AGENDA

Part I:
Screening of Selected pharmaceuticals and
Personal Care Products (PPCPs) in Missouri Source
and drinking waters by using LC-MS/MS

Dr. Yinfa Ma
Missouri University
Science & Technology

- Why do we care about PPCPs in the drinking water?

- Water resources
- Is Milli-Q water free from PPCPs?
- Conclusions

Part II:
Potential effect of high purity water on
environmental analysis
- Role of water in analytical chemistry
- Effect of water contaminants on LC-MS/MS
- Importance of TOC of ultrapure water on HPLC analyses
- Milli-Q water for ICP-MS analyses
- Conclusions

Dr. Maricar Tarun-Dube

Guest Speaker: Dr. Yinfa Ma

Curators Teaching Professor, Missouri University of Science and Technology

Biography
Yinfa Ma received his BS degree in chemistry at December 1981 at
Zhengzhou University in China. Ma received his Ph.D. in analytical chemistry
and minor Ph.D. in biochemistry in December 1990 from Iowa State
University. Ma has serviced as chemistry faculty at both Truman State
University (1990-2000) and Missouri University of Science and Technology
(2000-present).

Ma serves on the scientific committee since APCE was first initiated in 1996. He also serves on the
science committee for Chinese Capillary Electrophoresis (CCE) since 1993. Ma has received over 30
prestige awards for his teaching and research and was named Curator's Teaching Professor in
December 15, 2007 at Missouri University of Science and Technology.
He is currently associate editor for Global Journal of Analytical Chemistry. He has served as a guest
editor for Electrophoresis journal in 1998 and 2002.
Ma has published 105 peer-reviewed journal papers and 24 book chapters, gave 233 presentations
at national and international conferences, delivered 57 invited seminars, and filled 10 patents.

Study of Selected Pharmaceuticals and Personal

Care Products (PPCPs) in Missouri Source and
Drinking Waters
by Using LC-MS/MS
Yinfa Ma
Department of Chemistry
Missouri University of Science and Technology

http://www.sciencedirect.com/science/article/pii/S0043135410008237

How safe do you feel about our drinking water?

Algal Toxins?

Degradates?

Drug waste?

Degradates?

Where are these chemicals possibly traveling to?

These compounds may be excreted

unmetabolized, thereby leading to a wide
variety of drugs reaching water treatment
plants and the eventually humans.

The presence of pharmaceuticals in natural water systems and

drinking water supplies has raised public concerns because of

their potential adverse ecological effects on aquatic organism or
even on human health.

CCL 2 List

 The screening of PPCPs in source and drinking waters

is crucial in providing information for the disinfection
strategy in water treatment facilities.

Are there any PPCPs in

Missouri Water Resources?

PPCPs ---- Pharmaceuticals and Personal Care Products

Selection of compounds in this study:

Compounds
erythromycin
lincomycin
trimethoprim
sulfamethoxazole
ethynylestradiol
estradiol
acetaminophen
caffeine
carbamazepine
codeine
triclosan
ibuprofen
estrone
estriol
iopromide
clofibric acid

Formalua
C37H67NO13
C18H34N2O6S
C14H18N4O3
C10H11N3O3S
C20H24O2
C18H24O2
C8H9NO2
C8H10N4O2
C15H12N2O
C18H21NO3
C12H7Cl3O2
C13H18O2
C18H22O2
C18H24O3
C18H24I3N3O8
C10H11ClO3

CAS #
114-07-8
154-21-2
738-70-5
723-46-6
57-63-6
50-28-2
103-90-2
58-08-2
298-46-4
76-57-3
3380-34-5
15687-27-1
53-16-7
50-27-1
107793-72-6
882-09-07

MW
733.93
406.5
290.3
253.3
296.4
272.4
151.2
194.2
236.3
299.4
289.5
206.3
270.4
288.4
791.1
214.7

Class
antibiotics
antibiotics
antibiotics
antibiotics
hormone
hormone
analgesics
Stimulant
anticonvulsant
analgesics
antibiotics
analgesics
hormone
hormone
X-ray contrast media
lipid regulator

Mississippi river water

Missouri river water
Reservoir water
Lake water
Well water

Our work horse

Water purification
system used to provide
deionized water for
UPLC eluent, blank,
standard and sample
solution preparation:
Merck Millipore Milli-Q
Advantage fed by an Elix
water purification system

Polarity and precursor/product ion transitions

Compounds
erythromycin
lincomycin
trimethoprim
sulfamethoxazole
acetaminophen
codeine
carbamazepine
caffeine
ethynylestradiol
iopromide
clofibric acid
triclosan
ibuprofen
estrone
estriol
estradiol

