Assay for the detection of beet sugar adulteration of food products

• US 5,128,243 A
• Filed: 06/28/1990
• Issued: 07/07/1992
• Est. Priority Date: 06/28/1990
• Status: Expired due to Fees

• Current Assignee: University of South Florida (State University System of Florida)
• Original Assignee: University of South Florida (State University System of Florida)
• Inventors: Potter, Robert L., Mansell, Richard L. L.
• Primary Examiner(s): Kepplinger, Esther L.
• Assistant Examiner(s): Wortman, Donna C.
• Application Number:
  US07/544,890
• Time in Patent Office: 740 Days
• Field of Search: 435/7.92, 435/7.9, 530/587, 436/540, 436/825, 436/518, 426/231
• US Class Current: 435/7.92
• CPC Class Codes: G01N 33/5308 for analytes not provided for elsewhere, e.g. nucleic acids, uric acid, worms, mites
• CPC Class Codes: G01N 33/68 involving proteins, peptides or amino acids involving lipoproteins G01N33/92
• CPC Class Codes: Y10S 436/815 Test for named compound or class of compounds

Abstract

An immunoassay for the detection of beet sugar adulteration of sugar-containing materials such as fruit juices and fruit juice concentrates utilizes antibodies to proteinaceous components of beet sugar. These proteinaceous components have been found in all samples of beet sugar, tested, regardless of source or degree of refinement; these proteinaceous components have also been found in both crystal and invert sugars.

Claims

1. An antibody raised against a proteinaceous component of beet sugar, said proteinaceous component of beet sugar comprising at least one proteinaceous material, found in beet sugar, selected from the group consisting of those proteinaceous materials, found in beet sugar, having a molecular weight as determined by carrying our electrophoresis on said proteinaceous material and comparing its mobility with that of marker proteins of known molecular weight, of about 61,000 daltons, about 45,000 daltons, about 39,000 daltons, about 36,000 daltons, about 30,000 daltons, about 27,000 daltons, about 21,000 daltons, and about 16,000 daltons.

2. The antibody according to claim 1, wherein said proteinaceous component of beet sugar comprises more than about 50% by weight of amino acids linked by acid labile bonds.

3. The antibody according to claim 2, wherein said proteinaceous component is rich in alanine, serine, glycine and proline.

4. The antibody according to claim 2, wherein said proteinaceous component of beet sugar comprises a proteinaceous material, found in beet sugar, having a molecular weight of about 61,000 daltons.

5. The antibody according to claim 2, wherein said proteinaceous component of beet sugar comprises a proteinaceous material, found in beet sugar, having a molecular weight of about 45,000 daltons.

6. The antibody according to claim 2, wherein said proteinaceous component of beet sugar comprises a proteinaceous material, found in beet sugar, having a molecular weight of about 39,000 daltons.

7. The antibody according to claim 2, wherein said proteinaceous component of beet sugar comprises a proteinaceous material, found in beet sugar, having a molecular weight of about 36,000 daltons.

8. The antibody according to claim 2, wherein said proteinaceous component of beet sugar comprises a proteinaceous material, found in beet sugar, having a molecular weight of about 30,000 daltons.

9. The antibody according to claim 2, wherein said proteinaceous component of beet sugar comprises a proteinaceous material, found in beet sugar, having a molecular weight of about 27,000 daltons.

10. The antibody according to claim 2, wherein said proteinaceous component of beet sugar comprises a proteinaceous material, found in beet sugar, having a molecular weight of about 21,000 daltons.

11. The antibody according to claim 2, wherein said proteinaceous component of beet sugar comprises a proteinaceous material, found in beet sugar, having a molecular weight of about 16,000 daltons.

12. The antibody according to claim 2, wherein said proteinaceous component of beet sugar comprises a mixture of proteinaceous materials, found in beet sugar, having molecular weights of about 61,000 daltons, about 45,000 daltons, about 39,000 daltons, about 36,000 daltons, about 30,000 daltons, about 27,000 daltons, about 21,000 daltons and about 16,000 daltons.

13. The antibody according to claim 1, wherein said beet sugar is a crystalline sugar.

14. The antibody according to claim 1, wherein said beet sugar is at least partially inverted.

15. The antibody according to claim 14, wherein said at least partially inverted beet sugar has a degree of inversion of about 50 to 90%.

16. A method of detecting the presence of beet sugar in a sugar-containing material, said method comprising:

treating a test sample of said sugar-containing material to selectively recover proteinaceous materials therefrom to form a protein test sample;

contacting said protein test sample with antibody raised against a proteinaceous component of beet sugar; and

detecting the formation of complexes of said proteinaceous component and antibody.

17. The method of claim 16, wherein said treatment of said test sample to selectively recover proteinaceous materials therefrom comprises contacting said test sample with a weak anion exchange resin and then eluting the material absorbed on said weak anion exchange resin to form an eluant.

