Whisky analysis by electrospray ionization-Fourier transform mass spectrometry

Food Research International 51 (2013) 98–106

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Whisky analysis by electrospray ionization-Fourier transform mass spectrometry Jerusa S. Garcia a, b,⁎, Boniek G. Vaz a, Yuri E. Corilo a, Christina F. Ramires a, Sérgio A. Saraiva a, Gustavo B. Sanvido a, Eduardo M. Schmidt a, Denison R.J. Maia c, Ricardo G. Cosso c, Jorge J. Zacca d, Marcos Nogueira Eberlin a a

University of Campinas, UNICAMP, Institute of Chemistry, ThoMSon Mass Spectrometry Laboratory, 13084-971, Campinas, SP, Brazil Federal University of Alfenas, UNIFAL, Institute of Chemistry, 37130-000, Alfenas, MG, Brazil Brazilian Federal Police, Ministério da Justiça, Superintendência Regional de São Paulo, 05038-090, São Paulo, SP, Brazil d Brazilian Federal Police, Ministry of Justice, National Institute of Criminalistics — INC, 70390-145, Brasilia, DF, Brazil b c

a r t i c l e

i n f o

Article history: Received 15 October 2012 Accepted 15 November 2012 Keywords: Whisky MS fingerprinting Authenticity High resolution mass spectrometry

a b s t r a c t Electrospray ionization (ESI) coupled to ultra-high resolution and accuracy Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) was employed for the direct analysis of whisky samples. No pre-separation or extraction steps were employed and, owing to the gentleness of ESI, characteristic profiles of polar constituents of authentic whiskies from five different brands were obtained, which greatly contrast to those of counterfeit samples. Owing to the accuracy of FT-ICR MS mass measurements, elemental composition of the main ions detected were also securely determined, and related to a series of carboxylic acids, phenols, saccharides, fat acids and sulfur or nitrogen constituents of whisky. The accurate mass measurements and the number of detected ions, together with the direct, no sample preparation procedure employed, make FT-ICR MS a highly reliable approach to fingerprint whisky and to control its quality, and to screen for aging, counterfeiting and adulteration at the molecular level. © 2012 Elsevier Ltd. All rights reserved.

1. Introduction Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) is a powerful technique for complex mixture analysis due to its unsurpassed ability to determine chemical formulas with ultra high resolution and accuracy (better than 1 ppm) (Marshall et al., 2007). Due to its unique characteristics, FT-ICR MS is a powerful tool for resolving the elemental composition of large molecules and can be used in distinct applications. The use of the gentle ESI technique or ambient ionization techniques (Corilo et al., 2010) also permit the detection of constituents in intact molecular forms, and when FT-ICR MS analysis is employed, direct characterization of very complex mixtures such as of crude oils can be directly performed without pre-separation methods. For crude oils, ESI FT-ICR MS have allowed unambiguous assignments of polar heteroatom-containing organic components of crude oils with 20,000 or more distinct elemental compositions (Rodgers, Schaub, & Marshall, 2005) and for wine around 30 compounds were identified employing negative-ion ESI FT-ICR analysis (Cooper & Marshall, 2001). Herein, we describe an investigation aimed at testing the use of ESI FT-ICR MS in the analysis of whisky samples.

⁎ Corresponding author at: ThoMSon Mass Spectrometry Laboratory 3084-971, Campinas, São Paulo, Brazil. Tel./fax: +55 19 3521 3073. E-mail address: [email protected] (J.S. Garcia). 0963-9969/$ – see front matter © 2012 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.foodres.2012.11.027