Polarity
positive
positive
positive
positive
positive
positive
positive
positive
negative
negative
negative
negative
negative
negative
negative
positive

Q1
734.4
407.1
291.0
253.9
151.8
300
236.9
194.9
295.0
789.9
212.9
286.7
204.9
268.9
286.7
271.0

Quantification ion
158.1
126.2
230.2
156.0
110.0
215.0
194.0
138.0
145.1
126.7
126.8
35.0
159.0
144.8
171.1
144.8

Confirmer ion
576.4
359.1
123.1
108.1
65.1
165.1
193
110.1
142.9
NA
84.9
NA
161.1
143.1
144.9
142.9

Chromatographic conditions:
Column: Supelco C-18 1502.1 mm;
Flow rate: 0.25 mL/min;
Injection volume: 10 l.
Positive ionization mode:
Mobile phase A: 0.1% formic acid in Milli-Q H2O;
Mobile phase B: 0.1% formic acid in acetonitrile.
Negative ionization mode:
Mobile phase A: 5mM ammonium acetate in Milli-Q H2O;
Mobile phase B: acetonitrile

XIC of +MRM (22 pairs): 194.9/138.0 amu from Sample 29 (CS 5) of Curve of 15PPCPs on Supelco@07Dec@06Oct.wiff (Turbo Spray), ...

9.0e4

8.5e4
8.0e4
7.5e4
7.0e4
6.5e4

In te n s ity , c p s

6.0e4
5.5e4

8.24
5.0e4

4.5e4

4.0e4

Max. 5.2e4 cps.

acetaminophen

2
3
4

codeine
lincomycin
caffeine

5
6
7
8
9
10

trimethoprim
sulfamethoxazole
tylosin
carbamazepine
estradiol
ethynylestradiol

3.5e4
3.0e4

10

2.5e4

2.0e4

1.5e4

1.0e4
5000.0
0.0

10

11

12
13
Time, min

14

15

16

17

18

19

20

21

22

23

24

Total ion chromatogram of selected PPCPs at positive ionization mode.

XIC of -MRM (12 pairs): 789.9/126.7 amu from Sample 3 (Neg Compounds) of PPCPs.wiff (Turbo Spray), Smoothed, Smoothed, Smoothe...

4.2e4
4.0e4

3.8e4
3.6e4
3.4e4
3.2e4

3.0e4

Max. 1.1e4 cps.

iopromide

clofibric acid

estriol

ibuprofen

estrone

triclosan

2.8e4

In te n s ity , c p s

2.6e4

2.4e4

2.2e4
2.0e4
1.8e4
1.6e4

1.4e4
1.2e4

3.49

1.0e4

8000.0
6000.0
4000.0
2000.0
0.0

10

11

12
13
Time, min

14

15

16

17

18

19

20

21

22

23

Total ion chromatogram of selected PPCPs at negative ionization mode.

Is Milli-Q water free from PPCPs ?

Since we are dealing with parts per trillion (PPT) levels
of PPCPs in water samples, it is crucial that the purified
water from the Milli-Q water is free from any PPCPs we
are investigating, even before we started our real water
screening.

TIC of +Q1: from Sample 6 (MQ) of MQ 7 day old.wiff (Turbo Spray)

Max. 8.1e6 cps.

TIC of -Q1: from Sample 7 (JTB-H45 4L ) of JTB-H45 4L 1day old.wiff (Turb...

1.06

8.0e6

7.2e7
7.0e7

3.25

Max. 7.2e7 cps.

1.65
0.96

7.5e6
6.5e7
7.0e6
6.0e7
6.5e6
5.5e7

6.0e6

5.0e7

5.0e6

In te n s ity , c p s

In te n s ity , c p s

5.5e6

4.5e6
4.0e6
3.5e6
3.0e6

4.5e7
4.0e7
3.5e7
3.0e7
2.5e7

2.5e6

Q1 scan of Milli-Q water, positive mode

2.0e6

2.0e7
1.5e7

1.5e6

1.0e7

1.0e6

5.0e6

5.0e5
0.0

0.5

1.0

1.5

2.0

2.5
Time, min

3.0

3.5

4.0

-Q1: 2.494 min from Sample 3 (MQ) of MQ 7 day old.wiff (Turbo Spray)

0.0

4.5

Max. 3.5e6 cps.

1.5

2.0

2.5
Time, min

3.0

3.5

4.0

4.5

Max. 3.9e6 cps.

