macka talk pittcon 2016-v2016-03-06-given-watermarked

Gatherers and Foragers?

Analytical Scientists in the

Quest for Better Light Sources

Pittcon 2016, Atlanta, GA, USA

6-10 March 2016

Professor and Australian Research Council Future Fellow

Australian Centre for Research on Separation Science (ACROSS) and School of Chemistry,

University of Tasmania, Hobart, Australia

http://www.utas.edu.au/chem http://www.across.utas.edu.au

[email protected]

Mirek Macka

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http://www.utas.edu.au/chem


http://www.across.utas.edu.au/


mailto:[email protected]

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Pittcon

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6-10 March 2016





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Mirek Macka

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What is ’better’ ?

Depends on the purpose/use requirements

Light source properties, size, price, …

Wavelength(s)

Power

…so you may end up using:

‘BIG’: Synchrotron, nuclear reactor, …

‘Classical’: incandescent or discharge (W, W-hal., D2, Xe)

Some new ‘classical’ light sources

Solid state light sources: UV-vis-IR

New areas, strongest growth

Examples from own research Pittc

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Alternative VUV-vis light source?

IR laser driven light source: LDLS™ (Energetiq)

http://www.energetiq.com/index.php

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Solid state light sources

The light sources of the 21st century: SSLSs

SSLSs = LEDs + DLs

Synergies with new areas of strongest growth

Mobile technologies

Wearable technologies

Analysis

Platforms: Portable / mobile / remote

Areas:

Biomedical

Environmental

Food & agriculture

POC

on-site, in-field

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Mirek Macka

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Solid State Light Sources

Why SSLSs?

(Sydney skyline)

“It is expected that optics, also referred to as photonics, will surpass electronics in

the 21st century in terms of the size of the industry reliant on it.” http://www.sfi.ie/investments-achievements/research-showcase/shedding-light-on-many-subjects/

“In the next decade we will see a massive transformation of the lighting industry

towards energy efficient Solid State Lighting (SSL)”

http://www.photonics21.org/download/olae_sra.pdf

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http://www.sfi.ie/investments-achievements/research-showcase/shedding-light-on-many-subjects/
















- 10 -

SSLSs: ‘outside (analytical) chemistry’

They have many advantages

and still a few weaknesses

Benefits from large industries

(much larger then chemistry)

Consumer electronics

Lighting

IT & CT

LIFI

Medical

Automobile

Security & military

www.zoneray.com

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Looking ‘out of your box’

What is this?

iPad / Smartphone sterilisation device!

?

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Looking ‘out of your box’

Analytical science is small compared to defence +

‘bread + games’ industries = food, IT, medical...

Non-analytical usage of LEDs: 3D-agri- and aquaculture,

sterilisation

Vertical Farms, Silicon Chip, March 1012, pp.16-23 siliconchip.com.au

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http://www.siliconchip.com.au/

- 14 - 14

Solid state light sources

Why use solid state light sources (SSLSs) in science?

Conventional light sources

Incandescent, discharge lamps as light sources

SSLSs

Omnipresent, robust, inexpensive, miniaturisation compatible …

1880 2007

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SSLSs

LEDs

Laser diodes (LDs)

SLEDs

‘hybrid’ properties LED +LD

UV Vis NIR mid-IR

~ 200 nm - 7 µm

Electromagnetic spectrum

schematic representation

Light sources - typical properties:

Traditional: SSLS:

+++ Spectral coverage - + +

(deep-UV to NIR) (from 240 nm up)

+++ Mature well approved technology? - + +

(up to 200 years) (0 to ~40 years)

+ + - Luminosity - + -

- - - Energy conversion, heat production - ++

- - - Radiative heating +++

- - - Miniaturisation compatible? +++

- - - Robustness +++

- - - Life time +++

- - - $$$ +++

- + - Pulsed operation? +++

- + - Noise +++

- + - Future potential +++

0.01nm 1nm 100nm 1µm 1mm 1m 1km

!

Mirek Macka, Tomasz Piasecki Parmendu K Dasgupta, Light Emitting Diodes (LEDs) for Analytical Chemistry, Annual Review of Analytical Chemistry, 7, 183-207, 2014

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Dark

Red Gree

n Blue

i

"Full colour RGB" LED 621-419

maximum (half-width) nm

0

10

20

30

40

50

60

70

80

90

100

350 400 450 500 550 600 650 700 750

wavelength (nm)

rela

tiv

e in

ten

sit

y

Blue 438 (70) nm

Green 567 (25) nm

Red 627 (37) nm

SSLSs for analytical devices?

Advantages of LEDs

Small, reliable & robust => miniaturised & portable!