Scotch whisky is a prime spirit drink and a leading category of choice between spirit consumers (Gordon, 2003). The Scotch whisky in UK is on the range of several billions of dollars (Rhodes, Heaton, Goodall, & Brereton, 2009) and whisky is one of the top export products of Scotland. Due to its large commercialization and relatively high prices, Scotch whisky counterfeiting and/or adulteration is quite common worldwide. Whisky contains a great variety of constituents from different chemical classes such as alcohols, ethyl and isoamyl esters, acetates, ketones, fatty acids, monoterpenes, and phenols. This variable chemical composition is mainly influenced by the cereals used in fermentation and by distillation, maturation and blending regimes. Some of these compounds originate from raw materials and during the production steps (mashing, fermentation and distillation), while others are related to the spirit maturation in oak casks. Whisky constituents can be present in a wide range of concentrations, with contrasting volatilities and polarities, being common in distinct whisky brands but differing considerably in relative quantities (Câmara et al., 2007). Due to this complex chemical composition, whisky authenticity analysis normally requires sample extraction and pre-separation procedures. The most common strategy for establishing whisky brand authenticity is to determine the profile of volatile organic compounds (VOC congeners) such as acetaldehyde, methanol, ethyl acetate, propanol, isobutanol, diethyl acetal and the amyl alcohols: 2- and 3-methyl butanol (Aylott, Clyne, Fox, & Walker, 1994). Usually, VOC profiles of whisky are obtained via gas chromatography (GC) analysis (MacKenzie & Aylott, 2004; Nascimento, Cardoso, & Franco, 2008;

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whisky “brands” were also analyzed. The counterfeit street samples were provided by the Brazilian Federal Police.

Rhodes et al., 2009). However, spirit adulteration is becoming more sophisticated and sometimes counterfeit samples may display similar VOC profiles. GC coupled to isotope ratio mass spectrometry (GC/IRMS) has also been used to characterize whisky and to screen for adulteration (Parker, Kelly, Sharman, & Howie, 1998). Direct mass spectrometry (MS) analysis has also been employed to characterize spirits and screen for adulteration and counterfeiting. We, for instance, have employed direct “unit-resolution” MS analysis with electrospray ionization (ESI-MS) for quality control of spirits, including the Brazilian cachaça (de Souza, Augusti, et al., 2007; de Souza, Siebald, et al., 2007; de Souza et al., 2009), wine (Catharino et al., 2006), beer (Araujo et al., 2005) and whisky (Moller, Catharino, & Eberlin, 2005).

2.2. ESI FT-ICR MS analysis Samples were analyzed using a hybrid 9-T Fourier transform ion cyclotron resonance mass spectrometer (LTQ FT; Thermo Scientific, Bremen, Germany) equipped with a chip-based direct infusion nanoelectrospray ionization source (Triversa; Advion Biosciences, Ithaca, NY, USA). Nanoelectrospray conditions comprised a flow rate of 200 nL/min, backing pressure of ca. 0.3 psi, and electrospray voltages of 1.5 to 2.0 kV during 120 s, controlled by ChipSoft software (version 8.1.0, Advion Biosciences, Ithaca, NY, USA). Mass resolution was fixed at 200,000 at m/z 400. Data were obtained as transient files (scans recorded in the time domain). All the samples were evaluated in negative ESI(−) and positive ESI(+) ion modes and spectra were acquired in the m/z 100–800 range. For ESI(+), the samples were analyzed directly (without any sample treatment or dilution); for ESI(−), samples were diluted 1:1 (v/v) using a solution containing methanol (Mallinckrodt Baker, Phillipsburg, NJ, USA) and 0.1% (v/v) NH4OH (Mallinckrodt Baker, Phillipsburg, NJ, USA). The spectra were processed by the Xcalibur Analysis software package (version 2.0, Service Release 2, Thermo Electron Corporation).

