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At least 19 recordsLinked to original sources
PubMed2002

In vitro human scleral permeability of fluorescein, dexamethasone-fluorescein, methotrexate-fluorescein and rhodamine 6G and the use of a coated coil as a new drug delivery system.

PURPOSE: To determine the in vitro human scleral permeability of several dyes and drugdye combinations with varying molecular weights (MW) and lipid solubilities (fluorescein, dexamethasone-fluorescein, methotrexate-fluorescein, and rhodamine). Coils coated with rhodamine were also evaluated for scleral permeability and sustained release. METHODS: Scleral sections excised from moist chamber stored human globes were mounted in a 2-compartment perfusion chamber. A small depot of drug/dye (100 microl of 10(-4) M fluorescein, dexamethasone-fluorescein, methotrexate-fluorescein or rhodamine) or a coated coil in 100 microl of BSS was added to the episcleral surface while perfusing BSS to the choroidal side. The perfusate was collected and measured for fluorescence. Permeability was calculated as Ktrans from the flux measurements. RESULTS: Ktrans values (cm/sec, mean +/- SE) for the studied dyes and drug-dye combinations were 5.21 +/- 0.71 x 10(-6) for fluorescein, 1.64 +/- 0.17 x 10(-6) for dexamethasone-fluorescein, 3.36 +/- 0.62 x 10(-6) for methotrexate-fluorescein, 1.86 +/- 0.39 x 10(-6) for rhodamine and 2.18 +/- 0.23 x 10(-6) for the rhodamine from the coils. We found a significant difference between the permeability of the sclera to fluorescein and dexamethasone-fluorescein (P < 0.001), methotrexate-fluorescein (P < 0.05) and rhodamine (P < 0.001). Steady state flux was observed from the rhodamine coil. CONCLUSION: The rank order of scleral permeability to the studied dyes is as follows: fluorescein > methotrexate-fluorescein > rhodamine coil > rhodamine 6G > dexamethasone-fluorescein. Differences in scleral permeability are related to MW and lipid solubility. Prolonged transscleral diffusion of rhodamine delivered by solution and by coil are similar.

Coated Materials, Biocompatible↗
PubMed1986

Study of fluorescein glucuronide. II. A comparative ocular kinetic study of fluorescein and fluorescein glucuronide.

Comparative studies of fluorescein and fluorescein glucuronide were carried out. Binding to human serum protein was studied using an Amicon MPS-3 ultrafiltration unit; it averaged 63% for fluorescein glucuronide and 85% for fluorescein. Intracameral penetration of both compounds was studied in the human eye, and the concentration changes of both compounds in the plasma ultrafiltrate and in the anterior chamber were analyzed, based on Davson's equation. The coefficient of entry into the anterior chamber (ki) was 0.018 +/- 0.007 h-1 (mean +/- SD, n = 10) for fluorescein glucuronide and 0.054 +/- 0.033 h-1 for fluorescein, and the former was significantly lower than the latter (P less than 0.005). The rate of loss from the vitreous (kv) was studied by injecting each compound into the vitreous of the pigmented rabbit and following the fluorescein intensity changes in it. It was 0.042 +/- 0.008 h-1 (mean +/- SD, n = 8) for fluorescein glucuronide and 0.17 +/- 0.01 h-1 for fluorescein, and the former was significantly smaller than the latter (P less than 0.001). Intraperitoneal injection of probenecid significantly decreased the kv of fluorescein but had little effection that of fluorescein glucuronide. It was suggested that fluorescein glucuronide is lost from the vitreous mainly by a passive mechanism.

Adult↗
PubMed1991

The loss of fluorescein, fluorescein glucuronide and fluorescein isothiocyanate dextran from the vitreous by the anterior and retinal pathways.

