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Uremic Toxins pp 205-213 | Cite as

Biochemical Elucidation and HPLC Fractionation of Fluorescent Peptides in Patients with Chronic Renal Failure

  • Robert H. Williams
  • Alvin Dubin
  • Mashouf Shaykh
  • Theodore Musiala
  • Sarosh Ahmed
  • Nadine Bazilinski
  • George Dunea
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 223)

Abstract

We evaluated the biochemical characteristics of endogenous fluorescent substances. Ex 380 nm/Em 440 nm and Ex 400 nm/Em 460 nm, present in sera of patients with chronic renal failure (Clin. Chem. 31:1988, 1985). Sera from 23 patients with chronic renal failure (CRF) and from 10 normal subjects were filtered through ultrafiltration membranes (cutoff limit of 500 Da). Fluorescence intensity of the aforementioned substances was significantly elevated as compared to normals (p < 0.001). Fluorescence characteristics of these substances remained unaltered after ultrafiltration and treatment with beta-glucuronidase. Extraction of these fluorescent compounds with organic solvents (dichloromethane, ethyl acetate, chloroform:methanol) could not be achieved after ultrafiltrates were subjected to 6N hydrochloric acid (HCl) hydrolysis. In addition, treatment with 6N HCl enhanced fluorescence intensity without altering fluorescence excitation/emission maxima. Removal of fluorescence could be accomplished in toto by adsorption onto activated charcoal with subsequent recovery from charcoal by treatment with sodium hydroxide, pH 12 (Ex 380 nm: 51.1%, Ex 400 nm: 91.8%). Analysis of alkali-treated specimens by high performance liquid chromatography demonstrated that peptides associated with these fluorescent substances were denatured, although fluorescence at these previously described excitation/emission maxima persisted. Our studies indicate that the unique fluorescence observed in the sera of patients with CRF is not an intrinsic characteristic of a specific peptide or its amino acids, but rather an inherent property of fluorescent molecules which may bind to these peptides.

Keywords

High Performance Liquid Chromatography High Performance Liquid Chromatography Chronic Renal Failure Activate Charcoal Ultrafiltration Membrane 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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Copyright information

© Plenum Press, New York 1987

Authors and Affiliations

  • Robert H. Williams
    • 1
  • Alvin Dubin
    • 1
  • Mashouf Shaykh
    • 2
  • Theodore Musiala
    • 1
  • Sarosh Ahmed
    • 2
  • Nadine Bazilinski
    • 2
    • 3
  • George Dunea
    • 2
    • 3
  1. 1.Department of BiochemistryRush University and Hektoen Institute for Medical ResearchChicagoUSA
  2. 2.Division of NephrologyCook County Hospital and Hektoen Institute for Medical ResearchChicagoUSA
  3. 3.College of MedicineUniversity of Illinois at ChicagoChicagoUSA

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