Low-cost: from <$1 to ~$50, but typically ~ $101

Long life-time: ~105 h & no catastrophic failure

Very low noise 10-5 AU

Used in various types of optical detectors

(HPLC, FIA etc.)

Can be operated in a pulsed regime

Can be pulsed

At extremely fast rates => TRF

Single-, bi- or tri-coloured LED’s available

Quasi-monochromatic: w(h/2) ~ 20-70 nm

‘Cold light’

5 5.1 5.2 5.3 5.4

Migration time (min)

0.1

mA

U

a

b

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SSLSs in analysis: history

Blood oximeter 1972

Cohen A, Wadswort N

Red/NIR light absorption

Pulsed operation

www.medical-monitors.com

Deoxydated hemoglobin

Oxydated hemoglobin

oximeter.holisticphysio.com

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SSLSs in analysis: history

Blood oximeter 1972

Cohen A, Wadswort N

Red/NIR light absorption

Pulsed operation

1980s: LEDs: IR, RGB

1990s: explosive growth

Analytical detection

FIA, LC, CE, chip

1990-1995

Trojanovicz

Cardwell, Cattrall & Scollary,

Huang, Dasgupta, Hauser, Yeung,

Worsfold

www.medical-monitors.com Publications (Web of Science)

0

500

1000

1500

2000

2500

3000

3500

1940 1950 1960 1970 1980 1990 2000 2010

Year

No

. o

f p

ub

lica

tio

ns

TS= multidisciplin*

TS= miniaturi*

TS= MEMS

TS= microfluidic*

TI= electrophoresis

TI= liquid* chromatography

TI= gas* chromatography

TI= light emitting diode*

E&CE

HPLC

GC

Macka M., Andersson P., Haddad P.R., Electrophoresis, 17(12), 1898-1905, 1996 Mirek Macka, Tomasz Piasecki Parmendu K Dasgupta, Light Emitting Diodes (LEDs) for Analytical Chemistry, Annual

Review of Analytical Chemistry, 7, 183-207, 2014

Deoxydated hemoglobin

Oxydated hemoglobin

oximeter.holisticphysio.com

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‘My’ areas: analytical, separation science

Solid State Light Sources & Miniaturised platforms

SSLSs:

LEDs,

DLs

Platforms:

miniaturised,

portable,

remote

White LEDs:

broad spectrum

light sources

UV LEDs:

portable

fluorescence

microscopy

UV-vis LEDs: photopolymerisations

Chip-CE,

Chip-ITP

Portable

medium-pressure LC

Micro- and small UAV platforms

MEDIA:

Capillaries, chips,

paper(fluidics)

10-6/32 mol/ L Fluorescein Solution

LEDs:

on-capillary detection

Paperfluidics-inspired

sample preparation

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My ARC FT


SSLSs:

LEDs,

DLs

Platforms:

miniaturised,

portable,

remote

White LEDs:

broad spectrum

light sources

UV LEDs:

portable

fluorescence

microscopy


Chip-CE,

Chip-ITP

Portable

medium-pressure LC


MEDIA:

Capillaries, chips,

paper(fluidics)


LEDs:



sample preparation

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On-capillary photometric detection

Practical considerations

In-house designes: simple, robust

Capillary alignment in a Agilent CE optical interface

Vis-LEDs

Johns, C., et al. Journal of Chromatography A , 927, 237-241, 2001 Mirek Macka, Tomasz Piasecki Parmendu K Dasgupta, Light Emitting Diodes (LEDs) for Analytical Chemistry, Annual

Review of Analytical Chemistry, 7, 183-207, 2014

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On-capillary detection with LEDs

UV LED 370 nm

Buffered chromate electrolyte

LODs ~10x lower vs. Hg 254 nm

UV LED 370

nm

UV LED 370

nm

0

0.2

0.4

0.6

0.8

1

1.2

1.4

1.6

1.8

2

210 260 310 360 410 460

Wavelength (nm)

Ab

so

rba

nc

e U

nit

s

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1

Rela

tiv

e I

nte

ns

ity

LED

379.5nmHg line

254nm

chromate

absorption

spectrum

0.3

0.8

1.3

1.8

2.3

2.8

3.3

1.95 2.45 2.95 3.45

Migration Time (mins)A

bs

orb

an

ce (

mA

U)

Cl-

NO3-

HCO3-

(b)

SO42- F

-

King M., Macka M., Paull B., Haddad P. R., Analyst, 127(12), 1564-1567, 2002.

