Stable isotope ratio analysis for protecting foods with

denomination

Federica Camin

Strategies

paper traceability

(EU Reg. 178/2002)

analytical tests

robust, validated, official methods

(EU Reg, OIV, ISO/ENV, AOAC)

robust, validated, official limits or reference data

Stable Isotope Ratios of Bioelements

34

2H/1H

13C/12C

15N/14N

18O/16O

34S/32S

Stable isotope ratios methods as official standards

Year Method product Method Isotope Ratio Fraud

1987 OIV wine, must SNIF-NMR D/H sugar addition (beet, cane)

1990 EU Reg 2676/90, encl. 8 wine, must SNIF-NMR D/H sugar addition (beet, cane)

1991 AOAC 998,12 honey IRMS 13C/12C sugar addition (cane)

1993 ENV 12140, 13070 fruit juice IRMS 13C/12C sugar addition (cane)

1995 AOAC 995,17 fruit juice SNIF-NMR D/H sugar addition (beet, cane)

1996 OIV 2/96 wine, must IRMS 18O/16O addition of water/mislabelling

1997 EU Reg 2676/90, 822/97 wine, must IRMS 18O/16O addition of water/mislabelling

1997 ENV 12141 fruit juice IRMS 18O/16O addition of water/mislabelling

2000 AOAC 2000.19 mapple syrup SNIF-NMR D/H sugar addition (beet, cane)

2000 OIV 71/2000 vinegar SNIF-NMR, IRMS D/H, 13C/12C sugar addition (beet, cane)

2001 OIV 17/2001 wine, must IRMS 13C/12C sugar addition (cane)

2003 EU Reg. 2676/90, 440/03 wine, must IRMS 13C/12C sugar addition (cane)

2003 OIV MA-F-AS314-03 wine IRMS 13C/12C technogenic CO2

2004 AOAC 2004,01 fruit juice, maple syrup SNIF-NMR D/H sugar addition (beet, cane)

2006 AOAC 2006,05 vanillin SNIF-NMR D/H synthetic vanillin

2007 OIV-MA-AS312-07 wine IRMS 13C/12C addition of glycerol

2011 EU Reg 584/2011 Grana Padano DOP IRMS D/H, 13C/12C, 15N/14N, 34S/32S mislabelling

2013 EN 16466-1, 2, 3 vinegar SNIF-NMR, IRMS D/H, 13C/12C, 18O/16O water and sugar addition (beet, cane)

2013 OIV 510, 511/2013 vinegar IRMS 13C/12C, 18O/16O water and sugar addition (cane)

520 denomination for Italian wines (DOC/DOCG/PDO/IGP) Wine

OIV-MA-AS311-05 for site-specific analysis of the D/H ratio using 2H-SNIF-NMR (2H-Site-specif Natural Isotope Fractionation - Nuclear Magnetic Resonance)

OIV-MA-AS312-06 for analysis of the 13C/12C ratio (expressed as d13C‰) using Isotope Ratio Mass Spectrometry (IRMS)

OIV-MA-AS2-12 for analysis of the 18O/16O ratio (expressed as d18O‰) using Isotope Ratio Mass Spectrometry (IRMS).

Databank: EU reg 555/2008

Wine Databank EC Reg. Nº 2729/2000

Malta

4

Slovakia

15

Slovenia

20

Hungary

50 Czech Rep.

20

Cyprus

10

France

400

Germany

200

Italy

400

Spain

200 U. Kingdom

4

Austria

50

Greece

50

Luxembourg

4

Portugal

50

Wine data Maintenance of databank

Arbitration of disputes

Analysis of samples

Development and

validation of methods

Validation of data

Training

EC Reg. Nº 2120/2004

Romania

70

Bulgaria

30

EC Reg. Nº 2030/2006

Definition of limits and compliance

selection of reference data

computing 95% reference limits (t-student two tailed distribution)

c) the result is outside the limit, but, adding the measurement uncertainty, is inside the limit

d) the result and uncertainty are outside the limit

Situations a and b do not cause any consequences since the results are in compliance with the

a) b) c) d)

Upper limit

Lower limit

= result

= uncertainty

Balsamic vinegar of Modena PGI (Aceto balsamico di Modena)

obtained by mixing wine vinegar, cooked or concentrated grape must and caramel.

