HUMAN HEALTH • ENVIRONMENTAL HEALTH

What’s really in our food?

June 20152015

2

10/1

Food Integrity & Safety Issues…

Peanut Butter

- Salmonella

>€45M Fake Butter -

Non Food Oils

£100M

Spices -

Dye

Enhanced

Juices-

Fungicides

Eggs-

Dioxins

$1B

loss

Wine -

Dilution >18

dead

6 dead,

>300,000

sick

Milk Powder

Melamine

Spinach -

Salmonella

$350M

loss

Recalled

>250,000

Frozen Berries

- Hep A $1.7mil

3

Food industry trends in context to science …

Consumer Trust & Brand Value

Regulatory Safety

Quality & Consistency

Authenticity & Origin

Deter, Prevent, Detect

Fraud

Product Innovation

Value Chain Safety

Supply Chain Integrity

Brand Innovation

Industry Initiatives

4

10/1

A Focus on …

• Food authenticity

◦ Ingredient substitution

◦ Economic adulteration

◦ Incorrect labelling

◦ A question of provenance

• Food Safety

◦ Levels of toxic elements

◦ Residuals

◦ Contamination

5

Economically motivated adulteration

• “… the deliberate adulteration of

food… to inflate profits by fraudulent means” (FDA)

• Examples:

◦ Dilution of valuable oils with

cheaper ones

◦ Substitution of cheaper juices for

pomegranate juice

• Likely to involve high

concentrations of the adulterant

• The tools used to test for EMA may

also be useful for screening for

other types of contamination or

substitution, and in industries other

than food

6

Real-Time Detection Using Direct Sampling TOF Mass Spectroscopy

• Development of technique

known as “field-free APCI”

• A stream of Nitrogen gas is

heated

• Ionized by a corona discharge

needle.

• The ionized nitrogen flows over

the target sample, generating

ions from it.

• Readily swap between direct

sampling source and LC

introduction

Whole Foods

Liquids

Powders

7

How AxION DSA works

8

148.07 155.14

163.05

164.06

168.01 175.08

180.08

182.04 189.05

191.08

197.92

198.09

199.09

202.08

208.11

209.10

210.11

212.11

219.11

221.91

222.11

223.10

226.08 226.12

227.12

228.21

239.13

240.09

243.11

249.13

255.15

0 160 170 180 190 200 210 220 230 240 250 260m/z

Carbamates analysis in orange juice…

Spectrum of juice indicating presence of carbamates

9

Adulteration of our everyday commodities

10

No chromatography, No Sample Preparation

LIQUID EXTRACT SAMPLED DIRECTLY ON MESH ANSWER IN <30 SECONDS

Real-time Detection for Critical Authenticity Issues

Vanilla widely used AND the 2nd most

expensive spice in the world

12,000 Tons Synthetic Vanillin produced but only 20 tons extracted

from the bean Synthetic $12/kg vs.

$2,000/kg For Natural

4-Hydroxybenzaldehyde

ABSENT in Synthetic

Vanillin

11

No chromatography, No Sample Preparation, results in 30 seconds

LIQUID MILK SAMPLED DIRECTLY ON MESH

Real-time Detection – Dietary Reconstruction for Authenticity

Hippuric acid identified as a grass

feed marker in literature.

Shop bought organic and non-organic milk

analysed directly

Current USDA certification demands

that cows are fed grass for AT LEAST 120

days per year

12

Cow ß-LG A

Buffalo ß-LG

Cow

ß-LG B

Little Sample Preparation, LC-TOF measurement

LIQUID MILK SAMPLED AFTER PROTEIN PRECIPITATION

Real-time Detection – Species ID and Intact Protein Profiling

Protein development also key area of

research TOF-MS very suitable for measuring INTACT

PROTEINS

Increased interest in non-cows milk for health and taste

benefits

Cow ß-LG A

Buffalo ß-LG

Buffalo: 50% Cows Milk Substitution

Buffalo: 5% Cows Milk Substitution

Sheep ß-LG-A

Cow ß-LG-B

Sheep ß-LG-B Cow ß-LG-A

Sheep: 50% Cows Milk Substitution

Other protein work: Soy

Proteins, Camel Milk

Powder and Human Milk

13

Little Sample Preparation, DSA-TOF measurement

MILK SAMPLED AFTER PROTEIN PRECIPITATION

Detecting Fraud – Melamine in Liquid Milk & Milk Powder

Continued threat in milk powder but also

liquid milk. Rapid screening tool in DSA technology.