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Photometric

Deep-UV-LEDs: 255 nm

Photometric detection $200-300, ~20-300 mW

EOF

GDP

ADPAMP

0 2 4 6 8 10 12 14

time (min)

abso

rban

ce

5 m

AU

Johns C. et al., Electrophoresis,

2004, 25, 3145–3152

Noise ~0.1 mAU

no optical components

light utilisation <1%

Stefan Schmid, Mirek Macka, Peter Hauser, UV-absorbance detector for HPLC based on a light-emitting diode, Analyst, 133, 465-469, 2008 (DOI 10.1039/b715681b)

Lenka Krcmova, Anna Stjernlof, Sebastien Mehlen, Peter Hauser, Silvija Abele, Brett Paull, Mirek Macka, Analyst, 134, 2394 – 2396, 2009 (DOI:10.1039/ B916081G)

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Photometric

Deep-UV-LEDs

Performance

Baseline noise N~0.1mAU

0

5

10

15

20

0 0.1 0.2 0.3 0.4 0.5 0.6

absorbance (AU)

sensitiv

ity

(L/m

ol)

0

500

1000

1500

2000

2500

3000

3500

200 300 400 500 600 700

wavelength (nm)re

lative in

ten

sity

257.0

Lenka Krcmova, Anna Stjernlof, Sebastien Mehlen, Peter Hauser, Silvija Abele, Brett Paull, Mirek Macka, Analyst, 134, 2394 – 2396, 2009 DOI:10.1039/ B916081G

Need for better deep-UV-LEDs!

‘suboptimal’ linearity

parasitic vis-band emission

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New generation deep-UV-LED

High optical output in ‘deep-UV’ @ 255 nm

Radiometric power 0.57 mW (before ~0.015 mW)

Negligible parasitic visible range emission

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1

200 300 400 500 600 700

Re

lati

ve

inte

ns

ity

Wavelength (nm)

255 nm LED

NEW (Optan255H)

OLD (UV TOP 255)

350 450 550 650

Dvorak M. et al., in preparation, 2014

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New generation deep-UV-LED

High optical output in ‘deep-UV’ @ 255 nm

Radiometric power 0.57 mW (before ~0.015 mW)

Negligible parasitic visible range emission

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1

200 300 400 500 600 700

Re

lati

ve

inte

ns

ity

Wavelength (nm)

255 nm LED

NEW (Optan255H)

OLD (UV TOP 255)

350 450 550 650

7.1 V

~5 V

6.1 V

Dvorak M. et al., in preparation, 2014

0

1000

2000

3000

4000

200 400 600 800

0

1000

2000

3000

4000

200 400 600 800

0

1000

2000

3000

4000

200 400 600 800

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‘Our’ areas: analytical, separation sci.


SSLSs:

LEDs,

DLs

Platforms:

miniaturised,

portable,

remote

White LEDs:

broad spectrum

light sources

UV LEDs:

portable

fluorescence

microscopy


Chip-CE,

Chip-ITP

Portable

medium-pressure LC


MEDIA:

Capillaries, chips,

paper(fluidics)


LEDs:



sample preparation

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Microfluidics has gadgets!

Microfluidic platform

Modular, flexible

Pressure driven flow

Programable multi-channel HV source

Synchronised microscope 1

www.labsmith.com

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http://www.labsmith.com/

- 29 -

Visualization of the separation

Synchronized video microscope for visualization

available as part of the microfluidic platform

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Detectors: LED (in-house) +

electrochemical (BVT + eDAQ)

Syringe pumps

(LabSmith) Switchable valves

(LabSmith)

Injector (VALCO)

4-20 nL

Capillary column: monolith

(C18 - Merck Chromolith)

1

2

4

3

Mobile phase A

Switching valves

A1

A2

B2

B1

B

A

Mobile phase B

Waste

Sample

Injector

Data

acquisition

Capillary column

Pressure sensor

LED detector

~$700

~$1,000

~$700

Design

~$200

ca 25 cm

Li Y., Dvorak M, Nesterenko P., Stanley R., Nuchtachvorn N., Kujovska Krcmova L, Aufarova J., Macka M, Anal. Chim. Acta, 896,

166-176, 2015

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Performance of the LC system

Isocratic Separations (ion-pair RP LC)

Column: 300 x 0.1 mm ID

Eluent: 50 mM ammonium acetate -

acetonitrile 50/50 (v/v)

F=0.5 µL/min.

Detection: LED on-capillary photometric

detector (254 nm)

Sample: 60 µM methyl 4-hydroxybenzoate

(MHP), 0.16 mM ethyl 4

hydroxybenzoate (EHB), 0.16 mM

propyl 4-hydroxybenzoate (PHB),

and 0.17 mM butyl 4

hydroxybenzoate (BHB)

RSD < 1% (peak area), efficiency N ~ 80,000 TP

Li Y., Dvorak M, Nesterenko P., Stanley R., Nuchtachvorn N., Kujovska Krcmova L, Aufarova J., Macka M, Miniaturised medium

pressure capillary liquid chromatography system with flexible open platform design using off-the-shelf microfluidic components,

Anal. Chim. Acta, 896(8), 166-176, 2015

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Gradient Separations: MeOH


Eluent: A=50 mM NH4AC (pH 5),

B=methanol, linear gradient 0 to

100% B in 8.3 min,

F=1 µL/min.

