drastic effect on the pH of the boundary layer, an effect not seen for unbuffered solutions. This phenomenon does not invalidate the pH-partition hypothesis but can be explained by it.

Development of a potentially wearable glucose sensor for patients with diabetes mellitus: design and in-vitro evaluation.

A potentially wearable glucose sensor was developed, consisting of an oxygen electrode as detector and a dynamic enzyme perfusion system as selector. The selector is a hollow fibre, which can be placed subcutaneously and dialyses glucose from tissue fluid. In this design the problems of enzyme instability and oxygen limitation might be circumvented. The sensor measures glucose reliably for over two weeks, provided a new 10 ml syringe containing a glucose oxidase solution is connected to the system each day.

Lumen perfusion of the human rectum in situ: a method to study mechanisms for rectal drug transport in humans.

A lumen perfusion system was developed to study rectal transport mechanisms in humans. With this technique it is possible to perfuse a well-defined area of the rectum wall under single-pass and recirculation conditions. Sodium benzoate was used as a test drug. After absorption, sodium benzoate is conjugated with glycine to give hippuric acid which is rapidly eliminated (t 1/2 = 0.5 h). Due to the short half-life it is possible to reach a steady-state concentration within 2.5 h of perfusion. Plasma concentrations of hippuric acid were determined by HPLC. The absorption influx of sodium benzoate per unit area (phi in) could be calculated using the steady-state concentration of hippuric acid during rectal perfusion, the separately measured total body clearance after intravenous injection, and a designated absorption surface of the rectum. It was shown that with this technique reproducible phi in values within one subject could be obtained. Four volunteers were perfused with four different solutions of sodium benzoate, and it was found that phi in was proportional to the four concentrations used. In the case of recirculation perfusion (two volunteers), it was found that the amount of the perfusate lost equalled the amount absorbed into the general circulation. Therefore, the possibility of major drug accumulation in the rectal lumen or mucosa could be excluded. The perfusion technique elaborated in the present study can be used to investigate the mechanism of rectal absorption in humans as well as the factors that may influence this process.

Solution rate of crystals at fluid-fluid interface.

A model system was developed in which the dissolution behavior of a single crystal of potassium ferricyanide was studied at a liquid paraffin-water interface. Since the equilibrium position of a crystal at the interface is independent of its size, the lifetime of a crystal dissolving at the interface is determined entirely by its initial size and its dissolution rate in the water phase. The dimensions of every crystal were measured microscopically before dissolution. A continuous-flow recording dissolution apparatus was used to measure spectrophotometrically the mass flow of dissolved potassium ferricyanide. The dissolution cell in this system was mounted in an optical bench, making it possible to follow dissolving crystals visually by projecting them on a screen. The results show that the lifetime of a crystal is proportional to the shortest length of the crystal face in contact with the liquid paraffin and is rather independent of its form. Furthermore, crystal shape changes during dissolution, which is explained partly by the nonisometric dissolution of potassium ferricyanide crystal faces and partly by the nonconstancy of film thickness.

Contact angles and wetting of pharmaceutical powders.

Contact angles of pharmaceutical powders were determined by the h-epsilon method, which consists essentially of measuring the maximum height of a drop of liquid fomed on a presaturated compact of the material. Determinations with aspirin as the test material indicate that the measured value is independent of the particle size of the powder and the porosity of the cake. The method was extended to include determinations on mixed powder systems. The results show that the hydrophobic material dominates with large particle-size powders; with small particle sizes, a linear relationship between the cosine of the contact angle of the mixed system and the proportion of the components is obtained. Results are presented for a wide variety of materials of pharmaceutical interest.