The pathways by which fluorescein (F), fluorescein glucuronide (FG) and fluorescein dextran (FD) leave the vitreous body of the rabbit were examined by measuring the concentration distribution of the injected fluorophores in sections of the frozen eyes. The contours of F, as already known, show that it leaves the vitreous predominantly across the retinal surface. Mathematical analysis of the concentration gradient leads to an average outward permeability coefficient of 1.4 x 10(-3) cm min-1 for the retinal layers. The contours of FG and FD show that they leave predominantly by diffusion into the posterior chamber, encountering only a minor barrier at the anterior hyaloid membrane. The anterior contours indicate that there can be no substantial posteriorly directed fluid flow through the vitreous; if it occurs its velocity across the retinal surface must be less than 2 x 10(-5) cm min-1. The contours of FD near the posterior pole of the retina suggest that such a flow may be taking place. Some time after the systemic administration of F, an analysis of the rate of loss of fluorescence from the vitreous body shows that this corresponds to the movement of FG out through the anterior chamber. Its value bears little relationship to the condition of the blood-vitreal barrier.

Animals↗
PubMed1988

Fluorescein and fluorescein glucuronide in plasma after intravenous injection of fluorescein.

After intravenous injection of fluorescein the time-course of the plasma concentrations of fluorescein (F) and fluorescein glucuronide (FG) was studied in 18 insulin-dependent diabetics with various degrees of nephropathy, and in two non-diabetic subjects. Fifteen minutes after injection the molar concentrations of free (non-protein bound) F and FG were almost identical. After one hour the concentration curve integrals of the two substances were of the same magnitude. There was, however, considerable interindividual variation. In diabetic patients with renal insufficiency an increase was found in both integrals, the F integral being less increased (27%) than the FG integral (44%). It appears from the variation in absolute and relative concentrations of F and FG that a separate determination of the two fluorophores in plasma is desirable, when F is used by intravenous administration as an indicator of blood-ocular barrier function. Previous observations of an increase with time after injection of the free fraction of plasma fluorescence are explained by the finding of a higher free fraction of FG (34%) than of F (11%) and a change in relative concentrations of the two fluorophores.

Adult↗
PubMed1985

Charge transfer between fluorescein and tryptophan as a possible interaction in the binding of fluorescein to anti-fluorescein antibody.

Steady-state and time-resolved fluorescence studies of fluorescein (Fl) and 9-hydroxyphenylfluoron (HPF) bound to high-affinity rabbit anti-Fl IgG antibody (anti-Fl IgG) have been performed. The heterogeneity in the fluorescence properties observed for Fl bound to anti-Fl IgG is reduced for HPF bound to anti-Fl IgG. A charge transfer between a tryptophyl residue in the binding site and the hapten was considered as a possible binding interaction. Fl was observed to form complexes in solution with the amino acids tryptophan, tyrosine and methionine, probably due to charge transfer. Also, the fluorescence of tryptophyl residues of the protein is quenched on binding. While such charge transfer complexes may be present, there is no direct evidence that charge transfer complexes between Fl and tryptophan are necessarily present for Fl bound to all high-affinity anti-Fl IgG molecules.

Amino Acids↗
PubMed1986

Measurement of fluorescein and fluorescein monoglucuronide in the living human eye.

Fluorescein monoglucuronide is a fluorescent metabolite of fluorescein, and is 1/3 to 1/34 as fluorescent as fluorescein, depending on the wavelength of excitation. After systemic administration, fluorescein glucuronide reaches concentrations many times greater than fluorescein. In order to study the effect of fluorescein glucuronide on the measurement of ocular dynamics, we devised a technique to measure fluorescein and fluorescein glucuronide in the anterior segment of the living human eye. Concentrations of each fluorophore were determined by differential spectrofluorophotometry from measurements at excitation wavelengths of 457.9 nm and 488.0 nm. Measurements were made on normal volunteers after oral and intravenous administration of fluorescein. Fluorescein was the dominant fluorophore during the first hour, while fluorescein glucuronide became dominant after 3 hours. By 6 hours there was 10 to 30 times more fluorescein glucuronide than fluorescein in the anterior chamber after oral administration, and three to ten times more after intravenous administration. The blood aqueous diffusion coefficient kd estimated from the apparent concentration of fluorescein measured at 457.9 nm was consistently greater than kd estimated from measurements at 488.0 nm. Estimates of kd, which were made on the basis of concentrations of fluorescein determined from measurements at both wavelengths, were lower than estimates based on measurements at either wavelength. These results indicate that wavelength of excitation may influence the determination of ocular parameters when systemic fluorescein is used. Care must be taken in the interpretation of measurements when metabolites of a fluorophore can interfere with measurement of the fluorophore itself.