2. Experimental details 2.1. Whisky samples A total of 80 samples, divided into two groups (50 authentic and 30 counterfeit) were used. The authentic whiskies include Scotch whiskies from different brands: Johnnie Walker Red Label (n= 10), Johnnie Walker Black Label (n= 12), White Horse (n= 12), Buchanan's (n= 6) and J&B (n= 10). Counterfeit samples (n=30) from different 383.12

365.11 527.16 409.16 545.17

425.14

509.15 589.23

443.17 467.10

350

100

(A)

50 0 100

(B)

(D)

203.05

50 0 100

157.08 203.05 231.08

365.11

(E)

650

527.16 509.15 545.17

409.16

2 7 1 .0 8 347.09 311.07

623.23

689.21

737.52

527.16 509.15 545.17

6 2 3 .2 3 689.21

737.50

425.14 409.16 157.08 203.05 157.08 203.05 231.08

365.11 409.16 347.09 301.14

527.16

589.23 639.20

737.47 785.28

527.16 589.23 617.26

425.14

689.21 737.46

383.12

157.08 203.05 231.08

443.17

383.12

288.29 365.11 50

550

383.12

271.08 0 100

409.16

347.09 301.14

0 100 50

450

689.21 671.20

383.12

231.08

50

Relative Abundance

(C)

157.08

623.23

409.16

347.09

527.16 509.15 545.17

623.23 639.20

737.46 785.28

0 200

300

400

500

600

700

800

m/z Fig. 1. Representative ESI(+) FT-ICR MS of five distinct Scotch whisky brands: (A) Johnnie Walker Red Label, (B) Johnnie Walker Black Label, (C) J&B, (D) White Horse and (E) Buchanan's.

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ESI(−) FT-ICR MS of the same samples (ten spectra are shown in both figures). Both spectra demonstrate the complexity of the polar chemical profiles of whisky provided by the technique, with more than 500 ions observed with more than 1% of relative abundance. All authentic samples share many common ions which is beneficial for establishing a common chemical signature of Scotch whisky in terms of polar components. Another typical characteristic common to all the authentic whisky samples is a “grass ion” with a somewhat Gaussian distribution around m/z 300–700 for ESI(+) and m/z 250–600 for ESI(−), as shown by the insets in Figs. 1 and 2. But whisky samples from different brands also display characteristic features in terms of relative abundances and unique ions that permit differentiation between samples, using both ESI(+) and ESI(−), more particularly via ESI(−), as clearly demonstrated by the PCA plots of Fig. 3. Three principal components were necessary to describe more than 58.0% of the data variance in the ESI(+) results (PC1 = 40.4%, PC2 = 10.5% and PC3 = 7.1%). In the positive ion mode the different whisky brands are characterized by certain ions: m/z 269, 323 and 503 for Black Label; m/z 323, 431, 458 and 521 for Red Label; 239, 351 and 359 for Buchanan's; 172, 343, 399 and 519 for J&B; and, finally, m/z 255, 341, 421, 443 for White Horse. The total variability explained by three components in ESI(−) was 30.3% (PC1 = 14.6%, PC2 = 8.4% and PC3 = 7.8%). In the negative ion mode the characteristic ions are: m/z 359 (Black Label); m/z 503

The elemental compositions were generated using the Quali Browser software (version 2.0, Thermo Electron Corporation). Elemental composition was acceptable only when the average differences between calculated and experimental masses were less than 1.0 ppm. Molecular formula search was done using 12C, 1H, 16O, 14N, 23Na and 32S isotopologue ions. 2.3. Multivariate analysis To classify the whisky samples, principal component analysis (PCA) was performed using the Pirouette software (version 3.11, Infometrix Inc., Woodinville, WA, USA). The ESI-MS experimental data were compiled to generate a final matrix containing samples and variables (m/z values with their respective relative intensities). To construct the matrix, no pre-processing method was performed and only the 50 more abundant ions were considered. 3. Results and discussion 3.1. Profiles in positive and negative ion modes Fig. 1 exhibits the ESI(+) FT-ICR MS of a representative authentic sample from the five Scotch whisky brands investigated (one spectrum from each brand) whereas Fig. 2 shows, for comparison, the 341.11 359.12 301.00

239.08

503.16

255.23

517.32

379.23

323.10

485.15

401.13 431.14

250

50

(D)

Relative Abundance

(C)