3.8e6

3.2e6

3.6e6

3.0e6

3.4e6

2.8e6

3.2e6

2.6e6

3.0e6

2.4e6

2.8e6
2.6e6

In te n s ity , c p s

2.2e6
2.0e6
1.8e6
1.6e6

Sample mass spectrum of

water at positive mode

1.4e6
1.2e6
1.0e6

Milli-Q

2.4e6
2.2e6
2.0e6
1.8e6

Sample mass spectrum of Milli-Q

water at negative mode

1.6e6
1.4e6
1.2e6

8.0e5

1.0e6

6.0e5

8.0e5
6.0e5

4.0e5

92.8

100

4.0e5

312.6 337.1

94.6
198.4

50

1.0

60.1

3.4e6

2.0e5

0.5

-Q1: 2.819 min from Sample 8 (MQ) of MQ fresh 6.wiff (Turbo Spray)

59.5

In te n s ity , c p s

Q1 scan of Milli-Q water, negative mode

150

218.9

200

283.3

250

300

363.9

399.5

350
400
m/z, amu

440.4

450

493.1

500

528.7 557.2

550

587.8 610.5

600

2.0e5
650

700

268.2
58.8
83.4
63.5

50

120.3

100

146.1 155.3 165.7

150

218.9

200

236.1 288.8

250

318.4

339.3 361.7
373.0

300

350
400
m/z, amu

420.6442.3

450

486.0

501.4

500

584.6

550

619.2 630.0

600

650

679.7

700

How about the Milli-Q water after Solid phase

extraction (SPE)?
For PPCP screening in source and drinking
water, SPE procedure must be used to determine
PPCPs in the ppt levels.
Question: does Milli-Q water from Elix -3
contribute any of the PPCPs we are interested
in?

Water quality comparison

Extracted chromatogram of
Milli-Q water at positive
mode
Extracted
chromatogram of LCMS/MS grade water at
positive mode

After 500x concentration of Milli-Q

water and LC-MS/MS grade bottle
water by using SPE (EPA method)

 After a series of experiments, results

showed that the Milli-Q water purity is
excellent for our PPCP study.

Method Detection Limit of PPCPs

Compound

MDL/ppt in
MQ water

SD

MDL /ppt in
Rolla Tap water

SD

Tylosin

0.27

0.09

0.17

0.06

Lincomycin

0.12

0.04

0.11

0.04

Trimethoprim

0.32

0.11

0.37

0.13

Sulfamethoxazole

0.39

0.13

0.25

0.09

Ethynylestradiol

0.14

0.05

0.48

0.17

Estradiol

0.78

0.27

1.23

0.42

Acetaminophen

2.69

0.93

1.37

0.47

Caffeine

0.78

0.27

1.10

0.38

Carbamazepine

0.47

0.16

0.21

0.07

Codeine

0.98

0.34

1.45

0.48

Triclosan

0.99

0.34

1.18

0.39

Ibuprofen

1.02

0.35

1.63

0.54

Iopromide

3.49

1.20

9.89

3.42

Clofibric acid

1.27

0.44

0.61

0.21

Estriol

4.33

1.49

5.15

1.78

Estrone

1.35

0.46

1.00

0.34

Concentration of PPCPs in untreated water samples (ng/L)

100
90

80
70
60
50
40

River

30

Surface water

20

Deep well

10

Unconsolidated well and ground water

Concentration of PPCPs in treated water samples (ng/L)

70
60
50
40
30
20

10
0

River
Surface water
Deep well
Unconsolidated well and ground water

Concentration of PPCPs in Mississippi river in untreated and

treated water samples (ng/L) from Feb - May
50
45
40

Mississippi River Feb-Raw

35

Mississippi River Apr-Raw

Mississippi River May-Raw

30

Mississippi River Feb-Fin

25
20

Mississippi River Apr-Fin

Mississippi River May-Fin

15
10
5
0

Mississippi River May-Fin

Mississippi River Apr-Fin
Mississippi River Feb-Fin
Mississippi River May-Raw
Mississippi River Apr-Raw
Mississippi River Feb-Raw

Fig 4, The PPCPs levels of Mississippi river in 3 different months.

Concentration of PPCPs in Missouri river in untreated and treated

water samples (ng/L) from Feb - May
50
45
40
35
30
Missouri River Feb-Raw

25

Missouri River Apr-Raw

Missouri River May-Raw

20

Missouri River Feb-Fin

15

Missouri River Apr-Fin

10

5
0

Missouri River May-Fin

Missouri River May-Fin
Missouri River Apr-Fin
Missouri River Feb-Fin
Missouri River May-Raw
Missouri River Apr-Raw
Missouri River Feb-Raw

Fig 4, The PPCPs levels of Missouri river in 3 different months.

