Ten Second DART®-MS - IonSense€¢ Scan Settings: 1 µ-scan by 250 ms max inject time • AGC Target: Balanced (1e 6) ... • After pressure from FDA GMP Herbal Products, Inc. issued

Ten Second DART®-MS: Ultra-Fast QC Screening of

Dietary Supplements and Pharmaceuticals

with a Next Generation DART Source

IonSense, Inc.999 Broadway Suite 404

Saugus, MA 01906 USA

Elizabeth Crawford, Joseph LaPointe, and

Brian Musselman

125th AOAC Annual Meeting

September 18-21, 2011

Overview• Next Generation DART Source

for Rapid Screening:

• DART-SVP vs. ID-CUBE

• High Resolution/Accurate

Mass measurements with

benchtop Exactive Orbitrap

mass spectrometer

• Dietary Supplement and

Pharmaceutical Screening

• Are they safe and what’s

in them???

www.thecanarynews.com, Issue 52, August 26 – September 8, 2011

• Why take a dietary supplement?

• Support immune system

• Reduce risk of illness/age related conditions

• To support healing process

• To boost energy and performance in activities

• Weight control

DART-SVP Ionization Source

DART-SVP Ion Source

ID-CUBE: Built on DART Technology

ID-CUBE™ Ionization Source

MS InstrumentationThermo Exactive

• Accurate mass measurements within3 ppm for all work presented (external)

• Scan Parameters:• Positive & Negative Ion Modes

• Resolution: “High” - 50,000

• Fragmentation: None, HCD Gas Off

• Scan Settings: 1 µ-scan by 250 ms max inject time

• AGC Target: Balanced (1e6)

• Exactive Inlet Parameters:• Capillary Temp: 200° C 200° C

• Capillary Voltage: 25 V - 50 V

• Tube Lens Voltage: 120 V -120 V

• Skimmer Voltage: 26 V -25 V

• For all ID-CUBE experiments the Exactive Spray

Voltage was set to 4 kV and the following

parameters set to zero:

• Sheath Gas Flow, Aux Gas Flow, Sweep Gas Flow

ID-CUBE™ Settings• ID-CUBE Source:

• Desorption Temperature Settings:

• Low, Medium, High

• Sampling Time: Continuously heated for 10 seconds after initial signal observed

1. Spot 5 µL aliquot or powder

residue on the OpenSpot™ Card

2. Let

sample dry

for easy

transport

or analyze

directly

when wet

3. Insert

sample

card into

the slot on

ID-CUBE

Source

4. Set the desired heating setting

on ID-CUBE control box and press

button for instant desorption

DART-SVP vs. ID-CUBETemperature Profiling

DART-SVP - Temp. Ramp Method

RT: 0.00 - 5.71

0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5

Time (min)

0

10

20

30

40

50

60

70

80

90

100

Rela

tive A

bu

ndance

RT: 2.55

RT: 5.37

RT: 5.00RT: 2.91

RT: 0.71

RT: 0.31

2.52

4.97

5.032.880.70

2.49 5.32

NL: 8.65E7

m/z= 165.76838-166.22364 MS ICIS DART-SVP_Mesh_Sulfodene_Skin_Med_Temp_Ramp_NEG_20110810_02250⁰ C

350⁰ C

450⁰ C

Run time

of 5.5 min

0.00 0.05 0.10 0.15 0.20 0.25 0.30 0.35 0.40 0.45 0.50 0.55 0.60 0.65

Time (min)

0

10

20

30

40

50

60

70

80

90

100

Re

lative

Ab

un

da

nce

RT: 0.47SN: 1006RMS

RT: 0.42SN: 671RMS

RT: 0.15SN: 31RMS

0.40 0.52 0.55

0.60

NL: 1.29E5

m/z= 496.33259-496.48977 MS ICIS IDCUBE_Alli Capsule Extract #1_TEMP_RAMP_WET_POS_20110819

200 400 600 800 1000 1200 1400 1600 1800 2000

m/z

0

50

100

0

50

100

Re

lative A

bu

nda

nce

0

50

100171.11257

207.15888

171.11247

581.45114

991.78951

513.42491 991.78962

581.45101 1487.18105947.80029

469.43533

171.11243

NL: 3.97E5

IDCUBE_Alli Capsule Extract #1_TEMP_RAMP_WET_POS_20110819#13-18 RT: 0.15-0.21 AV: 6 T: FTMS + p NSI Full ms [100.00-2000.00]