Detection:

355 nm LED on-capillary photometric detector

Sample: Dichromate 5 mM, tartrazine 1 mM,

orange G 1 mM, naphthol yellow 1 mM

and brilliant yellow 1 mM, 20 nL

Compounds Tr

(min)

RSD (n=8)

Tr

Peak area

(mAU*t)

RSD (n=8)

Peak area

K2Cr2O7 1.19 1% 77 3%

Tartrazine 2.86 5% 45 5%

Orange G 3.45 4% 29 6%

Naphthol Yellow 3.76 3% 23 5%

Brilliant Yellow 6.17 1% 79 4%

-10

-5

0

5

10

15

20

25

0 2 4 6 8 10 12

mA

u

Time (min)

Tartrazine

Cr2O72-

Orange G Naphthol Yellow

Brilliant Yellow

100 %

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Gradient Separations: MeCN

-10

-5

0

5

10

15

20

25

0 2 4 6 8

mA

u

Time (min)

Coumarin

60%

100%

Cr2O72-

Tartrazine

Naphthol Yellow

Brilliant Yellow Sudan III Sudan IV

Martius Yellow


Eluent: A=50 mM NH4AC (pH 5),

B=acetonitrile, linear gradient 0 to 60%

B for 5 min, then 100% B for 3 min.

F=1 µL/min.

Detection:

355 nm LED on-capillary photometric detector

Sample: dichromate 4 mM, tartrazine 1.5 mM,

naphthol yellow 1.5 mM, brilliant yellow

0.5 mM, martius yellow 2 mM, coumarin 1

mM, sudan III 0.5 mM and sudan IV 0.5

mM, 20 nL

Li Y., Dvorak M, Nesterenko P., Stanley R., Nuchtachvorn N., Kujovska Krcmova L, Aufarova J., Macka M, Miniaturised medium

pressure capillary liquid chromatography system with flexible open platform design using off-the-shelf microfluidic components,

Anal. Chim. Acta, 896(8), 166-176, 2015

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LED z-cell

Ball lens Ball lens

Silicon photodiode

Optics?

LED-z-cell

LED-z-cell photometric detector

Considerations

• Li Y. et al., unpublished results

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LED-z-cell

LED-z-cell photometric detector

Eluent

Slit

Tubular LED holder

defining distance

LED

Silica photodiode


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-0.1

0.1

0.3

0.5

0.7

0.9

1.1

0 2 4 6 8 10 12

Ab

so

rban

ce

(m

AU

)

Time (min)

LED detector

Commercial detector 45 nL

LED-z-cell

Comparison of separation performance (254 nm)

The gradient separation of genetic amines: L-Dopa 9 ppm, L-tyrosine 18 ppm, norfenefrine 14 ppm,

phenylephrine 18 ppm and tyramine 18 ppm, linear gradient 5 mM to 35 mM methanesulfonic acid

(MSG) 0 to 12 min, flow rate 12 µL min-1. Column: CS19, 20 cm x 200 µm i.d., injection volume: 400 nL.

-0.1

0.1

0.3

0.5

0.7

0.9

1.1

0 2 4 6 8 10 12

Ab

sorb

ance

(m

AU

)

Time (min)

LED detector

Commercial detector 180 nL


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On-capillary detection with LEDs

Quality of detection optical setup easily checked

Effective pathlength

Stray light %

Instrument Detector linearity Effective

upper limit (AU) pathlength (mm)

Agilent 3DCE 1.2 64.6

AB 270A-HT 0.75 60.5

Waters CIA 0.175 49.7 0

2

4

6

8

10

12

14

16

18

20

1 10 100 1000 10000

absorbance (mAU)

Se

nsit

ivit

y [

AU

·L·m

ol-

1]

Agilent 3D-CE

Applied Biosystems 270A-HT

Waters CIA

Johns C., Macka M., Haddad P.R., King M., Paull B., J. Chromatogr. A, 927(1-2), 237-241, 2001 Johns C., Macka M., Haddad P.R., LC-GC Europe, 16(5), 290, 292, 294-295, 2003

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Photometry and photometric detection

Quality of detection optical setup easily checked:

Sensitivity vs. absorbance graph

Effective pathlength

Stray light %

Linearity evaluation

0

500

1000

1500

2000

2500

0 0.1 0.2 0.3 0.4 0.5 0.6

Chromate [mol/L]

Ab

so

rba

nce

[m

AU

]