Eye↗
PubMed1987

Movement of fluorescein and fluorescein glucuronide across the isolated rabbit iris-ciliary body.

Movement of fluorescein and fluorescein glucuronide, a fluorescent metabolite of fluorescein, across the isolated iris-ciliary body of the albino rabbit was determined under short-circuit conditions using a modified Ussing's chamber. The permeabilities of this tissue to these dyes were calculated. The outward permeability (from the aqueous to the stromal side) of the iris-ciliary body preparation averaged 6.63 +/- 0.86 for fluorescein and 1.51 +/- 0.47 X 10(-6) cm/sec for fluorescein glucuronide, and the inward permeability (from the stromal to the aqueous side) was 1.68 +/- 0.41 for fluorescein and 1.37 +/- 0.77 X 10(-6) cm/sec for fluorescein glucuronide, respectively. Application of probenecid or ouabain decreased the outward permeability of fluorescein, but it had no significant effect on the fluorescein glucuronide movement. Application of 10(-5) M 2,4-dinitrophenol showed no significant effect on the fluorescein or fluorescein glucuronide movement, but application of 5 X 10(-4) M 2,4-dinitrophenol decreased the outward fluorescein transfer, which was also markedly suppressed by incubation at 0 degrees C. It is possible that an active transport mechanism is involved in the outward fluorescein movement across the iris-ciliary body, while the inward movement of fluorescein and also the fluorescein glucuronide movement across this tissue is mainly by passive diffusion.

2,4-Dinitrophenol↗
PubMed1985

Studies on the pharmacokinetics of fluorescein and its dilaurate ester under the conditions of the fluorescein dilaurate test.

Some aspects of the pharmacokinetics of fluorescein have been studied under the conditions of the fluorescein dilaurate test (Pancreolauryl-Test) in healthy volunteers. Dependence of fluorescein excretion on urine volume was investigated in a retrospective study in 370 patients. For intravenously administered fluorescein mean Cmax was 10.9 micrograms/ml with a mean elimination half-life of 286 min. For orally administered fluorescein sodium mean Cmax was 3.5 micrograms/ml with a tmax of 120 min and a t 1/2 of 267 min. Bioavailability of fluorescein by oral administration was 99%. By contrast, fluorescein from fluorescein dilaurate showed a 56% bioavailability under the conditions of the test with a Cmax of 1.8 micrograms/ml, a tmax of 270 min and a tt 1/2 of 246 min. Following enteral absorption of fluorescein hepatic extraction and enterohepatic circulation via the bile occurs, but although the concentrations of fluorescein in the bile may exceed those in the urine the absolute amount is likely to be small. In spite of the enterohepatic circulation fluorescein cleared from the urine within 24 h indicating that no delay between Part 1 and Part 2 of the test seems necessary. However, an adequate urine flow must be maintained throughout the test since a renal clearance/urine flow relationship exists, with fluorescein excretion being increased with increasing urine volume.

Administration, Oral↗
PubMed1998

Efficacy and safety of fluorescein angiography with orally administered sodium fluorescein.

PURPOSE: To report the efficacy and safety of fluorescein angiography after oral administration of fluorescein solution in a large number of patients during a period of 8 years. METHODS: A total of 1,787 patients (2,625 eyes) underwent fluorescein angiography after oral administration of sodium fluorescein at Hara Eye Hospital, Utsunomiya, Japan, between January 1989 and March 1997. The ingestible solution was 10 ml of 10% sodium fluorescein, the same material generally used for injection in conventional fluorescein angiography. Retinal photography began 15 minutes after ingestion and continued for 1 hour. The camera and the photography and film processing techniques were the same as those used for conventional fluorescein angiography using injected sodium fluorescein. RESULTS: In 2,554 (97.3%) of 2,625 eyes, photographs adequate for clinical use were obtained. In 1,787 patients, no anaphylactic or other severe adverse effects were observed, and only 31 patients (1.7%) experienced minimal itching, discomfort, or nausea after oral sodium fluorescein intake. For conditions such as central serous chorioretinopathy, retinal vein occlusion, diabetic retinopathy, and cystoid macular edema, sufficient information for clinical use was obtained. CONCLUSIONS: Fluorescein angiography using orally administered sodium fluorescein is generally effective and safe in standard clinical practice.