NL: 3.08E5

IDCUBE_Alli Capsule Extract #1_TEMP_RAMP_WET_POS_20110819#28-36 RT: 0.32-0.41 AV: 9 SB: 6 0.02-0.07 T: FTMS + p NSI Full ms [100.00-2000.00]

NL: 1.45E5

IDCUBE_Alli Capsule Extract #1_TEMP_RAMP_WET_POS_20110819#41-51 RT: 0.47-0.57 AV: 11 SB: 6 0.02-0.07 T: FTMS + p NSI Full ms [100.00-2000.00]

MEDIUMHIGH

Run time of

0.65 minLOW

LOW

MEDIUM

HIGH

ID-CUBE – Temp. Gradient

Omega Fatty Acid &

Herbal Supplements

T: FTMS - p NSI Full ms [100.00-1000.00]

270 275 280 285 290 295 300 305 310 315 320 325 330 335 340 345 350

m/z

0

10

20

30

40

50

60

70

80

90

100

Rela

tive A

bun

dance

301.21678

281.24809

279.23248

327.23246

283.26371

275.20135

Nature Made Triple Omega Capsules 2% Dilution in Toluene

Eicosapentaenoic Acid (EPA)

Docosahexaenoic Acid (DHA)

Omega-3

Omega-6

Omega-9

T: FTMS - p NSI Full ms [100.00-1000.00]

275 276 277 278 279 280 281 282 283 284 285 286

m/z

0

10

20

30

40

50

60

70

80

90

100

Re

lative A

bunda

nce

281.24810C 18 H 33 O2

279.23248C 18 H 31 O2277.21684

C 18 H 29 O 2

283.26372C 18 H 35 O 2

282.25143275.20135C 18 H 27 O 2

280.23578278.22011

284.26710

α Linolenic Acid

Nature Made Triple Omega 2% Dilution in Toluene

Omega-3

Omega-6

Omega-9

Linoleic Acid

Oleic Acid

Stearidonic

Acid

Omega-3, -6, -9 Fatty Acids

in Dietary Oil Supplements

Nature Made Triple Omega Nature Made Flax Oil Target Origin Flax Oil

No

rma

lize

d A

ve

rag

e P

ea

k A

rea

Stearidonic Acid α-linolenic Acid Linoleic Acid Oleic Acid

n=5 per dietary oil brand

Replicate Samples – Triple Omega

RT: 0.00 - 3.54

0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5

Time (min)

0

50

100

0

50

100

0

50

100

Rela

tive A

bu

ndance

0

50

100

RT: 1.69SN: 1432RMS

RT: 0.21SN: 1310RMS

RT: 2.93SN: 1246RMS

RT: 1.02SN: 1094

RT: 2.30SN: 1001RMS

1.713.001.300.34 2.461.81

RT: 0.21SN: 2060RMS

RT: 1.69SN: 2017RMS

RT: 2.93SN: 1802RMS

RT: 1.02SN: 1562RMS

RT: 2.30SN: 1465RMS

1.710.251.30 3.000.34 1.77 2.461.25

RT: 1.67SN: 1284RMS

RT: 0.21SN: 1281RMS

RT: 2.93SN: 1169RMSRT: 2.30

SN: 932RMS

RT: 1.02SN: 910RMS 1.710.25

3.001.300.34 1.77 2.411.25

RT: 1.67SN: 657RMS

RT: 0.21SN: 635RMS

RT: 2.93SN: 563RMSRT: 2.30

SN: 431RMSRT: 1.02SN: 414RMS 1.710.25

3.00

NL: 3.49E5

m/z= 281.23996-281.25792 MS ID-Cube_Nature Made Triple Oemga_Cap 1_High_WET_NEG_1_Different Card_20110819