Conclusions
Some selected PPCPs have been found in Missouri source and treated water samples.

The levels of PPCPs are water source dependent. There were higher levels of caffeine
and other PPCP compounds in surface water than those in the well water.
Most of the PPCPs are not detectable after the water treatment. Even though some PPCPs
can be detectable after treatment, the concentrations are much lower than the untreated
water.
The quality of deionized water is very crucial for this study because it was used as eluent
for UPLC, blank checking (QA/QC purpose), and preparation of standard PPCPs and
samples preparation.

The water quality from Milli-Q water purification system is excellent for PPCP study in
source water and drinking water samples.

Guest Speaker: Dr. Maricar Tarun-Dube

Biography
Maricar Tarun-Dube received her PhD in chemistry from the University of
Connecticut in 2006. She worked on the LC-MS/MS analysis of DNA adducts
formed on DNA-polyion films upon incubation with organic chemicals.
Maricar joined Millipore Corporation (now Merck Millipore) after graduation, as
an Applications Scientist in the Lab Water business. At Millipore, she used her
experience and knowledge in LC-MS/MS to support the sales and marketing
organizations.
Maricar is currently a global speaker and freelance scientific writer and expert
in Life Sciences and Lab Water industries.

37

The benefits of using high purity water

on environmental analyses

The role of water in analytical chemistry

Mobile phase
Liquid Chromatography

Blanks

Preparation of standards

Analytical Method

Washing, rinsing
of sample & reagent
containers

Sample
Preparation
- extraction
- dilution
- rinsing

Avoid interference with measured analytes

Optimize analytical instrument operation

39

Contaminants

Na+
2+
NO3- Ba

WATER
bacteria

Cl-

40

The benefits of using high purity water

on LC-MS/MS

Effects of water contaminants on LC-MS/MS:

Particles, bacteria, ions
Particles
Increase column backpressure - decrease column lifetime,
poor resolution and peak shape, poor reproducibility
Damage pump and injector

Could contaminate the ion source component of the MS

Bacteria
Behave as particulates
Release of organic by-products resulting in extraneous peaks
and unstable baselines
Cl-

2+
NO3- Ba
42

Ions
Metal ions, especially Na+, K+ form adduct peaks

Using Milli-Q water gives cleaner mass spectrum

Bradykinin m/z 757

Na adduct peaks
Additional peaks

96% fresh ultrapure water : 4% ACN

96% fresh ultrapure water with 1 ppm NaCl : 4% ACN

43

Effects of water contaminants on LC-MS/MS:

Organics
Noisy, drifting background
Appearance of spurious/contaminant peaks
Distorted peak shapes
Shift in retention time
Presence of measured analytes in ultrapure water could compromise the
quality of data

44

Experiments designed to show suitability of

Milli-Q water for LC-MS/MS

Tap water
(hormones,
pesticides)

Milli-Q water
purification

Ultrapure
water

Spain, France,
China, India,
Finland

Study 1. Hormones in tap water and MQ water by LC-MS/MS

Study 2. Pesticide in tap water and Milli-Q water by LC-MS( (preliminary investigation)

45

Significant removal of hormones by Milli-Q

x102
1.1
1

Tap water France

0.9
0.8

France tap

0.7

Spain Tap

Tap water Spain

Milli-Q water France

0.6
0.5

Milli-Q water Spain

0.4
0.3
0.2
0.1

France Milli-Q
Spain Milli-Q

-0.1
6.2 6.226.246.266.286.3 6.326.346.366.386.4 6.426.446.466.486.5 6.526.546.566.586.6 6.626.646.66
Counts (%) vs. Acquisition Time (min)

MRM chromatogram of estradiol (273 255)

46

Significant removal of hormones in Milli-Q

47

Hormone

Tap Water

Milli-Q
water

Estadiol

Detected

ND

Androsterone

Detected

ND

Corticosterone

Detected

ND

Significant removal of pesticides*

x102 +/-ESI EIC(254.3530) Scan Frag=135.0V EDS.d
1
0.9

255.48 g/mol

0.8

Milli-Q water Spain

0.7
0.6
0.5
0.4

Tap Spain
Milli-Q Spain

0.3
0.2
0.1
0

-0.1
9.4

9.5

9.6

9.7

9.8

9.9
10
10.1
10.2
10.3
10.4
Counts (%) vs. Acquisition Time (min)

