**4. Real-time monitoring of renal clearance**

In vivo noninvasive real-time monitoring of renal clearance, with eventual translation to commercial development, has been demonstrated in the rodent model. A schematic of an apparatus is shown in Fig. 6. A 445 nm solid state laser was directed into one leg of a silica

Exogenous Fluorescent Agents for the Determination of Glomerular Filtration Rate 257

bifurcated fiber optic bundle, with the common end of this bifurcated bundle placed approximately 2 mm from the rat ear. The second leg of the bifurcated fiber optic bundle was fitted with a collimating beam probe. A long pass filter and narrow band interference filter were placed in front of a photosensor module. A chopper was placed after the laser and before the launch into the bifurcated cable. The output of the photosensor was connected to a lock-in amplifier. The lock-in output was digitized and the digitized data was

Anesthesized Sprague-Dawley rats of weight ~ 400 g were used. A volume of 1 mL of a 0.4 mg/mL concentration in PBS of compound 12 was administered to a rat with normal functioning kidneys and to a rat with a recent bi-lateral nephrectomy. The continuously monitored fluorescent signal is shown in Figure 5. An increase in fluorescence at the ear is immediately seen in both rats. In the normal rat, the fluorescence decreases back to baseline as the kidney removes compound 12 from the body. In the ligated rat, the fluorescence remains elevated with time as the body is unable to remove compound with the kidneys not functioning.

Fig. 7. Continuous Monitoring of Pyrazine **12** in Normal and Partially Nephrectomized Rats.

On the basis of the fluorescence properties, plasma protein binding data, the injected dose recovered in urine, the plasma clearance data, and the renal tubular secretion studies, the pyrazine deriviatives **12-15** are promising candidates as exogenous fluorescent tracer agents for the determination of GFR under both chronic and acute settings. In the rat model, these compounds display superior properties compared to iothalamate, which is currently an

A prototype instrument for clinical trials has been developed based on the apparatus in Figure 4. A clinical trial with one of the pyrazine compounds is currently being planned.

acquired by computer using data acquisition software.

**5. Conclusions** 

accepted standard for the measurement of GFR.

Fig. 5. Optical Image of Pyrazine **13** at 1 Hour Post Administration.

Fig. 6. Apparatus for non-invasive in vivo detection of fluorescence.

bifurcated fiber optic bundle, with the common end of this bifurcated bundle placed approximately 2 mm from the rat ear. The second leg of the bifurcated fiber optic bundle was fitted with a collimating beam probe. A long pass filter and narrow band interference filter were placed in front of a photosensor module. A chopper was placed after the laser and before the launch into the bifurcated cable. The output of the photosensor was connected to a lock-in amplifier. The lock-in output was digitized and the digitized data was acquired by computer using data acquisition software.

Anesthesized Sprague-Dawley rats of weight ~ 400 g were used. A volume of 1 mL of a 0.4 mg/mL concentration in PBS of compound 12 was administered to a rat with normal functioning kidneys and to a rat with a recent bi-lateral nephrectomy. The continuously monitored fluorescent signal is shown in Figure 5. An increase in fluorescence at the ear is immediately seen in both rats. In the normal rat, the fluorescence decreases back to baseline as the kidney removes compound 12 from the body. In the ligated rat, the fluorescence remains elevated with time as the body is unable to remove compound with the kidneys not functioning.

Fig. 7. Continuous Monitoring of Pyrazine **12** in Normal and Partially Nephrectomized Rats.