NL: 1.99E5


NL: 1.93E5


NL: 6.28E4


Oleic Acid

Linoleic Acid

α Linolenic Acid

Stearidonic

Acid

Omega-3 Omega-6 Omega-9

Omega-3 and -9 Fatty Acids

Found in Marine Oil Samples

Nature Made Fish Oil Nature's Bounty Fish Oil Nature Made Triple

Omega

Schiff's Megared Krill Oil

No

rma

lize

d A

ve

rag

e P

ea

k A

rea

EPA DHA Oleic Acid

n=5 per dietary oil brand

QC Monitoring of Anthocyanins in an

Elderberry Herbal Supplement

0

0.05

0.1

0.15

0.2

0.25

0.3

0.35

0.4

1 2 3 4 5 6 7 8

No

rma

lize

d A

ve

rag

e P

ea

k A

rea

Elderberry Capsules (n=3 per Capsule)

Delphinidin Pelargonidin Cyanidin8.6 % CV 11.9 % CV27.2 % CV

Performance Drug

Scraping Contents of “Mexican sildenafil”

This tablet was sold by a drug pusher

in Mexico as a male enhancement drug

that resembles the appearance of the

well known erectile dysfunction (ED)

drug, Viagra.

The tablet was sliced open and using a

standard glass capillary tube a small

amount of material was scarped from

the inside of the tablet, transferred

onto the mesh and directly screened

for the FDA regulated API sildenafil.

“Mexican sildenafil” Tablet Scraping

474 475 476 477 478 479 480

m/z

0

20

40

60

80

100

0

20

40

60

80

100

Rela

tive

Abunda

nce

475.21220

476.21497

477.20881478.21363 479.21464

475.21122

476.21513

477.21716

NL:1.70E4

C 22 H 31 O 4 N 6 S: C 22 H 31 O 4 N 6 S 1

p (gss, s /p:40) Chrg 1R: 50000 Res .Pwr . @FWHM

NL:3.04E3

IDCUBE_Mexican Viagra_High_Tablet Scrape_POS_1_20110822#106 RT: 0.99 AV: 1 SB: 16 0.75-0.83 , 1.26-1.29 T: FTMS + p NSI Full ms [100.00-1000.00]

Elemental composition search on mass 475.21

m/z= 470.21-480.21

m/z Theo. Mass Delta (ppm)

RDB equiv.

Compositionm/z Theo. Mass Delta (ppm)

RDB equiv.

Composition

475.21122 475.21220 -2.06 10.5 C 22 H 31 O4 N 6 S

[M+H]+

Elemental Composition

Simulation

[M+H]+

DART Spectrum

Weight Loss Aid/Supplements

Pai You Guo “Herbal” Slim Capsules

Article cut from Boston Globe newspaper, August 16, 2011.

November 12, 2009

• After pressure from FDA GMP Herbal Products, Inc. issued a voluntary

nationwide recall for Pai You Guo, which was found to contain

undeclared drug ingredients.

http://www.fda.gov/Safety/Recalls/ArchiveRecalls/2009/ucm190403.htm

Pai You Guo Slim Capsules

• Listed Ingredients on package insert:

• Evil powder of, low gather the sugar,

malt, chitin, the, meal fiber, apple

withdraws the thing, Living the plain C,

vitamin E, vitamin B2

Pai You Guo Tablet – Direct PowderThe capsules were twisted open and a

very small amount of the fine powder

was transferred with a glass capillary

tube onto the mesh sampling area of an

OpenSpot card for rapid screening for

the FDA regulated API sibutramine.

The capsule powder was also

dissolved in ACN and directly

analyzed from the sample cards.

Pai You Guo “Herbal” Slim CapsulesClearly contains Sibutramine

279 280 281 282 283 284 285 286

m/z

0

20

40

60

80

100

0

20

40

60

80

100

Rela

tive A

bu

ndance

0

20

40

60

80

100280.18265

282.17976

281.18593

283.18313284.18633

280.18204

282.17913

281.18540

283.18237

280.18191

282.17887

281.18525

283.18213

NL:7.35E4

C 17 H 27 N Cl: C 17 H 27 N 1 Cl 1

p (gss, s /p:8) Chrg 1R: 50000 Res .Pwr . @FWHM

NL:1.68E5

ID-Cube_Pai You Guo_POWDER_POS_High_20110821#45-51 RT: 0.46-0.53 AV: 7 SB: 5 0.01-0.05 T: FTMS + p NSI Full ms [100.00-1000.00]

NL:1.72E6

ID-Cube_Pan You Guo_20 ug-mL_WET_POS_Low_3_5 Capsules_20110821#232-247 RT: 2.18-2.32 AV: 16 SB: 39 0.10-0.46 T: FTMS + p NSI Full ms [100.00-1000.00]


m/z= 275.18-285.18

m/z Theo. Mass Delta

(ppm)

RDB

equiv.

