New Technologies Addressing Challenges in Fruit Juice … · New Technologies Addressing Challenges in Fruit Juice Testing ... Fructose, 2) Glucose, 3) Sucrose, 4) Maltose, 5)

©2008 Waters Corporation

New Technologies Addressing

Challenges in Fruit Juice Testing

Antonietta Gledhill

Waters Corporation

http://10.2.240.144/cgi-bin/ImageFolio31/imageFolio.cgi?direct=_Global_Branding/Logos&img=

©2008 Waters Corporation 2

Waters CorporationA Brief Introduction

Founded in 1958

Number of Employees: >5,000

Operate in 27 countries

—11 Manufacturing Facilities

—Products available >50 countries



Waters CorporationTechnology Focus



Company’s Philosophy: Innovate


http://www.waters.com/WatersDivision/ContentD.asp?watersit=JDRS-6EMRYQ








Application Focus

Biopharmaceuticals

Chemical materials

Clinical

Environmental

Food & Beverage

Pharmaceutical

Proteomics



Use of Analytical Tools to Support Business Decisions

Research & Innovation

Manufacture Quality

Food safety & QA

Established

products

Reformulated

products

New

products

Incoming

Material

Production

monitoring

Product

release

Breakthrough

innovation

Product

stability

Functional

benefit



Routine Analysis


Manufacture Quality

Food safety & QA



Routine Analysis Workflow

SAMPLE PREPARATION

Generic, L-L, SPE…

CHROMATOGRAPHY

Alliance, ACQUITY UPLC, GC

DETECTOR

UV, FLR, ELSD, MS

SOFTWARE

Empower, TargetLynx

SEPARATION CHEMISTRY

SunFire, XBridge, ACQUITY



Routine Analysis Separation Chemistry

BEH Amide Column (2009)

— Carbohydrates & Sugars

Detector: ELSD

* *

5 Food Sugars1

2

3

0.0

Minutes

2.0 4.0 6.0 8.0 10.0 12.0 14.0 16.0 18.0 20.0

4

5

Low Salt Retention = No Salt Interference

1) Fructose, 2) Glucose, 3) Sucrose, 4) Maltose, 5) Lactose (each 1mg/mL), * Salt Interferences

with Salts



Gain Switching

Routine Analysis Separation Chemistry

Minutes

0.0 1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0 10.0 11.0 12.0 13.0

Malto-tri, tetra, penta, hexa, hepta-ose

Double Chocolate Stout

Food Sugar Standard

Xylo

se

Arab

inose

Glu

cose

Malt

ose

Lacto

se

Fu

cose

Malt

otr

iose

Malt

ote

traose

Malt

op

en

taose

Malt

oh

exaose

Malt

oh

ep

taose

1.3µL injection volumeSamples: 100% Beer (No dilution or filtering)



Routine AnalysisLiquid Chromatography: Acquity UPLC®

Soft drinks

Example 1: Established HPLC methods

Speed Advantage




PolyphenolsVitamin C

Advantages: Resolution and Speed

Soft drinks




Benefits of Acquity UPLC

Chromatographic resolution

— Interferences / closely eluting

peaks

— …Selectivity from Core Detectors

Reduction in run-times

— Routine analysis

— Method development

Financial savings

— Solvent

— No of Instruments in lab

(servicing…)



Routine AnalysisDetectors - MS

ACQUITY SQD

Complementary tool for core detector

applications

Mass analysis for chromatographer

Selective & sensitive detector

IntelliStart with automated

— Set-up

— Calibration

— Method development

— Instrument diagnostics



Time0.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50 5.00 5.50

AU

0.0

5.0e-3

1.0e-2

1.5e-2

2.0e-2

2.5e-2

3.0e-2

3.5e-2

4.0e-2

4.5e-2

5.0e-2

5.5e-2

6.0e-2

6.5e-2

7.0e-2

1.73

1.52

0.35

0.22

1.24

0.51

0.84

2.35

2.02

1.90

2.17

2.66

2.46

5.50

3.17

3.08

2.79

2.95

3.41

3.28

5.16

4.81

5.73


Additional security of selectivity & confirmation using MS

m/z100 120 140 160 180 200 220 240 260 280 300 320 340 360 380 400 420 440 460 480 500

%

0

100

%

0

100443

293

257

191

147134131121105

179173149 187

209193 256214

233229 247 264

277293

385

341294

323315299 311

329

355347 367 375

407386

406396 442408 433421

444

445

491475458 483 497

353

191

176

141

119109125 161

158173 179

192353201

198217 218 333233 319301269

239253 283 286

315

375

354

355

358

433411408

376 395431 451438

452463

491481 493

Advantage 1: Selectivity of MS in full scan mode




Time0.20 0.40 0.60 0.80 1.00 1.20 1.40 1.60 1.80 2.00 2.20 2.40 2.60 2.80

%

0

1.08

MS: m/z 271

Additional security of selectivity & confirmation using MS

Advantage 1: Selectivity of MS in SIR mode



Routine AnalysisDetectors – ELSD and MS

Stevia(5mg/mL)

