Perfusion Flow Rate

In practice, estimation of Laq requires information on the rate of solute removal at the membrane since aqueous resistance is calculated from experimental data defining the solute concentration profile across this barrier [7], Mean /.aq values calculated from the product of aqueous diffusivity (at body temperature) and aqueous resistance obtained from human and animal intestinal perfusion experiments in situ are in the range of 100-900 pm, compared to lumenal radii of 0.2 cm (rat) and 1 cm (human). These estimates will necessarily be a function of perfusion flow rate and choice of solute. The lower Laq estimated in vivo is rationalized by better mixing within the lumen in the vicinity of the mucosal membrane [6],... 

Yabannavar et al. [89] employed Eq. (11) to calculate the maximum perfusion flow rate attainable before cell wash-out, which occurs when cell growth is compensated by cell passage through the spin-filter or, according to Eq. (12), when Fapp = 0- Under these conditions ... 

Approach to zero Vary the perfusion flow rate (Q) and Requires steady-state concentrations at Possible, if it can be confirmed that... 

Perfusion flow-rates ranges between 0.1 and 5 til/min. The tendency is to use lower flow-rates as this may increase recovery, provided that an analytical technique is available to deal with the smaller sized samples. 

Since the first reports on microdialysis in living animals, there have been efforts to estimate true (absolute) extracellular concentrations of recovered substances (ZetterstrOm et al., 1983 Tossman et al., 1986). Microdialysis sampling, however, is a dynamic process, and because of a relatively high liquid flow and small membrane area, it does not lead to the complete equilibration of concentrations in the two compartments. Rather, under steady state conditions, only a fraction of any total concentration is recovered. This recovery is referred to as relative or concentration recovery, as opposed to the diffusion flux expressed as absolute or mass recovery. The dependence of recovery on the perfusion flow rate is illustrated in Figure 6.2. As seen, relative recovery will exponentially decrease with increasing flow as the samples become more... 

A number of variables, such as the perfusion flow rate, the membrane surface area and geometry, the MWCO of the membrane, the diffusion characteristics of the collected analyte, the composition of the perfusion medium, and the temperature, influence recovery parameters. When the microdialysis is carried out in vivo, the recovery can also be affected by some tissue properties, including tissue tortuosity, the extracellular space volume, the tissue blood fluid and the tissue metabolism of the substance. 

Skin pre-treatment Skin thickness Composition of perfusate Perfusate flow rate, temperature... 

Reasons for the discrepancies in fatigue resistance may stem from factors related to the nature of the experiment, including the in vitro or in vivo aspect of muscle perfusion, flow rate and blood/plasma contents (Schaafsma et al, 2002 Vedsted et aL, 2003). The hypothyroidism-induced reduction in cardiac output and blood perfusion during exercise (Bausch and McAllister, 2003 McAllister et al, 1991, 1995) may limit the supply of oxygen and metabolic substrates, and thus result in reduced fatigue resistance when measured in vivo. 

Consistent with the suggestion that removal rate is proportional to flow. Cross et al. (1994) showed that, after dermal application in the perfused rat limb preparation, doubling the perfusate flow rates led to 4, 2.3, and 2.6 times the outflow... 

Cross, S.E., Wu, Z.-Y., and Roberts, M.S., Effect of perfusion flow rate on the dermal tissue uptake of solutes after dermal application using the isolated perfused rat hindlimb preparation, J. Pharm. Pharmacol, 1994, 46, 844-850. 

Pang, K.S., Lee, W.F., Cherry, W.F, Yuen, V., Accaputo, J., Fayz, S., Schwab, A.J., and Goresky, C. A., Effects of perfusate flow rate on measured blood volume, disse space, intracellular water space, and drug extraction in the perfused rat liver preparation characterization by the multiple indicator dilution technique, J. Pharmacokinet. Biop-harm., 1988, 116, 595-632. 