Compositionm/z Theo. Mass Delta

(ppm)

RDB

equiv.

Composition

280.18204 280.18265 -2.19 4.5 C 17 H 27 N Cl

280.18401 -7.04 1.5 C H O N


m/z= 275.18-285.18

m/z Theo. Mass Delta

(ppm)

RDB

equiv.

Compositionm/z Theo. Mass Delta

(ppm)

RDB

equiv.

Composition

280.18191 280.18265 -2.66 4.5 C 17 H 27 N Cl

280.17997 6.91 0.0 C H O Cl

[M+H]+

Elemental Comp. Simulation

[M+H]+ Capsule Powder Direct

DART Spectrum

[M+H]+ Capsule Contents in ACN

DART Spectrum

HCD Experiment - Pai You Guo Capsule

T: FTMS + p NSI Full ms2 [email protected] [100.00-1000.00]

100 120 140 160 180 200 220 240 260 280

m/z

0

10

20

30

40

50

60

70

80

90

100

Re

lativ

e A

bun

da

nce

125.01518

139.03071

111.11687

149.02315

280.18225179.06198

165.04634199.16905

Sibutramine

[M+H]+

HCD Gas on, 20 eV

Major

Fragment Ion

Minor

Fragment Ion

FDA Warning – Jan. 18, 2010

Counterfeit Alli™

http://www.fda.gov/NewsEvents/Newsroom/PressAnnouncements/2010/ucm197857.htm

The 120 capsule Alli

weight loss aid refill

pack, which was

subject of an FDA

warning in Jan. 2010

was purchased from an

online vendor. Upon

receipt the product

appeared to be

authentic based on the

appearance of the

packaging and capsule

contents.

Orlistat Detected

480 485 490 495 500 505 510 515 520 525 530

m/z

0

10

20

30

40

50

60

70

80

90

100

Rela

tive A

bun

dance

513.42529

496.39883

514.42853

497.40229

515.43161498.40547 527.38057


m/z= 491.40-501.40


RDB equiv.


RDB equiv.

Composition

496.39883 496.39965 -1.65 3.5 C 29 H 54 O5 N

496.39697 3.75 -1.0 C H O


m/z= 508.43-518.43


RDB equiv.


RDB equiv.

Composition

513.42529 513.42620 -1.77 2.5 C 29 H 57 O5 N 2

513.43022 -9.61 6.5 C H O

Orlistat

[M+NH4]+

Orlistat

[M+H]+

Conclusions• ID-CUBE on the Exactive Orbitrap platform provides extremely fast accurate mass

measurements (< 3 ppm with external calibration) to complete rapid screening in 10 seconds or less per sample for liquids and powders

• Optimization of the desorption settings with the ID-CUBE is 8x faster than with the DART-SVP source since sample heating and cooling is nearly instantaneous

• The listed Omega-3, -6, -9 fatty acids in several oil supplements were easily identified and relative levels per capsule were compared

• “Mexican sildenafil” tablet purchased on the street did indeed contain sildenafil, however dosage level is unknown

• Sibutramine was clearly detected in the banned Pai You Guo “herbal” slim capsule product from both the powder directly and from solution

• The Alli weight loss aid capsules purchased from an online vendor appeared to be legitimate and did not show any signs of gross contamination or adulteration

AcknowledgementsThe authors would like to kindly thank Maciej Bromirski at Thermo Fisher Scientific for

coordinating the collaborative efforts for the loan of an Exactive Orbitrap Mass Spectrometer.

Documents

Ten Second DART®-MS - IonSense€¢ Scan Settings: 1 µ-scan by 250 ms max inject time • AGC Target: Balanced (1e 6) ... • After pressure from FDA GMP Herbal Products, Inc. issued