Administration, Oral↗
PubMed1992

Fluorescein and fluorescein glucuronide in plasma.

The evaluation of the blood-ocular barrier for fluorescein requires the measurement of free and unconjugated fluorescein in plasma. This study introduces a new and simple method for the determination of free fluorescein in plasma on the basis of determined total free plasma fluorescence and the free fraction of fluorescence. An excellent good correlation between differential spectrofluorophotometry and this new method is demonstrated. After intravenous administration of sodium fluorescein, the contribution of fluorescein glucuronide to total free plasma fluorescence was evaluated on basis of the areas under the plasma concentration/time curves for fluorescein and fluorescein glucuronide, respectively. After 1 h 8.2% of total free fluorescence in plasma was found to originate from fluorescein glucuronide and after 24 h 18.3% originated from this metabolite. It was concluded that although plasma fluorescein glucuronide measurements are important in the exact evaluation of the blood-ocular barrier, the contribution of fluorescein glucuronide to vitreous fluorescence after intravenous fluorescein administration seems to be of minor magnitude.

Adult↗
PubMed1989

Time-resolved fluorescence properties of fluorescein and fluorescein glucuronide.

The use of fluorescein as a tracer in the study of the blood-ocular barriers is complicated by the metabolic production of fluorescein monoglucuronide, the excitation and fluorescence spectra of which overlap with those of fluorescein. Time-resolved fluorescence measurements provide a means of detecting the two substances in a mixture. Fluorescein and fluorescein glucuronide have different fluorescence lifetimes, 4.0 nsec and 2.3 nsec, at 37 degrees C and pH 7.35. Hence the two substances can be distinguished from the biexponential fluorescence decay in the mixture. The lifetimes are identical in buffered water, in hyaluronic acid and in human vitreous in vitro. The method is suggested for estimation of fluorescein and fluorescein glucuronide in the human vitreous in vivo. The time-resolved and steady state fluorescence anisotropies, the fluorescence lifetimes and the quantum yields strongly suggest that binding of fluorescein or fluorescein glucuronide to hyaluronic acid or other macromolecules in the human vitreous is weak, if present at all, and that static quenching of fluorescence does not occur in the vitreous.

Fluorescein↗
PubMed1993

Ocular fluorometry methodological improvements and clinical studies--with special reference to the blood-retina barrier permeability to fluorescein and fluorescein glucuronide.

The measurement of fluorescence in the human eye can be made using relatively simple instruments. Fluorescence is evoked when illumination is absorbed by intrinsic fluorophores in the eye or by artificially introduced extrinsic fluorophores. Intrinsic fluorescence is evidence of important molecular characteristics of the ocular tissues, whereas the extrinsic fluorophores are used primarily in the study of the barriers between the anatomical and physiological compartments of the eye. Blood-retina barrier leakage of fluorescein can be examined after the intravenous injection of fluorescein by quantitative determination of fluorescence in plasma and in the vitreous. From these measurements of the distribution of fluorescein, the permeability of a hypothetical spherical interface between the blood and the retina can be estimated using a mathematical model of the barrier. The use of fluorescein as a tracer is problematic because of its rapid metabolic conversion to fluorescein glucuronide. This metabolite disturbs ocular fluorescence measurements because it fluoresces over the same part of the spectrum as the parent compound. Additionally, the glucuronide occurs in markedly different concentrations depending upon the patient's renal function. With the previously used fluorometry techniques it has been impossible to determine the contribution of fluorescein glucuronide to the vitreous fluorescence. The primary objective of the studies described in this thesis was to develop a method for the determination of fluorescein and fluorescein glucuronide in the human eye and in plasma, and to calculate the blood-retina barrier permeabilities of the two substances. The necessary methodological improvements included a detailed description of the geometrical optics of the eye and the optical filter properties of the lens. A new method was developed for the determination of the spatial locations of ocular fluorescence measurements and the intrinsic lens fluorescence was used to estimate lens transmittance. The new techniques were applied to clinical studies in patients with diabetic retinopathy. It was shown that in insulin-dependent diabetes mellitus, the apparent rate whereby fluorophores are accumulated in the lens is increased in inverse proportion to the quality of metabolic control, i.e. patients who have had consistently poor control have higher fluorescence than patients who have been in good control. An increase in lens fluorescence was also found in the presence of diabetic nephropathy. The results support the assumption that lens fluorometry can provide a rough estimate of cumulative glycaemia and that glucose is involved in certain age-related changes in the lens.(ABSTRACT TRUNCATED AT 400 WORDS)