Melamine is a nitrogen rich compound added to diluted milk to falsify

the test for protein content.

Detection Limit:

adult food of 2.5ppm

Infant food 1ppm

DSA method readily

detects down to 1ppm

500

1000

1500

2000

2500

3000

3500

4000

4500

5000

32.3 32.4 32.5 32.6 32.7 32.8 32.9 33.0 33.1 33.2 33.3 33.4

EICm/z (127.069:127.077)

Amplitude

Time (min)

Cu

EIC of varying concentrations of

melamine spiked in milk powder

127.0724

0

50

100

150

200

250

300

350

400

126.5 127.0 127.5 128.0 128.5 129.0

Mass

Amplitud

e

m/z

Cur

Exact mass = 127.0727

ppm error = 2.3

Mass spectrum of melamine

spiked in milk powder (1 ppm)

50 ppm spike

5 ppm spike 1 ppm spike control

Calibration curve of melamine spiked in liquid milk y = 0.0013x - 0.011

R² = 0.9997

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0 100 200 300 400 500 600Conc of melamine (ng/mL)

Are

a r

atio

14

No chromatography, No Sample Preparation

LIQUID MILK SAMPLED DIRECTLY ON MESH ANSWER IN <30 SECONDS

Deterring Fraud – Screening for Future Adulterants

List of 5 additional future adulterants to

melamine well documented:

Urea, Biuret, Cyanuric Acid, Dicyandiamdie,

Cyromaxin,

Common Adulterants Sodium Thiosulphate,

Hydrolyzed leather meal,

calcium hydroxide, salicylic

acid, gelatin

Future threats Cyanoguanidine,

Amidinourea,

Aminotriazole,

Isobutylidenodiurea,

caprolactam, biuret

15

10/1

Pomegranate Juice Adulteration

DSA-TOF No sample prep; results in seconds

Tartaric acid is only present in grape juice; so it can be used as a marker for the detection of pomegranate juice adulteration with grape juice.

Can easily identify as low as 1% grape juice mixed in pomegranate juice

Pomegranate juice No Tartaric acid

detected

1 3 3 . 0 1 4 1

1 4 9 . 0 0 8 8

1 9 1 . 0 2 0 2

0

1 0 0 0 0

2 0 0 0 0

3 0 0 0 0

4 0 0 0 0

5 0 0 0 0

6 0 0 0 0

7 0 0 0 0

8 0 0 0 0

9 0 0 0 0

0 1 0 0 2 0 0 3 0 0 4 0 0 5 0 0 6 0 0 7 0 0

M a s sR E T ( 0 2 . 2 9 1 3 3 )

Am

pli

tu

de

m / z

Pomegranate juice with 1% grape juice

1 3 3 . 0 1 4 8

1 4 9 . 0 1 0 5

1 9 1 . 0 2 0 1

0

2 5 0 0 0

5 0 0 0 0

7 5 0 0 0

1 0 0 0 0 0

1 2 5 0 0 0

0 1 0 0 2 0 0 3 0 0 4 0 0 5 0 0 6 0 0 7 0 0

M a s sR E T ( 0 2 . 0 4 7 9 1 )