In microdialysis, the recovery of analyte from the sample depends on a number of factors, including the chemistry of the analyte, temperature, perfusion rate, membrane surface area, membrane characteristics, and the nature of the sample (including its fluid volume percentage and whether it is in motion). MiCTodialysis is typically done at low perfusion flow rates (0.5 to 2 /d/min). As flow rate increases, relative recovery decreases, but absolute recovery increases. Relative recoveries from small membrane probes are in the 1-20% range. SubstantiaUy higher recoveries in the 50-80% range can be obtained with longer loop-type probes. 

There have only been a few reports of coupling microdialysis to microchip-based separation systems. Ideally, the microchip system should allow the injection of discrete sample plugs from a continuously flowing stream of dialysate without disturbing the separation element of the analysis. This allows maximal temporal resolution and limits the effect of perfusion flow rate on system performance. 

External microdialysis probes fabricated in-house were used in these experiments. Initial studies found that the addition of ethylenediamine tetraacetic acid (EDTA) to the derivatizing reagent was necessary to prevent divalent cations (Mg + and Ca +) present in the cerebral spinal fluid from decreasing the EOE. Using the device, separations could be carried out at 1.8 min intervals however, the effective temporal resolution was estimated to be between 2 and 4 min due to delay times attributed to the dead volume in the system. Increases in perfusion flow rates led to a decreased delay time but reduced recovery through the probe was observed. 

In order to conduct microdialysis experiments, several other components are required. Syringe pumps are often used to control the perfusate flow rate. The pump has to be able to deliver flow rates precisely in the microliter per minute range. Tubing is needed to connect between the probe and the pump which drives the perfusion flow and, in some cases, between the probe and a sample collector as well. The total dead volume of tubing should also be maintained as small as possible to have better time resolution. The perfusion fluid is a medium resembling the composition of extracellular fluid with minimal or zero concentration of the molecules of interest. Dial-ysate exiting from the outlet of the microdialysis probe is usually collected in a vial for later analysis. It is also possible to coimect the outlet directly to an analysis instrument without using a collector, which is usually preferred, if possible, for its convenience and usually faster analysis results. 

By plotting the left side of Eq. 8 as a function of perfusion flow rate (1/2), the permeability of the membrane to the molecule of interest (the slope is KA) may be determined to characterize the system functionality. 

Because recovery is determined by a competition between a diffusional and perfusion convective flux, it is perfusate flow rate dependent. 

Perfusate Flow Rate Variation Calibration is accomplished by placing a probe in the tissue of interest and varying the perfusate flow rate. At low flow rates, the dialysate concentration will reach a plateau, which is assumed to be near 100 % recovery. Recoveries at different perfusate flow rates can then be calculated. 

This is a method enabling the estimation of the in vivo recovery of an analyte of interest introduced by Loimroth in 1987. The procedure involves adding the anal3 te of interest into the perfusion solution at different concentrations at a fixed perfusate flow rate and measuring the difference between perfusate and dialysate analyte concentration. The recovery represented by the left term of Eq. 3 can be assumed to be an unknown value P since the perfusate flow rate and resistance are left unchanged. When it is rearranged... 

By varying Cd,m and determining Cd,out> and plotting Eq. 12 as a function of Cd,in, the x-axis intercept is the actual recovery concentration C, and the slope reflects the recovery at the selected perfusate flow rate. 

By coupling microdialysis to capillary electrophoresis, much shorter sampling times can be used, since the separating step can be very fast. As mentioned in Chapter 6, injection volumes are in the nanoliter range. This allows not only short sampling times but also a low perfusate flow rate that vhll give better detection limits (Section 9.6.1). 

Because recovery is determined by a competition between a diffusional and perfusion convective flux, it is perfusate flow rate dependent. As demonstrated in Eq. (3), recovery increases as the flow rate decreases. Even though low flow rates result in high recovery, it is often restricted hy the reproducihUity of low flow rates supphed hy syringe pumps and the sample volume needed for solution analysis. Therefore, extremely low flow rates are often not apphcahle due to limited analysis time resolution because of the long times needed to collect the appropriate solution volume for analysis. 

---