Agilent Technologies 3D CE

Applied Biosystems

Waters CIA

0

2

4

6

8

10

12

14

16

18

20

0.1 1 10 100 1000

Chromate [mmol/L]

Sen

sit

ivit

y [

AU

·L·m

ol-1

]

Agilent Technologies 3D CE

Applied Biosystems

Waters CIA

0

2

4

6

8

10

12

14

16

18

20

1 10 100 1000 10000

Absorbance [mAU]

Sen

sit

ivit

y [

AU

·L·m

ol-

1]

Agilent Technologies 3D-CE

Applied Biosystems

Waters CIA

Johns C., Macka M., Haddad P.R., King M., Paull B., J. Chromatogr. A, 927(1-2), 237-241, 2001 Johns C., Macka M., Haddad P.R., LC-GC Europe, 16(5), 290, 292, 294-295, 2003

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Beckm

an Instr

um

ents

Ag

ilen

t Te

cn

olo

gie

s

deltaD

OT

Pri

nce T

echnolo

gie

s

Sebia

Mic

ro T

ech*

Lum

ex

Unim

icro

Technolo

gie

s*

CE

Resourc

es**

0

10

20

30

40

50

60

Brand

We

igh

t [k

g]

0

0.05

0.1

0.15

0.2

0.25

0.3

Vo

lum

e [

m 3

]

W [kg] V [m3]

Portable platforms - still rare species…

Example: CE

Markéta Ryvolová, Jan Preisler, Dermot Brabazon, Mirek Macka, Portable capillary-based (non-chip) capillary electrophoresis: current state, instrumentation and future development, TRAC, 29(4), 339-353, 2010

Analytical instrumentation: Portability? Pi

ttcon

201

6, A

tlant

a, U

SA, 6

Mar

ch 2

016

Portable platforms - still rare species…

How portable is commercial instrumentation?

Example: CE

Markéta Ryvolová, Jan Preisler, Dermot Brabazon, Mirek Macka, Portable capillary-based (non-chip) capillary electrophoresis: current state, instrumentation and future development, TRAC, 29(4), 339-353, 2010

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- 41 -



SSLSs:

LEDs,

DLs

Platforms:

miniaturised,

portable,

remote

White LEDs:

broad spectrum

light sources

UV LEDs:

portable

fluorescence

microscopy


Chip-CE,

Chip-ITP

Portable

medium-pressure LC


MEDIA:

Capillaries, chips,

paper(fluidics)


LEDs:


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- 42 -

Research chips in a commercial chip-CE

Agilent Bioanalyzer 2010

DNA-chip vs. in-house-designed chips

DNA-chip In-house-chips

Ch

ip c

ad

die

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- 43 -


Chips: Complex challenges

Optical detection: Fluorescence primarily

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- 44 -


Details to be found in these publications:

1. Petr Smejkal, Ákos Szekrényes, Markéta Ryvolová, Frantisek Foret, András Guttman, Fritz Bek, Mirek Macka, Chip-based CE for rapid separation of 8-aminopyrene-1,3,6-trisulfonic acid (APTS) derivatized glycans, Electrophoresis, 31(22), 3783-3786, 2010, http://dx.doi.org/10.1002/elps.201000457

2. Nantana Nuchtavorn, Fritz Bek, Mirek Macka, Worapot Suntornsuk, Leena Suntornsuk, Rapid separations of Nile blue stained microorganisms as cationic charged species by chip-CE with LIF, Electrophoresis, 33 (9-10), 1421-1426, 2012, http://dx.doi.org/10.1002/elps.201100698

3. Nantana Nuchtavorn, Petr Smejkal, Michael C Breadmore, Philip Doble, Fritz Bek, Rosanne M Guijt, Frantisek Foret, Leena Suntornsuk, Mirek Macka, Exploring chip-CE-LIF platform flexibility: Separations of fluorescent dyes by chip-based non aqueous CE, J. Chromatogr.A, 1286, 216–221, 2013, http://dx.doi.org/10.1016/j.chroma.2013.02.060

4. Petr Smejkal, Michael C. Breadmore, Rosanne M. Guijt, Frantisek Foret, Fritz Bek, Mirek Macka, Isotachophoresis on a chip with indirect fluorescence detection as a field deployable system for analysis of carboxylic acids, Electrophoresis, 33(21), 3166-3172, 2012, http://dx.doi.org/10.1002/elps.201200141.R1

5. Petr Smejkal, Michael C. Breadmore, Rosanne M. Guijt, Jakub Grym, Frantisek Foret, Fritz Bek, Mirek Macka, Separation of carboxylic acids in human serum by isotachophoresis using a commercial field-deployable analytical platform combined with in-house glass microfluidic chips, Anal. Chim. Acta, 755, 115– 120, 2012, http://dx.doi.org/10.1016/j.aca.2012.10.022