Am

pli

tu

de

m / z

Tartaric Acid

[M-H]-

Mass Accuracy = 8.72 ppm

Pomegranate juice with 2% grape juice

Tartaric Acid

[M-H]-

Mass Accuracy = 2.68ppm

1 3 3 . 0 1 4 9

1 4 9 . 0 0 9 6

1 9 1 . 0 2 0 4

0

2 5 0 0 0

5 0 0 0 0

7 5 0 0 0

1 0 0 0 0 0

1 2 5 0 0 0

0 1 0 0 2 0 0 3 0 0 4 0 0 5 0 0 6 0 0 7 0 0

M a s sG E N ( 0 1 . 6 5 4 5 8 : 0 1 . 7 0 4 5 8 )

Am

pli

tu

de

m / z

Tartaric Acid

[M-H]-

Mass Accuracy = 2.68 ppm

Pomegranate juice with 5% grape juice

16

10/1

Fruit

Fungicide screening Organic Authenticity Mycotoxin analysis

DSA-TOF No sample prep; results in seconds

Surface analysis of fungicides on oranges with a swab test. Can easily analyze for pesticides and fungicides on food surfaces with DSA.

DSA-TOF No sample prep; results in seconds

LC-UV Quantitative analysis

Fungicides such as imazalil are banned in organic production and MRL’s are in place for conventional fruits. Rapid analysis of organic and non-organic citrus fruits can detect the presence of any fungicides.

EIC for imazalil in two organic and five conventional fruits

1% Grape juice in pomegranate juice

Patulin in apple juice

Patulin is a naturally occurring toxin that is not destroyed by the pasteurisation process. Due to its potential toxicity MRL have been set.

Thiabendazole

[M+H]+

0.5ppm

Imazalil

[M+H]+

2.36 ppm

17

No chromatography, Little Sample Preparation, results in 30 seconds

Olive oil diluted to 1 % in IPA with 10 mM Ammonium acetate then SAMPLED DIRECTLY ON MESH

Rapid Authenticity Checks - Olive Oil

Profile of fatty acids and their ratios can be used to identify

olive oil, other edible oils and any potential

adulteration.

A recent study showed 70% of olive oils did not meet their

label claims Oleic acid exhibits the highest response L/O = 0.18 Ln/O = 0.017

18

No chromatography, Little Sample Preparation, results in 30 seconds

Soya oil diluted to 1 % in IPA with 10 mM Ammonium acetate then SAMPLED DIRECTLY ON MESH

Rapid Authenticity Checks - Olive Oil

The ratio of Linoleic acid to Oleic Acid

response can be used as a way to detect

adulteration of olive oil.

Cheaper vegetable based oils are a

common adulterant. Linoleic acid has highest response in soy based vegetable oil. L/O = 1.86 Ln/O = 0.29

Olive oil is composed of ~ 75% mono-

unsaturated oleic acid. Soy based veg oil

consists of polyunsaturated

linoleic acid, ~ 55 % & linolenic acid ~ 5%.

19

No chromatography, minimal sample preparation

RAPID CONFIRMATION OF OLIVE OIL ADULTERATION – RESULTS IN SECONDS

Rapid Authenticity Checks - Olive Oil

The ratio of Linoleic acid to Oleic Acid response can be used as a way to

detect adulteration of olive oil.

Effect of Soy based Vegetable oil adulteration in olive oil on Linoleic to Oleic acid response

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0 20 40 60

Ra

tio

Re

spo

nse

of Li

no

leic

Ac

id

to O

leic

Ac

id

Vegetable Oil Adulteration in Olive Oil/%

Mass Spectra of Olive oil adulterated with 10 % Soy based Vegetable oil

Olive oil is composed of ~ 75% mono-

unsaturated oleic acid. Soy based veg oil

consists of polyunsaturated

linoleic acid, ~ 55 % & linolenic acid ~ 5%.

L/O = 0.30 Ln/O = 0.031

20 10/1

Adulteration of Fraud of Spices?

• Potential food safety issue with allergens

present

• The opportunity for adulteration?