50

341.11 255.23 323.10

143.11

0 100

(B)

550

401.13

503.16 485.15 521.17

575.18 665.22

737.52 775.21

575.18 665.22

737.45 777.23

603.01 665.22

737.47 783.77

575.18 665.22

737.45 793.22

171.14 359.12 143.11

0 100

239.08 301.00

379.23

503.16 485.15 521.17

171.14 199.17

50

143.11 255.23 301.00 341.11 419.17

0 100 50

143.11

0 100 50

503.16

171.14 199.17

255.23

(E)

450

171.14 199.17

100

(A)

350

341.11 323.10 359.12

431.14

503.16 521.17

171.14 239.08 301.00 341.11 379.23

143.11

503.16

0 100

200

300

400

500

545.17 600

679.37 737.45 777.23 700

800

m/z Fig. 2. Representative ESI(−) FT-ICR MS of five distinct Scotch whisky brands: (A) Johnnie Walker Red Label, (B) Johnnie Walker Black Label, (C) J&B, (D) White Horse and (E) Buchanan's.

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3.2. Elemental composition and formula

(A) Buchanan’s

8 6

White Horse

4

Red Label

PC3

2 0

Black Label

J&B

-2

10 5

-4

0 10

8

-5 6

4

2

0

PC2

PC 1

-6

-10 -2 -4

-6

-15

-8

(B) 10

Black Label

Buchanan’s

5

Red Label J&B

PC3

0

White Horse f

-5 -10

15 10

-15

10

5

PC2

0

-5 -10 -5

-10

PC 1

5 0

-20

-15

Fig. 3. PCA from the five authentic Scotch whisky samples for the (A) ESI(−) and (B) ESI(+) FT-ICR MS data.

(Red Label); m/z 177, 197, 239 and 517 (Buchanan's); m/z 200 and 347 (J&B) and m/z 251 (White Horse). Some of these ions are listed in Table 1 (m/z 239 and 503) and Scheme 1 (m/z 197 and 359).

Table 1 Elemental compositions assigned to ions in ESI(−) FT-ICR MS data (20 most abundant ions). m/z

Error (ppm)

RDBa

Elemental composition

143.10784 161.04568 171.13918 179.05625 199.17052 221.06689 227.20189 239.07750 255.23324 283.26457 323.09883 341.10943 351.20294 359.12001 431.14141 467.14151 485.15215 503.16264 521.17323

0.64 0.8 0.75 0.79 0.85 0.97 1.04 0.35 1.12 1.12 0.88 0.94 0.92 0.94 −0.22 0.01 0.17 0.02 0.07

1.5 2.5 1.5 1.5 1.5 2.5 1.5 10 1.5 1.5 12 11 10 10 1.5 4.5 3.5 2.5 1.5

C8H16O2 C6H10O5 C10H20O2 C6H12O6 C12H24O2 C8H14O7 C14H28O2 C15H14NS C16H32O2 C18H36O2 C19H18O2NS C12H22O11 C23H30NS C19H22O4NS C16H32O9S2 C19H32O9S2 C19H34O10S2 C19H36O11S2 C19H38O12S2

a

RDB — ring/double bond equivalents.