6. Petr Smejkal, Michael C. Breadmore, Rosanne M. Guijt, Frantisek Foret, Fritz Bek, Mirek Macka, Analytical isotachophoresis of lactate in human serum using dry film photoresist microfluidic chips compatible with a commercially available field-deployable instrument platform, Anal. Chim. Acta, 803, 135–142, 2013, http://dx.doi.org/10.1016/j.aca.2013.01.046

7. Petr Smejkal, Danny Botteus, Michael C Breadmore, Rosanne M Guijt, Cornelius F Ivory, Frantisek Foret, Mirek Macka, Chip-ITP: a review, Electrophoresis, Special Issue Bioanalysis, 34(11), 1493-1509, 2013, http://dx.doi.org/10.1002/elps.201300021

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http://dx.doi.org/10.1002/elps.201000457












http://dx.doi.org/10.1016/j.chroma.2013.02.060










http://dx.doi.org/10.1002/elps.201200141.R1








http://dx.doi.org/10.1016/j.aca.2012.10.022
















- 45 -

Detection: Concurrent multi-detection

LED photometric on-capillary

250-660 nm, baseline noise ~0.0001 AU

Macka M., Andersson P., Haddad P.R., Linearity evaluation in absorbance detection: The use of light emitting diodes for on-

capillary detection in capillary electrophoresis, Electrophoresis, 17(12), 1898-1905, 1996

Johns C., Macka M., Paul R. Haddad, Design and performance of a light-emitting diode detector compatible with a

commercial capillary electrophoresis instrument, Electrophoresis, 25(18-19), 3145-3152, 2004,


Lenka Krcmova, Anna Stjernlof, Sebastien Mehlen, Peter Hauser, Silvija Abele, Brett Paull, Mirek Macka, Deep-UV LEDs in

photometric detection: A 255 nm LED on-capillary detector in capillary electrophoresis, Analyst, 134, 2394 – 2396, 2009,

http://dx.doi.org/10.1039/B916081G

Marketa Ryvolová, Jan Preisler, Pavel Krásenský, František Foret, Peter C. Hauser, Brett Paul, Mirek Macka, Single Point of

Detection Combined Contactless Conductometric, Photometric and Fluorimetric on-Capillary Detector for Capillary

Separation Methods, Anal.Chem., 82(1), 129-135, 2010, http://dx.doi.org/10.1021/ac902376v

C4D electrodes

capillary

Second fluorescence

excitation (400 mm OF)

Fluorescence emission

pick-up (300 mm OF)

Optical fiber (OF)

SMA adapter

PD detector

(optical fiber)

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http://dx.doi.org/10.1021/ac902376v







Triple detection C4D+PD+FD

Identical light source for PD and FD: 470 nm

Noise~<0.1mAU

1,24

1,25

1,26

1,27

1,28

1,29

1,3

0 2 4 6 8 10 12

Migration Time (min)

Sig

nal (

AU

)

PD

tartrazine

3,5

4

4,5

5

5,5

0 2 4 6 8 10 12


Sig

nal (

V)

C4D

CO32- MES

His

EOF

K+

tartrazine

28

33

38

43

48

53

0 2 4 6 8 10 12


Sig

nal (

mV

)

fluorescein FD

3-in-1 combined C4D + PD + FD

CONDITIONS:

Sample: standard mix

Capillary: 75 mm, 31.5/39 cm

BGE: 20 mM CHES buffer pH 9

Voltage : 12 kV, Injection: 7 cm,10 s

C4D: 100 kHz

PD: LED 470nm, no filter @ 30 mA (2.7 mW), 300 mm /50mm in/out fibres

FL: Ex.: LED 470nm, no filter @ 20 mA (2.7 mW),

300 mm /300 mm excit./pick-up fibre

Markéta Ryvolová, Jan Preisler, Pavel Krásenský, František Foret, Peter C. Hauser, Brett Paul, Mirek

Macka, Anal.Chem., 82(1), 129-135, 2010, DOI 10.1021/ac902376v

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Design

1 point of detection

Fibre optics

SMA adapter

C4D electrodes

capillary

Second fluorescence

excitation (400 mm OF)

Fluorescence emission

pick-up (300 mm OF)

Optical fiber (OF)

SMA adapter

PD detector

(optical fiber)

C4D

electrodes

Capillary

SMA adapter

(PD input & pick-up, FD

input)

Fluorescence pick-

up (optical fiber)

Detector assembly platform Detector assembly cover

A B

3-in-1 combined C4D + PD + FD

Markéta Ryvolová, Jan Preisler, Pavel Krásenský, František Foret, Peter C. Hauser, Brett Paul, Mirek