◦ Poor cumin harvest

◦ Abundance of nut shells

◦ Bulking out of raw material

• Products recalled

◦ Incumbent method ELISA

◦ Tests showed positive for peanut

and almond

21

Accurate mass and isotope ratio data

for147mw using EC ID gave a hit for

coumarin; 177mw is methoxycoumarin.

MeOH EXTRACTION, LIQ ANALYSED ANSWER IN <30 SECONDS

INVESTIGATING QUANTITATION BY GCMS

Deterring Fraud – Screening for Unknown Adulterants

Potential adulteration of ground cumin following a bad

harvest .

Power of the TOF to detect unknown

unknowns used to identify markers of

adulteration.

Cumin

Mehlab

y = 3415.8x + 2404.3

R² = 0.9994

0.000

5000.000

10000.000

15000.000

20000.000

25000.000

30000.000

35000.000

40000.000

45000.000

50000.000

0.00 2.00 4.00 6.00 8.00 10.00 12.00 14.00

Pe

ak

Are

a

Conc (ppm)

Non-target analysis of pesticides

Utilizing LC TOF analysis with AxION EC ID Software

Inputting accurate mass and isotope

ratio data using EC ID gave a list of

potential compounds.

PESTICIDE CARBARYL IDENTIFIED AS THE UNKNOWN

Ensuring Food Safety – Detecting Contamination

Pesticides linked to cancer, birth defects, liver, kidney damage

EU directive: EU 2002/657/EC

regulation on MRL of pesticides in or on food

and feed of plant intended for human

consumption

2 0 2 .0 8 6 6

2 0 2 .5 7 1 1

2 0 3 .0 8 8 7

2 0 4 .0 9 1 2 2 0 4 .1 5 8 3

0

1 0 0 0

2 0 0 0

3 0 0 0

4 0 0 0

5 0 0 0

6 0 0 0

7 0 0 0

8 0 0 0

9 0 0 0

1 0 0 0 0

1 1 0 0 0

2 0 1 .5 2 0 2 .0 2 0 2 .5 2 0 3 .0 2 0 3 .5 2 0 4 .0 2 0 4 .5

M a s sR E T ( 0 2 .8 8 6 0 0 )

Am

pli

tu

de

m / z

Identifying an unknown analyte in QuEChERS extract of artichokes

Unknown peak

eluting at 3mins

Accurate mass of unknown peak in artichoke extract

Based on accurate mass information and AxION EC ID prediction the likely

candidate is Spinosad A.

Retention time matching of the celery extract with a standard

confirms the pesticide indentity.

Ensuring Food Safety – Contaminated Organic Produce?

Spinosad A is an insecticide sprayed

on vegetables

EPA registered pesticides are

prohibited in organic production; however residues up to 5% EPA

tolerance allowed (environmental effects)

Identifying an unknown analyte in QuEChERS extract of organic celery

Accurate mass of extract of organic celery

0

5 0

1 0 0

1 5 0

2 0 0

2 5 0

3 0 0

3 5 0

4 0 0

4 5 0

0 1 2 3 4 5 6 7

E I Cm / z ( 7 3 2 . 3 0 0 : 7 3 2 . 5 0 0 )

Am

pli

tu

de

T i m e ( m i n )

7 3 2 . 4 7

7 3 3 . 4 7

7 3 4 . 5 3

0

1 0

2 0

3 0

4 0

5 0

6 0

7 0

7 2 5 . 0 7 2 7 . 5 7 3 0 . 0 7 3 2 . 5 7 3 5 . 0 7 3 7 . 5 7 4 0 . 0 7 4 2 . 5

M a s sG E N ( 0 3 . 8 3 5 2 5 : 0 4 . 2 0 1 9 1 )

Am

pl

it

ud

e

m / z

C u

31

Summary

• DSA is an effective tool for rapid screening

◦ Real time detection for authenticity

◦ Fast screen for contamination detection and identification

◦ Targeted and non-targeted analysis

• Continued focus on food adulteration and food safety

• Innovation is key to support customers

• Extending collaborations with food companies

• Solutions and applications