10.5

10.6

10.7

EIC of 2,4,5-Trichlorophenoxyacetic acid (2,4,5-T) in

tap and Milli-Q in the Spain laboratory
*Preliminary investigation
48

Tap water Spain

Significant removal of pesticides*

Pesticide

Tap Water

Milli-Q water

Atrazine

Detected

ND

Simazine

Detected

ND

Cyanazine

Detected

ND

2,4,5-T

Detected

ND

*Preliminary investigation

49

Lower levels of TOC in ultrapure result to

cleaner chromatograms

20 ppb
13 ppb
9 ppb
5 ppb
2 ppb

Gradient profiles of ultrapure water with different TOC levels (214 nm)
50

High TOC levels in ultrapure water is not

suitable for trace analyses
x10 2 +ESI EIC(202.0000) Scan Frag=135.0V IndiaMQ_scan+
1.1 1
1

TOC monitor displays 48 ppb

0.9
0.8
0.7
0.6

0.5

Tap India Simazine

Milli-Q India

0.4

0.3
0.2

0.1
0
-0.1
0.5 1 1.5 2 2.5 3 3.5 4 4.5 5 5.5 6 6.5 7 7.5 8 8.5 9 9.5 10 10.511 11.512 12.5
Counts (%) vs. Acquisition Time (min)

EIC of m/z 202 (simazine) in water samples from India (TOC level was
above the specification of < 5 ppb)
51

The benefits of using high purity water

on ICP-MS

Effects of water contaminants on ICP-MS analysis

Particles

Could clog sample line tubing and nebulizer

Bacteria and by-products

Act like particulates
By-products include ions and organics
Organics
Instrument contamination
Organo-metallic species

Na+
Cl-

53

2+
NO3- Ba

Ions
Inaccurate measurement of analyte

Milli-Q water for ICP-MS using the Q-POD

Element showed very low BEC and detection
limits
Element

Isotope

BEC [ppt]

DL[ppt]

Cr

52

< DL

0.03

0.94

Co

59

< DL

0.01

0.04

Ni

60

< DL

0.10

0.99

Cu

65

< DL

0.03

5.32

Cd

111

< DL

N.D.

0.13

Pb

208

< DL

N.D.

0.22

Ultrapure water source: Milli-Q Integral + QPOD Element

Instrument: ICP-MS Agilent 7700

Consistency of Milli-Q water quality

40

Cd-111
Cr-52
Cu-63
Ni-60
Pb-208
Co-59

30
20
10
0

Date of analysis
Ultrapure water source: Milli-Q Integral 5
Instrument: Perkin Elmer Elan DRCII
55

Apr-13

Feb-13
Mar-13

Jan-13

Dec-12

Nov-12

Oct-12

Sep-12

Aug-12

Jul-12

Jun-12

May-12

Apr-12

Feb-12
Mar-12

Jan-12

Dec-11

Nov-11

Oct-11

Sep-11

Aug-11

Jul-11

Jun-11

-10

May-11

Concentration (ppt)

50

Water purification: The right combination of

technologies is critical

850 ppb
300-700 S/cm

1000 ppb
300-700 S/cm

56

50-100 ppb
10S/cm

30 ppb
5 Mcm
30 ppb
5 Mcm

50 ppb
1Mcm
5 ppb
18.2 Mcm

Online monitoring of organic and ionic purity of

ultrapure water

Ionic purity
(Resistivity)

Organic level
(TOC)

57

A global approach to water purification

according needs of the laboratory
Water quality and quantity

Monitoring quality parameters

Reporting
Validation

Maintenance of the purification unit

Training of the personnel

Millitrack

58

Summary and Conclusions

Milli-Q efficiently removed the hormones and pesticides that were
detected in the tap water feed.
Extremely low BEC and detection limits were obtained from ultrapure
water delivered from a Q-POD Element, under clean room conditions.
The levels of heavy metals in ultrapure water from a Milli-Q system were
consistently low over a 22-month monitoring period.
Milli-Q systems meet the stringent requirement of trace LC-MS/MS and
ICP-MS analyses for the highest purity water.

59

Acknowledgments
We would like to thank Univeristy of Helsinki, Finland for their help and support, in particular:
Mikael Tilli
Jevgeni Parshintsev
Matias Kopperi
Raymundo Lizcano
Marja-Liisa Riekkola
As well as our colleagues from Merck KGaA, Darmstadt, Germany, in particular:
Fernando Cabaleiro-Nogueira
Ingrid Heiderstaedt
Evelyn Trueller
Marcus Claassen
Matthias Ohm

And our colleagues from Merck Millipore, in particular:

Anastasia Khvataeva-Domanov
Jeffrey Lei
Gora Sharangi
Vasanthy Rajamannar
Juan Carmona
Patrik Appelblad
60

Thank you

61