Adult↗
PubMed1995

Plasma fluorescein binding and transcapillary fluorescein escape rate in renal failure associated with diabetes.

Microvascular damage, often resulting in renal failure, is a common complication of diabetes. Transcapillary fluorescein escape rate (TCFER) as monitored by intravital microscopy has been used as an indicator of the extent of capillary damage in diabetes and to assess improvement in microvascular function after combined kidney-pancreas transplant. However, fluorescein anion binds to plasma albumin, and albumin-ligand binding may be altered in the presence of renal disease. The purpose of this study was to compare fluorescein binding by plasma from diabetics with renal failure with plasma from healthy nondiabetics. Fluorescein binding by plasma from seven type I diabetics awaiting kidney-pancreas transplant and seven healthy adults of similar age and sex was studied using ultrafiltration and dialysis. There was no significant difference in the apparent albumin binding of fluorescein at physiologically relevant fluorescein concentrations, even though the TCFER was significantly increased in the diabetics as compared with the controls. Hippurate, a ligand that accumulates in renal failure, did alter fluorescein binding in a defatted albumin solution but not sufficiently to account for the differences in TCFERs. These data indicate that impaired albumin binding of fluorescein does not contribute significantly to the TCFER in diabetics with renal failure.

Adult↗
PubMed1993

Role of electrostatic interactions in the binding of fluorescein by anti-fluorescein antibody 4-4-20.

Anti-fluorescein antibodies are excellent model systems for studying the biochemical basis of molecular recognition because a prodigious amount of both physico-chemical and structural information is available for these antibodies. Furthermore, recombinant single-chain antibodies have been produced for several anti-fluorescein antibodies, and site-specific mutagenesis studies have defined the energetic contributions of a number of key active-site residues. In previous studies, we determined the three-dimensional structure of an antigen-binding fragment of a high-affinity anti-fluorescein antibody (4-4-20) in complex with fluorescein. These studies showed that fluorescein binds tightly in an aromatic slot and participates in a network of electrostatic interactions. In this report, we examine the role of electrostatic interactions in the 4-4-20 antigen-combining site by observing the effects of pH on the fluorescence of fluorescein and antigen-binding affinity. These studies showed that the salt link between fluorescein and Arg-L34 in 4-4-20 probably accounts for about -1.5 kcal/mol-1 of the observed free energy of interaction. Furthermore, at pH 10 and higher, the affinity decreases by more than 100-fold (delta delta G degrees approximately equal to 3 kcal mol-1). We attributed this decrease to the ionization of Tyr-L32, which probably disrupts a hydrogen bond between tyrosine's hydroxyl group and fluorescein's phenylcarboxylate group. The fluorescence lifetime of the 4-4-20/fluorescein complex was determined at both pH 8 and pH 10.6. Only one lifetime component (0.38 ns) was observed at pH 8, while two components (0.3 and 3.4 ns) were observed at pH 10.6.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1987

Human corneal endothelial permeability to fluorescein and fluorescein glucuronide.