Tables 1 and 2 show the ESI FT-ICR MS elemental composition assignments of major ions (20 ions with higher relative abundance) detected in the whisky samples. In the negative ion mode (Table 1), most ions correspond to deprotonated forms of carboxylic acids, phenols, saccharides derivates and other sulfur or nitrogen compounds. Scheme 1 proposes the chemical structures for such ions; those of m/z 143.1078 and 171.1391 correspond to caprylic and capric acid, respectively. These compounds were also detected in cachaça samples (Moller et al., 2005), and are common in food processing (Beare-Rogers, Dieffenbacher, & Holm, 2001). other identified fatty acids may be related to hydrolysis of fatty acid esters or lactones (Poisson & Schieberle, 2008). The ion of m/z 197.04574 was attributed to ethyl gallate (3), a food additive also found naturally in a variety of plants. Most of the compounds identified in Table 1 are degradation products of maltose and glucose or products from their reaction with phenolic compounds, or carboxylic acids extracted from the wood casks during the aging and maturation process, such as 11 (ellagic acid) and 15 (an isoprenoid derivate). Oxidoreductions and polymerizations occurring during maturation involve compounds present in the raw distillate and wood-derived compounds, such as the lignin-derived p-coumaryl alcohol, coniferyl alcohol and sinapyl alcohol, and could account for formations of 12 and 17 (Scheme 1). The ions of m/z 179.05625 and 341.10943 were also detected via ESI(−)-MS as abundant ions of Scotch whisky samples (Moller et al., 2005). These ions correspond to deprotonated molecules of monosaccharides such as glucose (180 Da) and disaccharides such as sucrose (342 Da). The spectra in the positive ion mode (Table 2) provided composition information complementary to that obtained in the negative ion mode. The majority of compounds were detected by ESI(+) as sodium adducts [M + Na] + and Scheme 2 presents propositions for their structures. As discussed before, major compounds are likely formed from oxidoreduction processes such as 8 and 9; ligninderivate products were also found such as 7 and 8, which are derivates of the sinapyl alcohol (Scheme 2). In some cases, secure molecular formula determination was not possible particularly for the heavier ions and the exponential increase of elemental composition variation that is particularly severe above 450 Da (Cooper & Marshall, 2001). 3.3. Adulteration and counterfeiting The capability of ESI FT-ICR MS to identify whisky counterfeiting was tested by the addition of a common low cost “distillate whisky” commercialized in Brazil, herein referred as “Brazilian distilled whisky”. This “whisky” is known to be frequently used in Brazil to adulterate and counterfeit the much more valuable Scotch whiskies and was added at different amounts to an authentic sample of Johnnie Walker Red Label. Fig. 4 displays the resulting ESI(−) spectra. Note that adulteration is easily perceived by the appearance of some characteristic “marker” ions already at 25% of adulteration, most particularly that of m/z 269.09. The PCA analysis (Fig. 5) shows that ESI(−) FT-ICR is indeed promising as an effective technique for adulteration and counterfeiting screening of whisky and also to estimate the level of adulteration. The PCA scores plot clearly splitting the authentic and adulterated samples into two distinct groups. In ESI(+), PC1, PC2 and PC3 were found to account for 76.7%, 9.0% and 4.2% of the total variance, respectively. Therefore, the total variance explained by the three first PCs is 89.8% at a confidence level of 95%. Additionally, in the ESI(−) results PC1, PC2 and PC3 accounts for 81.8%, 7.8% and 5.5% of the total variance, respectively. Fig. 6(A) and (B) shows a representative pair of spectra for an authentic and counterfeit sample of the same brand (Red Label). Note the immediate recognition of counterfeiting such as via the lack of