Macka, Anal.Chem., 82(1), 129-135, 2010, DOI 10.1021/ac902376v

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- 48 -

Rapid design using 3D-printing

CAD 3D-printing characterisation

Extrusion (Felix)

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- 49 -

LED-IF: on-capillary detection - comparison

3-in-1

High Brightness LED 460 nm

@ 700 mA, 3.1 lm, optical fibre 1 mm

~27 mW

LOD = 4 × 10-8 M LOD = 1 × 10-9 M Ultra bright 3mm LED 470 nm

@ 30 mA, 3.6 V, 5000 mcd, 35º

~ 2.7 mW

LEDs vs. lasers

LODs 10-10

M vs. 10-12

M

Prices ~ € 10 vs. ~ € 5,000

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- 50 -

LEDIF: µLEDs

LEDs - the smallest light sources ever

Micro LEDs with diameter ~15 μm

LED chip – 1 mm2

LED Cluster – 450 μm diameter

170 µLEDs/ 400 μm optical fibre

Integrated filters, micropackaged

15 mm

Tanriseven, S., Maaskant, P., Corbett, B., Applied Physics Letters, 92, 123501, 2008

Vaculovičová, M., Akther, M., Maaskant, P., Brabazon, D., Macka, M., Fibre coupled micro-light emitting diode array light source with integrated band-pass filter for fluorescence detection in miniaturised analytical systems, Anal.Chim.Acta, 871, 85-92, 2015

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- 51 -

Back metal Filter

n -

contact

Light generating

active area

Dielectric

p - contact

Semiconductor (GaN)

Bond metal

Optical fibre

SiO2/HfO

LEDIF: µLEDs

Microfabrication & micropackaging

Custom designed integrated interfrence filters

400 450 500 550 0.0

0.5

1.0

S16 (without filter)

S19 (with filter)

Norm

alis

ed c

ou

nts

Wavelength, nm

Normalised emission spectra

Tanriseven, S., Maaskant, P., Corbett, B., Applied Physics Letters, 92, 123501, 2008

Vaculovičová, M., Akther, M., Maaskant, P., Brabazon, D., Macka, M., Fibre coupled micro-light emitting diode array light source with integrated band-pass filter for fluorescence detection in miniaturised analytical systems, Anal.Chim.Acta, 871, 85-92, 2015

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- 52 -

LEDIF: µLEDs

LOD (fluorescein) = 5x10-9 mol/L @ ~50 µW

Outlook: SLEDs!

35

40

45

50

55

60

200 400 600 800 1000 1200 1400

Migration Time [s]

I F [

a.u

.]

Sample: Maltooligosaccharide ledder

(Diluted by H2O 1:10)

Capillary: 75 mm, 30/40 cm

BGE: 50 mM acetate pH 4.7 + 20% DNA gel

Voltage : -10 kV, Injection: 7 cm, 15 s

300 mm pick-up fibre

µLED Device: S 19 (with filter) @ 20 mA

capillary

mLED fibre mLED chip

pick-up fibre

to PMT

Vaculovičová, M., Akther, M., Maaskant, P., Brabazon, D., Macka, M., Fibre coupled micro-light emitting diode

array light source with integrated band-pass filter for fluorescence detection in miniaturised analytical

systems, Anal.Chim.Acta, 871, 85-92, 2015

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- 53 -



SSLSs:

LEDs,

DLs

Platforms:

miniaturised,

portable,

remote

White LEDs:

broad spectrum

light sources

UV LEDs:

portable

fluorescence

microscopy


Chip-CE,

Chip-ITP

Portable

medium-pressure LC


MEDIA:

Capillaries, chips,

paper(fluidics)


LEDs:



sample preparation

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- 54 -

LEDs as a broad-spectrum source?

UV-vis-NIR LEDs as a multi-chip

‘all-wavelengths-in-one’ LEDs

$?

White LEDs?

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1

200 300 400 500 600 700 800 900 1000 1100

Line I

nten

sity

Wavelength, nm

Normalized Emission Spectrum of UV-Vis-NIR LEDs355 nm390 nm415 nm458 nm490 nm540 nm570 nm595 nm625 nm660 nm690 nm720 nm750 nm780 nm820 nm850 nm880 nm920 nm980 nm

Wavelength [nm] 200 300 400 500 600

Lam

p C

ou

nts

0 2000 4000 6000 8000

10000 12000 14000 16000 18000

Deutrium Lamp

Luxeon LED

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- 55 -

LED as a broad-spectrum light source

White LED as a broad-spectrum light source ?

