Bioconjugates and antibodies for immunodetection assisted by derivatization of the mycotoxin patulin

Patent metadata

Field	Value
Publication number	US-12534447-B2
Application number	US-202117800401-A
Country	US
Kind code	B2
Filing date	Feb 17, 2021
Priority date	Feb 17, 2020
Publication date	Jan 27, 2026
Grant date	Jan 27, 2026

Abstract

The present invention relates to bioconjugates and labeled derivatives of the adducts formed by the reaction of patulin with benzene-1,2-dithiols or their salts. Said conjugates are suitable for the production of antibodies capable of recognizing the aforementioned patulin-(benzene-1,2-dithiols) adducts with high affinity and specificity. Likewise, the present invention also relates to the use of bioconjugates of patulin-(benzene-1,2-dithiols) adducts and of labeled derivatives thereof as assay antigens. Furthermore, the present invention also relates to immunochemical methods for the analysis of patulin, using the bioconjugates and labeled derivatives and antibodies of the invention, which include a previous step to quantitatively convert patulin into these adducts. This invention also provides a kit for analyzing patulin comprising a benzene-1,2-dithiol or its salts, antibodies against patulin-(benzene-1,2-dithiols) adducts, and assay antigens.

First claim

The invention claimed is:

1 . A bioconjugate of general formula (I):
[T-L-Z] m —P  (I)
where: T is selected from the group consisting of the radicals R-I and R-II wherein R 1 , R 2 , R 3 , and R 4 are independently selected from H, substituted or unsubstituted aryl groups, linear or branched alkyl groups of 1 to 6 carbon atoms, OMe, OEt, OPr, C≡N, CO 2 H, CO 2 -alkyl of 1 to 6 carbon atoms, NO 2 , NH 2 , F, Cl, Br, and I

L is a hydrocarbon chain, saturated or unsaturated, of 0 to 14 carbon atoms, where the chain is linear or branched, and said hydrocarbon chain comprises the substitution of between 0 and 7 carbon atoms by heteroatoms, which are selected from the group consisting of S, O and N,
Z is a functional group selected from:
—NH—, —NH(C═O)—, —NH(C═O)O—, —(C═O)NH—, —(C═O)O—, —O(C═O)—, —O(C═O)O—, —O(C═O)NH—, —S—,

P is a natural or synthetic peptide or polypeptide, or a protein, with a molecular weight greater than 2000 daltons,
m is a number with a value between 1 and 500.

2 . The bioconjugate of formula (I) according to claim 1 wherein L is a linear hydrocarbon chain of 0 to 7 carbon atoms and said hydrocarbon chain comprises the substitution of between 0 and 3 carbon atoms by heteroatoms selected from the group consisting of O and N.

3 . The bioconjugate of formula (I) according to claim 1 , characterized in that Z is selected from the group consisting of:
—NH—, —NH(C═O)—, —(C═O)NH—,

4 . The bioconjugate of formula (I) according to claim 1 characterized in that L is a linear hydrocarbon chain of 0 to 3 carbon atoms and said hydrocarbon chain comprises the substitution of between 0 and 2 carbon atoms by heteroatoms selected from the group consisting of O and N.

5 . The bioconjugate of formula (I) according to claim 1 characterized in that Z is —(C═O)NH—.

6 . The bioconjugate of formula (I) according to claim 1 , characterized in that P is selected from the group consisting of albumin, thyroglobulin, hemocyanin, beta-galactosidase, phosphatase, peroxidase, and oxidase.

7 . The bioconjugate of formula (I) according to claim 1 , characterized in that m is a number with a value between 1 and 50.

8 . The bioconjugate of formula (I) according to claim 1 , having the formula (Ia)

where P is selected from the group consisting of albumin or peroxidase, and m is a value selected between 1 and 50.

9 . The bioconjugate of formula (I) according to claim 1 , having the formula (Ib)

where P is selected from the group consisting of albumin or peroxidase, and m is a value selected between 1 and 50.

10 . The bioconjugate of formula (I) according to claim 1 , having the formula (Ic)

where P is selected from the group consisting of albumin or peroxidase, m is a value selected between 1 and 50, and where R 2 is CH 3 and R 3 is H, or R 2 is H and R 3 is CH 3 .

11 . The bioconjugate of formula (I) according to claim 1 , having the formula (Id)

where P is selected from the group consisting of albumin or peroxidase, and m is a value selected between 1 and 50.

12 . The bioconjugate of formula (I) according to claim 1 , having the formula (Ie)

where P is selected from the group consisting of albumin or peroxidase, and m is a value selected between 1 and 50.

13 . The bioconjugate of formula (I) according to claim 1 , having the formula (If)

where P is selected from the group consisting of albumin or peroxidase, and m is a value selected between 1 and 50, and where R 2 is CH 3 and R 3 is H, or R 2 is H and R 3 is CH 3 .

14 . The bioconjugate of formula (I) according to claim 1 , having the formula (Ig)

where P is selected from the group consisting of albumin or peroxidase, and m is a value selected between 1 and 50.

15 . The bioconjugate of formula (I) according to claim 1 , having the formula (Ih)

where P is selected from the group consisting of albumin or peroxidase, and m is a value selected between 1 and 50.

16 . The bioconjugate of formula (I) according to claim 1 , having the formula (Ii)

where P is selected from the group consisting of albumin or peroxidase, m is a value selected between 1 and 50, and where R 2 is CH 3 and R 3 is H, or R 2 is H and R 3 is CH 3 .

17 . A labeled derivative of formula (II):
[T-L-Z] n -Q  (II)
wherein, T and L have been defined according to claim 1 ; Z has been defined in claim 1 ; Q is a detectable label and n is a number with a value between 1 and 500.

18 . The labeled derivative of formula (II) according to claim 17 , where Q is selected from the group consisting of enzymes, biotin, fluorescein or any one of its derivatives, a cyanine fluorophore, a rhodamine fluorophore, a coumarin fluorophore, a ruthenium bipyryl, luciferin or any one of its derivatives, an acridinium ester, quantum nanoparticles (quantum dots), and micro- or nanoparticles of colloidal gold, carbon, or latex.

19 . The labeled derivative of formula (II) according to claim 17 characterized in that Z is selected from the group consisting of
—NH—, —NH(C═O)—, —(C═O)NH—,

20 . The bioconjugate of formula (II) according to claim 17 , characterized in that m is a number with a value between 1 and 50.

21 . The labeled derivative of formula (II) according to claim 17 , having the formula (IIa)

where Q is selected from the group consisting of peroxidase, biotin, fluorescein, or gold nanoparticles, and n is a value selected between 1 and 10.

22 . The labeled derivative of formula (II) according to claim 17 , having the formula (IIb)

where Q is selected from the group consisting of peroxidase, biotin, fluorescein, or gold nanoparticles, and n is a value selected between 1 and 10.

23 . The labeled derivative of formula (II) according to claim 17 , having the formula (IIc)

Assignees

• Consejo Superior Investigacion
• Univ Valencia

Inventors

• Abad Fuentes Antonio
• Mercader Badia Josep Vicent
• Duncan Hadyn
• Abad Somovilla Antonio
• Agulló Blanes Consuelo

Classifications

• G01N33/025

  Fruits or vegetables · CPC title

• C07K16/44

  against material not provided for elsewhere {, e.g. haptens, metals, DNA, RNA, amino acids} · CPC title

• C07D339/08 Primary

  Six-membered rings · CPC title

• C12Q1/34

  involving hydrolase · CPC title

• C12Q1/26

  involving oxidoreductase · CPC title