102

J.S. Garcia et al. / Food Research International 51 (2013) 98–106 O

OH

OH

HO

O

O

-

[ 1 - H] m/ z 143.10785

[ 2 - H]- m/ z 171.13919

OH

OH

OH

OH OH

2, 172.14633

1, 144.11503

O

HO

O

O

3, 198.05282

4, 200.17763

[ 3 - H] - m/ z 197.04574

[ 4 - H]- m/ z 199.17053

5, 220.05830

OH OH

HO

OH

O

OH

OH

HO O

O

OH OH

O

HO

OH

OH

6, 228.20893

7, 256.24023

8, 270.09508

9, 270.07395 OH

[ 6 - H]- m/ z 227.20190

[ 7 - H]- m/ z 255.23326

[ 8 - H]- m/ z 269.08814

[ 9 - H]- m/ z 269.06698

OH OH

O

HO

HO O

OH O

O

O

O

HO

O

OH

HO

OH

OH

O

HO

OH

OH

OH

OH

O HO

O

HO

HO

OH

OH O

HO

OH

OH

O

O

H 3C

HO

HO

O

OH O

O

HO

OH

OH OH

OH

10, 298.10525

11 302.00627

12 323.09885

13 342.11621

14 360.12678

[ 10 - H]- m/ z 297.09836

[ 11 - H]- m/ z 300.99938

[ 12 - H]- m/ z 324.10565

[ 13 - H]- m/ z 341.10946

[ 14 - H]- m/ z 359.12005

OH

OH

OH

O

O

HO O

O O

HO

OH

O

OH

O

O

O

HO

OH

HO

OH HO

O

O

HO

OH

O

HO

OH

HO

OH OH

O

OH

O

[ 5 - H]- m/ z 219.05125

O

OH HO

O

OH

O

O OH

O

OH O

OH HO

OH

OH OH

OH OH

16, 380.11073

17, 400.12169

18, 414.13734

15, 380.13186 [ 15 - H]- m/ z 379.12518

[ 16 - H]- m/ z 379.10404

[ 17 - H]- m/ z 399.11515

[ 18 - H]- m/ z 413.13080

Scheme 1. Structures proposed for the major compounds detected in the whisky samples by ESI(−) FT-ICR MS.

Table 2 Elemental compositions assigned to ions in ESI(+) FT-ICR data (20 most abundant ions). m/z

Error (ppm)

RDBa

Elemental composition

157.08350 203.05261 213.07336 219.02655 231.08393 271.07884 301.14106 347.09490 365.10548 383.11602 409.16215 411.14730 425.13613 443.16766 509.14777 527.15842 545.16894 589.22561 623.23115 689.21181

0.09 0.01 0.06 0.71 0.08 0.05 0.1 0.09 0.12 0.07 0.03 0.01 0.44 0.05 0 0.03 0.07 0.11 0.06 0.21

0.5 0.5 1.5 5.5 0.5 2.5 5.5 2.5 1.5 0.5 9.5 0.5 14.5 8.5 19 11 17 9.5 24 2.5

C6H14O3Na C6H12O6Na C8H14O5Na C9H8O5Na C8H16O6Na C10H16O7Na C16H22O4Na C12H20O10Na C12H22O11Na C12H24O12Na C22H26O6Na C14H28O12Na C25H22O5Na C22H28O8Na C31H26O2NS2 C25H30O8NSNa C31H31O4NS2 C28H38O12Na C41H36ONS2 C25H46O16S2Na

a

RDB — ring/double bond equivalent.

the Gaussian “grass” of polar compounds around m/z 300–700 and the presence of an abundant sugar ion of m/z 311.07 corresponding to sodiated saccharose. Counterfeit samples (n = 30) from different whisky brands were also analyzed, and the resulting spectra, as expected, displayed quite contrasting and variable profiles of polar constituents (Fig. 6(C)). 3.4. Aging and blending The ability of ESI FT-ICR MS to evaluate whisky aging and blending was also tested (Fig. 7) using samples of whisky from Johnnie Walker Black, Gold and Blue Labels. Note that aging and blending is easily recognized by changes in the polar profiles particularly in the relative abundances of the heavier ions and that of the “ion grass” around m/z 300–500. 4. Conclusion ESI FT-ICR MS has been shown to be a powerful technique for direct analysis at the molecular level of Scotch whiskies, being able to