Stevia Standards

Iso

ste

vio

l

Ste

vio

sid

e

Reb

au

dio

sid

e A

Reb

au

dio

sid

e C

Minutes0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5 6.0

0100200300400500600700800900

1000

ELSD (77.5% ACN with 0.2% TEA)

MS (77.5% ACN with 0.1% NH4OH) SIR of 5 Channels ES-TIC

1.15e6

Minutes

0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5 6.00.0

%

0

100

m/z = 966.1(Rebaudioside A)

m/z = 950.1(Rebaudioside C)

m/z = 803.8(Stevioside)

Stevia(50µg/mL)

m/z = 318.5(Isosteviol)

Advantage 2: Sensitivity of MS



Summary for Routine Analysis

Technological Solutions to support Beverage Analysis

Separation Chemistries – Continual

research and development efforts here to

support challenging applications

ACQUITY UPLC – Excellent resolution &

increased speed of analysis

SQD MS – Additional confirmatory tool

benefitted from selectivity & sensitivity of

MS



Research:

Investigative / Troubleshooting


Manufacture Quality

Food safety & QA



Research: Investigative Analysis, Troubleshooting

Examples include:

New Product Development

— Ingredient / product profiling and characterisation

— Structural elucidation

— Degradation / transformation process

— Stability studies

Troubleshooting

— Authenticity and origin

— Manufacturing: Process optimisation

— Final Product: Contaminant screening

— Customer complaints & Competitor analysis



Research: Investigative Analysis…Published Data

“…To advance our understanding of the benefits

of GADF8 it is important to thoroughly

characterize its putatively active phenolic

components.

Mass spectrometry coupled to liquid

chromatography (LC/MS) has been increasingly

used in the structural characterization…”



Research: Investigative / Troubleshooting

SAMPLE PREPARATION

Generic, L-L, SPE

CHROMATOGRAPHY

ACQUITY UPLC, GC

DETECTOR

Tof (Time of flight) MS

SOFTWARE

ChromaLynx, MarkerLynx, MetaboLynx, MassFragment


ACQUITY BEH, ACQUITY HSSGENERIC

METHOD

CONDITIONS



Research: Xevo QTof MS

Xevo QTof MS

DATA ACQUISITION

MODE

Low collision energy mode produces M+H or M-H

Ramp collision energy mode produce fragmentation data



Research: Software Tools

• Deconvolute Chromatograms: ID components in samples

• Sample Comparison: common & unique components

• Library search

QTof MS

DATA

• Multivariate statistical tool (PCA, PLS-DA)

• Perform database search (e.g. ChemSpider, user database) to ID potential markers

• Enables rapid metabolite identification (expected & unexpected)

• Integration with MassFragment for structural elucidation

• Structural Elucidation of observed exact mass ion fragments




SAMPLE PREPARATION

Generic, L-L, SPE

CHROMATOGRAPHY

ACQUITY UPLC, GC

DETECTOR


SOFTWARE



ACQUITY BEH, ACQUITY HSS

• Dilute and shoot

• ACQUITY column

• ACQUITY UPLC

• Xevo QTof MS (MSE mode)

• ChromaLynx XS• MassFragment




• Deconvolute Chromatograms: List components in sample

• Sample Comparison: Common & unique components

• Library search




[C16H11N2O4S]-

Precursor Information



Research: Elemental Composition & Chemspider

[C16H11N2O4S]-

Structural Elucidation Tools



Research: MassFragment

Product Ion Fragments

[C16H11N2O4S]-

Structural Elucidation Tool



Research:Investigative / Troubleshooting

SAMPLE PREPARATION

Generic, L-L, SPE

CHROMATOGRAPHY

ACQUITY UPLC, GC

DETECTOR


SOFTWARE



ACQUITY BEH, ACQUITY HSS

• Dilute and shoot

• ACQUITY BEH C18

• ACQUITY UPLC

• Xevo QTof MS (MSE mode)

• MarkerLynx• MassFragment



Research: MarkerLynx XS (Chemometric Software)

Sample List

Marker Table

S-Plot results

Trend-PlotChromatogram



Research:Chemometrics visual result

Kenya

Colombia

Colombia

Ethiopia

JavaBlend

Nicaragua

Costa Rica



Research: Grouping within a region



Research: Marker Identification



Research:Marker Database



Summary for Research Analysis

Technological Solutions to support Beverage Analysis

ACQUITY UPLC – Excellent resolution

and speed of analysis / method

development

Xevo QTof MS – Confirmatory exact

mass tool allowing experimental diversity

Software Tools – Provide intelligent

information to support investigative analysis

in research and troubleshooting environments



Thank-you for your attention

Any Questions?


Documents

New Technologies Addressing Challenges in Fruit Juice … · New Technologies Addressing Challenges in Fruit Juice Testing ... Fructose, 2) Glucose, 3) Sucrose, 4) Maltose, 5)