The corneal endothelial permeability coefficient (Pac) for fluorescein and fluorescein glucuronide was determined in ten normal young volunteers. After oral administration of fluorescein, the apparent concentrations of both dyes in the corneal stroma and the anterior chamber were measured by differential fluorometry. The apparent dye levels calculated directly from the in vivo fluorometric measurements were converted to the true ones, based on the result of a normalization experiment performed in rabbit eyes. The value of Pac averaged 5.44 +/- 1.77 X 10(-4) cm/min for fluorescein and 3.77 +/- 1.10 X 10(-4) cm/min for fluorescein glucuronide (mean +/- SD, N = 20); the former was significantly greater than the latter (paired t-test, P less than 0.001). The aqueous-cornea distribution ratio was 0.50 +/- 0.14 for fluorescein and 0.66 +/- 0.16 for fluorescein glucuronide; the latter was significantly greater than the former (paired t-test, P less than 0.001). It was suggested that the previously reported values of Pac for fluorescein in the human eye were underestimates.

Administration, Oral↗
PubMed2002

Discrete bathochromic shifts exhibited by fluorescein ligand bound to rabbit polyclonal anti-fluorescein Fab fragments.

Eleven individual hyperimmune rabbit polyclonal anti-fluorescein Fab fragment preparations were resolved into heterogeneous subfractions based on differential dissociation times from a specific adsorbent. Four Fab subfractions (i.e., 0.1-, 1.0-, 10-, and 100-day elutions) that differed in affinity were characterized and classified according to the extent of the bathochromic shift in the absorption properties of antibody-bound fluorescein ligand. Absorption maxima of bound fluorescein were shifted in all cases to two distinct narrow ranges, namely, 505 to 507 nm or 518 to 520 nm relative to 491 nm for free fluorescein. There was no direct correlation between the two spectral shift populations and antibody affinity, fluorescence polarization, fluorescence quenching, or fluorescence lifetimes of bound ligand. Fluorescence emission maxima varied with the bathochromic shift range. Bound fluorescein ligand, with absorption maxima of 505 to 507 nm and 518 to 520 nm showed fluorescence emission maxima of 519 to 520 nm and 535 nm, respectively. The two spectral shift ranges differed by approximately 14 to 15 nm and/or energies of approximately 1.5 kcal mol(-1) relative to each other and to the absorption maximum for free fluorescein. Spectral effects on the antibody-bound ligand were discussed relative to solvent-water studies and the atomic structure of a high-affinity liganded anti-fluorescein active site.

Animals↗
PubMed2001

Comparison of fluorescein break-up time measurement reproducibility using standard fluorescein strips versus the Dry Eye Test (DET) method.

PURPOSE: To compare the repeatability of fluorescein break-up time (FBUT) measurements determined with either a standard fluorescein strip or the Dry Eye Test (DET) modified fluorescein strip methods. METHODS: This was a prospective, randomized contralateral study of 100 patients, in which FBUT measurements were determined with a standard FUL-GLO fluorescein strip (Akorn, Inc., Buffalo Grove, IL, U.S.A.) in one eye and a DET strip (Akorn, Inc., Buffalo Grove, IL, U.S.A.) for the contralateral eye. Three consecutive measurements were made immediately after fluorescein instillation. The second eye was evaluated 1 minute after completion of the first eye. Data from patients with FBUT values less than 20 seconds were included in the data analysis, because measurements greater than 20 seconds are not diagnostically significant. RESULTS: Seventy-five patients met enrollment and FBUT measurement criteria. For three consecutive FBUT measurements, the DET values were within 3 seconds for 72 of the 75 patients (96%). Eighty percent of patients reported no sensation with the DET strip, 20% reported mild sensation, and no patient reported moderate sensation. With FUL-GLO strips, measurements were within 3 seconds for 53 of the 75 patients (71%) (p <0.005). The standard fluorescein strip method elicited reports of no sensation from 31% of patients, mild sensation from 60%, and moderate sensation from 9% (p <0.001). CONCLUSIONS: The DET strip provides a significant reduction in sensation upon application, improved single measurement reliability, and enhanced measurement precision, compared with a conventional fluorescein strip.

Adult↗