J.S. Garcia et al. / Food Research International 51 (2013) 98–106

103

OH HO

O HO

O HO

HO HO

H 3C

OH

O

H OH

O

H

OH

H H OH OH

O H 3 CO OH

1, 134.094294

3, 180.06339

2, 150.06808

[1 + Na +] m/ z 157.08350

[2 + Na + ] m/ z 173.05774

[3 +

Na +

] m/ z 203.05261

[4 + Na + ] m/ z 223.05768

[5 + Na +] m/ z 227.0895

O

O

OH

OH O

O

HO

O

HO

OH

H 3CO

6, 208.09468

OCH 3

HO

7, 242.07904

[6 + Na + ] m/ z 231.08393

HO

OH

OH

OH

8, 324.10564

9, 324.10564

10, 342.11621

Na +

Na +

] m/ z 347.09490

[9 +

] m/ z 347.09490

[10 + Na+ ] m/ z 365.10548

OH

OCH 3

H3 CO

OCH3 H 3C

OH O

CH 3

H 3C O H3 C

O

OH

OH

OH

O

HO

OH

HO

HO

O

OH

[8 +

[7 + Na + ] m/ z 265.06827

OH

O

O

HO

O

H OH

HO

OH O

HO

HO

OH

HO H OH H O

5, 204.02974

HO

O

H OH

4, 200.06847

O

O

HO

HO HO

OCH 3

O

HO

O

O

O

O

O

O

O O

H 3CO

OH OH

12, 386.17294

11, 360.126776 [11 + Na+ ] m/ z 383.11602

OCH 3

[12 + Na+ ] m/ z 409.16215

13, 386.17294

14, 402.14673

[12 + Na+ ] m/ z 409.16215

[14 + Na+ ] m/ z 425.13614

Scheme 2. Structures proposed for the major compounds detected in the whisky samples by ESI(+)FT-ICR MS.

100

171.14 199.17

(A)

50

(B)

(C)

Relative Abundance

0 100

485.15

323.17

401.13 431.14 385.14

521.17 485.22

0 100

563.18

359.12

285.08

50

503.16 521.17

375.12 401.13

323.10

171.14 199.17 269.09 239.08

563.18 593.19

269.09

50

179.06 0 100

(D)

341.11

219.05 255.23

239.08

431.14 359.12 285.08 401.13 323.17

485.22

521.17 563.18 593.19

269.09 179.06

50

195.00

251.08

431.14

341.11 277.02

375.12

0 200

300

400

449.15

521.17 500

593.20 600

m/z Fig. 4. ESI(−) FT-ICR MS of (A) authentic Johnnie Walker Red Label sample and adulterated samples with different proportions of the “Brazilian distilled whisky”, (B) 25%, (C) 80% and (D) 100% (v/v).

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de Nível Superior (CAPES) and Petrobras for financial support. We also thank ABRAPE — Associação Brasileira de Bebidas for providing authentic whisky samples.

(A) Brazilian Distilled Whisky

4

References

3 90%

2 Red Label ed

at

er

lt du

A

-1

30 25 20 15 10

25%

-2 -3

5

8

6

0

4

2

PC2

0

-2

-4

1

0

PC

PC3

1

-5 -10

Adulterated

(B)

Brazilian Distilled Whisky

10 8

PC3

6 4 Red Label

2

2 0

0 -2 -4 6

1

8

4

2

PC2

0

-6 -2

-4

-6

PC

-2

-8

Fig. 5. PCA for the (A) ESI(−) and (B) ESI(+) FT-ICR MS data from authentic Johnnie Walker Red Label whisky samples (n = 12) and samples adulterated with different proportions of the “Brazilian distilled whisky” (25, 50, 70, 80 and 90%).

provide a characteristic profile of polar constituents both in the negative and positive ion modes. Samples from different brands and with different ages and blending were also shown to display contrasting profiles. Adulteration and counterfeiting were also promptly identified. This strategy is also probably applicable to other valuable spirits and beverages. Acknowledgments The authors thank the State of São Paulo Research Foundation (FAPESP), Brazilian National Council for Scientific and Technological Development (CNPq), Coordenação de Aperfeiçoamento de Pessoal

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Fig. 6. (A) Authentic spectrum, (B) counterfeit spectrum and (C) PCA plots for the ESI(−) FT-ICR data of Johnnie Walker Red Label whisky samples (authentic and counterfeit).

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Fig. 7. ESI(−) FT-ICR for samples of Johnnie Walker (A) Black, (B) Gold and (C) Blue Label whisky samples.