18. The method of claim 17, further comprising ultrafiltration of said eluant to produce a retentate.

19. The method of claim 18, wherein said ultrafiltration produces a retentate containing materials having a molecular weight of greater than about 10,000 daltons.

20. The method of claim 18, further comprising concentration of said retentate by centrifugation.

21. The method of claim 16, wherein said proteinaceous component of beet sugar comprises at least one proteinaceous material, found in beet sugar, selected from the group consisting of those proteinaceous materials, found in beet sugar, having a molecular weight, as determined by carrying out electrophoresis on said proteinaceous material and comparing its movement with that of marker proteins of known molecular weight, of about 61,000 daltons, about 45,000 daltons, about 39,000 daltons, about 36,000 daltons, about 30,000 daltons, about 27,000 daltons, about 21,000 daltons and about 16,000 daltons.

22. The method of claim 21, wherein said proteinaceous component of beet sugar comprises more than about 50% by weight of amino acids linked by acid labile bonds.

23. The method of claim 22, wherein said proteinaceous component is rich in alanine, serine, glycine and proline.

24. The method of claim 21, wherein said proteinaceous component of beet sugar comprises a mixture of proteinaceous materials, found in beet sugar, having molecular weights of about 61,000 daltons, about 45,000 daltons, about 39,000 daltons, about 36,000 daltons, about 30,000 daltons, about 27,000 daltons, about 21,000 daltons and about 16,000 daltons.

25. The method of claim 16, wherein said detection of said complexes is effected by enzyme-linked immunosorbent assay.

26. The method of claim 16, further comprising removing at least a major portion of the sugar present in a sample of said sugar-containing material to form said test sample.

27. The method of claim 26, wherein at least 80% of the sugar present in said sample of sugar-containing material is removed from said test sample.

28. The method of claim 27, wherein at least 85% of the sugar present in said sample of sugar-containing material is removed to form said test sample.

29. The method of claim 16, wherein said removal of said sample of said sugar-containing material is effected by dialysis.

30. The method of claim 29, wherein said dialysis removes materials having a molecular weight of not more than about 6,000 daltons.

31. The method of claim 26, further comprising forming an aqueous sugar solution of said sugar-containing material prior to removal of said major portion of the sugar therefrom.

32. The method of claim 31, wherein said aqueous sugar solution is filtered to remove particulate material therefrom prior to removal of said major portion of the sugar therefrom.

33. The method of claim 32, wherein said filtration removes particulate material having an average diameter greater than about 0.2 micron.

34. The method of claim 16, wherein said sugar-containing material is a fruit juice or fruit juice concentrate.

35. The method of claim 34, further comprising filtering said fruit juice or fruit juice concentrate prior to said treatment to selectively recover proteinaceous materials therefrom.

36. The method of claim 35, wherein said filtration removes particulate material having an average diameter greater than about 0.2 micron.

37. The method of claim 34, wherein said sugar-containing material is apple juice or apply juice concentrate.

38. The method of claim 34, wherein said sugar-containing material is orange juice or orange juice concentrate.

39. Rabbit IgG specifically binds to a proteinaceous component of beet sugar, said proteinaceous component of beet sugar comprising at least one proteinaceous material, found in beet sugar, selected from the group consisting of those proteinaceous materials, found in beet sugar, having a molecular weight, as determined by carrying out electrophoresis on said proteinaceous material and comparing its mobility with that of marker proteins of known molecular weight, of about 61,000 daltons, about 45,000 daltons, about 39,000 daltons, about 36,000 daltons, about 30,000 daltons, about 27,000 daltons, about 21,000 daltons and about 16,000 daltons.

40. The rabbit IgG according to claim 39, wherein said proteinaceous component of beet sugar comprises more than about 50% by weight of amino acids linked by acid labile bonds.

41. The rabbit IgG according to claim 40, wherein said proteinaceous component is rich in alanine, serine, glycine and proline.

42. A test kit for the detection of beet sugar in a sugar-containing material comprising:

a predetermined quantity of a first mammalian IgG specifically binds to a proteinaceous component of beet sugar, said proteinaceous component of beet sugar comprising at least one proteinaceous material, found in beet sugar, selected from the group consisting of those proteinaceous materials, found in beet sugar, having a molecular weight, as determined by carrying out electrophoresis on said proteinaceous material and comparing its mobility with that of marker proteins of known molecular weight, of about 61,000 daltons, about 45,000 daltons, about 39,000 daltons, about 36,000 daltons, about 30,000 daltons, about 27,000 daltons, about 21,000 daltons and about 16,000 daltons; and

a predetermined quantity of a labelled second mammalian anti-first mammal IgG.

43. The test kit according to claim 42, wherein said mammalian IgG is rabbit IgG and said labelled second mammalian anti-first mammal IgG is labelled goat anti-rabbit IgG.

44. The test kit according to claim 43, wherein said labelled goat anti-rabbit IgG is goat anti-rabbit IgG-alkaline phosphatase conjugate.