Apparatus for the quantitative determination of saccharides

• US 4,247,647 A
• Filed: 03/14/1978
• Issued: 01/27/1981
• Est. Priority Date: 03/14/1978
• Status: Expired due to Term

• Current Assignee: Technicon Instruments Corporation (Bayer AG)
• Original Assignee: Technicon Instruments Corporation (Bayer AG)
• Inventors: Keyes, Melvin H., Barabino, Raymond C.
• Primary Examiner(s): Wiseman, Thomas G.
• Application Number:
  US05/886,503
• Time in Patent Office: 1,050 Days
• Field of Search: 195/103.5 R, 195/103.5 C, 195/127, 195/115, 195/31 R, 204/195 B, 204/1 T, 435/14, 435/18, 435/22, 435/27, 435/28, 435/288, 435/176, 435/291
• US Class Current: 435/287.9
• CPC Class Codes: C12Q 1/54 involving glucose or galactose

Abstract

A method for the quantitative determination of sugars in a fluid sample. The sugar sample, which may contain free glucose, is passed through a scavenger stage where substantially all of the glucose is removed, so that it does not interfere with the determination. The glucose-free sample is then passed through an immobilized glucose generating stage to produce glucose in molar proportion to the total initial concentration of the sugars present. The generated glucose is passed through an immobilized hydrogen peroxide generating stage to form gluconic acid and hydrogen peroxide. The generated hydrogen peroxide is detected in suitable detection means, typically a polarographic cell. Total free glucose is determined by measuring the total sugars in the sample by bypassing the scavenger stage, determining total sugar, then determining the glucose in the sample without bypassing the scavenger and measuring free glucose as the difference.

Claims

1. An apparatus for the quantitative determination of sugars contained in a fluid sample containing free glucose as a contaminant which comprises, in combination, a reservoir containing a buffer diluent solution and means for flowing said buffer solution as a stream, means for injecting said fluid sample into said flowing stream of buffer solution, first means for passing said flowing stream and including a scavenger enzymatic reagent comprising glucose oxidase and catalase immobilized upon a first solid support for concurrently reacting said free glucose and also hydrogen peroxide resulting from the reaction of said free glucose, the respective quantities of said glucose oxidase and said catalase immobilized on said first solid support being at least sufficient to substantially completely react said free glucose and said resulting hydrogen peroxide, second means for receiving said flowing stream directly from said first means and comprising a glucose generating enzymatic reagent immobilized on a second solid support for reacting with sugars remaining in said flowing stream to produce glucose reaction products, third means for receiving said flowing stream from said second means and comprising a hydrogen peroxide generating enzymatic reagent to react with certain ones of said glucose reaction products to generate hydrogen peroxide, and polarographic detection means for measuring said hydrogen peroxide generated in said flowing stream.

2. The apparatus of claim 1 wherein said glucose generating emzymatic reagent is glucoamylase.

3. The apparatus of claim 1 wherein said glucose generating enzymatic reagent is maltase.

4. The apparatus of claim 1 wherein said first solid support is alumina.

5. The apparatus of claim 1 wherein said second solid support is alumina.

6. The apparatus of claim 1 wherein third solid support is alumina.