PGI status (Protected Geographical Indication) with EC Regulations 583/2009

6° PDO/PGI Italian product for sales (260 million € /year)

The geographical origin of vinegar and must is not specified

Modena

Stable Isotope ratio analysis in vinegar

CEN (European Commitee for Standardisation) from 01/03/2013 EN 16466-1 D/H SNIF NMR acetic acid EN 16466-2: 13C/12C IRMS acetic acid EN 16466-3: 18O/16O IRMS water

OIV from 06/2013

RISOLUZIONE OIV-OENO 510-2013: 13C/12C IRMS acetic acid

RISOLUZIONE OIV-OENO 511-2013: 18O/16O IRMS water

Dordevic et al., 2013

(D/H)1 (D/H)2 δ13C δ18O

Wines, 95% CI 98.8 / 106 124.5 / 135.5 -29.3 / -24.3 -1.3 / 8.9

Beet sugar 92.5 -27.5

Cane sugar 109.5 -12

Water (north Italy) -9

Water (south Italy) -5

Stable Isotope ratios for the autenticity of ABM

-34

-32

-30

-28

-26

-24

-22

-20

-18

-16

-14

-12

-10

90 92 94 96 98 100 102 104 106 108 110 112 114 116 118 120

(D/H)I ppm ethanol

d13C

‰ e

than

ol

beet sugar

synthesis

cane sugar

Wine/must/

vinegar

(D/H)I

(D/H)II

The “top ten” Italian PDO and PGI for sales 2012

milions €

Grana Padano 1.395

Parmigiano Reggiano 1.357

Prosciutto di Parma 992

Prosciutto di San Daniele 302

Mozzarella di Bufala Campana 288

Aceto Balsamico di Modena 260

Gorgonzola 249

Mortadella Bologna 224

Bresaola della Valtellina 215

Mela Alto Adige 170

Total (production costs) 5.453

% of total 83,8

Source: Osservatorio Ismea prodotti Dop e Igp

Random Forest Model (2011) for the traceability of Grana Padano cheese

98.5% correct classification, in cross validation

Validation of the methods

International collaborative study:

IUPAC protocol and the ISO Standards 5725/2004 and 13528/2005.

7 types of cheeses in blind duplicate

20 laboratories

13 labs answered

Sr and SR are available

Camin et al., RCM, 2015

Specification for PDO Grana Padano cheese

Isotopic ratios as official parameters for verifying the authenticity of PDO Grana Padano cheese in grated and shredded form

The variability ranges have been deposited at the Italian Ministry of Agriculture, Food and Forestry

Passate di pomodoro made in Italy

D.M. 23 settembre 2005 (GU n. 232 del 5 ottobre 2005):

‘Definizione di passata di pomodoro: la denominazione di vendita ‘Passata di Pomodoro’ è riservata al prodotto ottenuto direttamente dal pomodoro fresco, sano e maturo…, per spremitura, eventuale separazione di bucce e semi e parziale eliminazione dell’acqua di costituzione in modo che il risiduo ottico rifrattometrico risulti compreso tra 5 e 12 gradi Brix ….

Non è consentito concentrare il succo di pomodoro al di sopra dei 12 gradi brix e provvedere alla successiva diluizione per la preparazione della passata di pomodoro…

Per la determinazione dell’acqua aggiunta ai prodotti che riportano tale denominazione di vendita … si applica il metodo di analisi UNI ENV 12141 (1997) e successive modifiche = Determinazione del rapporto 18O/16O nei succhi di frutta e vegetali

Bontempo et al., Food Control 2014

d18O passate > 7,5°Bx da evaporazione

Bontempo et al., Food Control 2014

Limite: -1.4‰

Conclusion

SIRA, sometime in combination with other analytical techniques, is able to detect the authenticity of food with denomination:

DOC/DOCG/PDO wine

Aceto Balsamico di Modena PGI

Grana Padano and Parmigiano Reggiano PDO

passata ‘Made in Italy’

Other examples: Extravirgin olive oil PDO, PGI, PDO ham, organic/conventional products, …

Thank you for your kind attention!

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