US 12,656,257 B2Granted

Method and device for the rapid quantification of aflatoxin M1 (AFM1) in milk

AssigneeUniversita' Degli Studi Di Padova

InventorsTeofilo Vamerali, Giuseppe Barion, Stefano Tiozzo Netti

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Abstract

A device and its use for the direct quantification of Aflatoxin M1 (AFM1) in a sample of milk, the use includes a) providing a sample of milk; b) irradiating the sample of milk of step a) with a first electromagnetic radiation which includes at least one wavelength in a range from 340 to 400 nm; c) detecting a second electromagnetic radiation, emitted by the sample of milk of step b), in a range from 410 to 860 nm; d) selecting at least 9 wavelengths in a range from 410 to 860 nm of the second electromagnetic radiation detected in step c); and e) quantifying the Aflatoxin M1 (AFM1) in the sample of milk, on the basis of an intensity of the radiation emitted by the wavelengths selected in step d).

A method for the quantification of Aflatoxin M1 (AFM1) in milk, comprising the steps of:

a) providing a sample of milk;
a′) irradiating the sample of milk with an electromagnetic radiation comprising at least one wavelength in a range from 200 to 280 nm, suitable to photo-derivatize Aflatoxin M1 (AFM1) so as to obtain photo-derivatized Aflatoxin M1 (AFM2a);
a″) adding trifluoroacetic acid, or at least one source of bromine selected from the group consisting of alkali bromine salts and pyridinium bromide-perbromide (PBPB), or iodine, and mixtures thereof, to the sample of milk obtained in step a′) so as to chemically derivatize Aflatoxin M1 and increase fluorescence emission;
b) irradiating the sample of milk obtained in the step a″) with an electromagnetic radiation comprising at least one wavelength in a range from 340 to 400 nm;
c) detecting a second electromagnetic radiation emitted by the sample of milk in response to the irradiation of step b), said second electromagnetic radiation being detected in a wavelength range from 410 to 860 nm;
d) selecting fluorescence intensity values detected at the following wavelengths of the second electromagnetic radiation:
410 nm, 435 nm, 560 nm, 585 nm, 645 nm, 705 nm, 760 nm, 810 nm, and 860 nm;
e) calculating standardized fluorescence intensity values Is410 nm, Is435 nm, Is560 nm, Is585 nm, Is645 nm, Is705 nm, Is760 nm, Is810 nm and Is860 nm according to the following equations (III1-III9):
Is
⁢
410
⁢
nm
=
(
I
⁢
410
⁢
nm
-
52.
7
⁢
37
)
/
2.468
(
III1
)
Is
⁢
435
⁢
nm
=
(
I
⁢
435
⁢
nm
-
52.503
)
/
2.141
(
III2
)
Is
⁢
560
⁢
nm
=
(
I
⁢
560
⁢
nm
-
2
⁢
4
.
2
⁢
67
)
/
0.928
(
III3
)
Is
⁢
585
⁢
nm
=
(
I
⁢
585
⁢
nm
-
14.
8
⁢
00
)
/
0.563
(
III4
)
Is
⁢
645
⁢
nm
=
(
I
⁢
645
⁢
nm
-
13.
4
⁢
37
)
/
0.523
(
III5
)
Is
⁢
705
⁢
nm
=
(
I
⁢
705
⁢
nm
-
1
⁢
8
.
5
⁢
40
)
/
0.759
(
III6
)
Is
⁢
760
⁢
nm
=
(
I
⁢
760
⁢
nm
-
20.033
)
/
2.972
(
III7
)
Is
⁢
810
⁢
nm
=
(
I
⁢
810
⁢
nm
-
24.233
)
/
4.183
(
III8
)
Is
⁢
860
⁢
nm
=
(
I
⁢
860
⁢
nm
-
12.387
)
/
1.574
(
III9
)
wherein I410 nm, I435 nm, I560 nm, I585 nm, I645 nm, I705 nm, I760 nm, I810 nm and I860 nm are fluorescence intensity values detected in step c);
f) calculating a parameter F1 according to the following equation (II):
and
F1=2.533× Is410 nm−0.132×Is435 nm+5.527× Is560 nm+1.911× Is585 nm+7.225× Is645 nm−1.082× Is705 nm−8.849× Is760 nm−11.337× Is810 nm+1.874×Is860 nm (II);
g) quantifying a concentration of Aflatoxin M1 (AFM1) in the sample of milk by applying the following equation (I):
AFM
⁢
1
⁢
(
ppt
)
=
0.0092
×
(
F
⁢
1
)
2
+
1.0694
×
(
F
⁢
1
)
+
31.635
(
I
)
wherein AFM1 (ppt) is the concentration of Aflatoxin M1 expressed in ng kg−1.

241 more elements

Facts

Application

17/906,911Filed 23 Mar 2021

Priority

IT 10202000000637026 Mar 2020

Published

US 2023/0126620 A127 Apr 2023

Granted

16 Jun 2026

Term ends

Not estimatedThe source records don’t support an estimate.

Classification5 codes

G01N 21/6486
Fluorescence; Phosphorescence: measuring fluorescence of biological material, e.g. DNA, RNA, cells
G01N 33/04
Food: dairy products
G01N 21/645
Fluorescence; Phosphorescence: specially adapted constructive features of fluorimeters
G01N 2021/6421
Spectrofluorimetric devices: measuring at two or more wavelengths

Abstract

Claims17