White phosphorus LED

Mounted on a

D2-lamp base

Tomasz Piasecki

Piasecki T., Breadmore M.C., Macka M., White LEDs as broad spectrum light sources for spectrophotometry: Demonstration in the visible spectrum range in a diode-array spectrophotometric detector, Electrophoresis, 31(22), 3737-3744, 2010 (DOI 10.1002/elps. 201000341)

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White LED as a broad-spectrum light source for a DAD

- photometric detection for CE

Luxeon LED powered to currents of 0-400 mA.

0 mA

50 mA

100 mA

200 mA

400 mA

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- 57 -


Spectra

Luxeon (colour-coded)

Deuterium lamp (black)

Wavelength [nm]

200 300 400 500 600

La

mp

Co

un

ts

0

2000

4000

6000

8000

10000

12000

14000

16000

18000

Deutrium Lamp

Luxeon LED

current: 50-400 mA

~5nm (thermal effect)

Time [min]

0 2 4 6 8

Ab

so

rban

ce [

mA

U]

0

2

4

6

8

10

Deuterium Lamp

Time (min) 0 2 4 6 8

Ab

so

rba

nc

e (

mA

U)

0

2

4

6

8

10

Luxeon LED

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- 58 -

White LED as a broad-spectrum source

Photometric detection in CE – D2 Lamp

Photometric detection in CE - LEDs

Better spectra quality for the LED lamp!

Only vis-range available

Wavelength [nm] 200 300 400 500 600

Ab

so

rban

ce (

mA

U)

0

2

4

6

8 FITC

Wavelength [nm] 200 300 400 500 600

0

2

4

6 F

Ab

so

rban

ce (

mA

U)

Wavelength [nm]

200 300 400 500 600 0

4

8

12

16

20

24 PR

Ab

so

rban

ce (

mA

U)

200 300 400 500 600 0

2

4

6

8 OG

Wavelength [nm]

Ab

so

rban

ce (

mA

U)

Wavelength (nm)

200 300 400 500 600

Ab

so

rban

ce (

mA

U)

0

2

4

6

8 FITC

Wavelength (nm) 200 300 40

0

500 600 0

2

4

6

8 F

Ab

so

rban

ce (

mA

U)

Wavelength (nm) 200 300 400 500 600

0

4

8

12

16

20 PR

Ab

so

rban

ce (

mA

U)

Wavelength (nm) 200 300 400 500 600

0

2

4

6 OG

Ab

so

rban

ce (

mA

U)

Piasecki T., Breadmore M.C., Macka M., White LEDs as broad spectrum light sources for spectrophotometry: Demonstration in the visible spectrum range in a diode-array spectrophotometric detector, Electrophoresis, 31(22), 3737-3744, 2010 (DOI 10.1002/elps. 201000341)

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- 59 -



SSLSs:

LEDs,

DLs

Platforms:

miniaturised,

portable,

remote

White LEDs:

broad spectrum

light sources

UV LEDs:

portable

fluorescence

microscopy


Chip-CE,

Chip-ITP

Portable

medium-pressure LC


MEDIA:

Capillaries, chips,

paper(fluidics)


LEDs:



sample preparation

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- 60 -

LED-Imaging

Microscopy

Portable

Low-cost

USB Microscopes

Dino-Lite http://www.bigc.com/

Cheap (~$50) Chinese

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http://www.bigc.com/

- 61 -

LED-Imaging

Microscopy

Portable

Low-cost

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- 62 -

LED-Imaging

Microscopy

Portable

Low-cost

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- 63 -

SSLSs: ‘non-analytical’

LEDs as light sources in many areas

Plant science

Wavelength-selective light

Biotechnology

Green algae

Photochemistry

Photolithography – chip microfabrication

e. g. Breadmore MC et al.

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- 64 - 64

Where use SSLSs?

LEDs, LDs, SLEDs

Optical detection

Fluorimetric

Photometric

Single-colour

White

Photoinitiated

polymerisations

of monoliths

Microphotochemistry

White LEDs:

broad spectrum

light sources

Single-colour or white LEDs:

photopolymerised monolith

LED-IF and diode LIF

Single-colour LEDs:

quasi-monochromatic

light sources

Single-colour LEDs:

photochemistry

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- 65 -

Where use SSLSs?

LEDs, LDs, SLEDs

Optical detection

Fluorimetric

Photometric

Single-colour

White

Microphotochemistry

Photoinitiated polymerisations

of monoliths

White LEDs:

broad spectrum

light sources

Single-colour or white LEDs:

photopolymerised monolith

LED-IF and diode LIF

Single-colour LEDs:

quasi-monochromatic

light sources

Single-colour LEDs:

photochemistry

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- 66 -

Photopolymerisations

LEDs as light sources

UV: 255 nm: Silvija Abele, Fu-Qiang Nie, František Foret, Brett Paull, Mirek

Macka, UV-LED photopolymerised monoliths, Analyst, 133, 864 - 866,

2008, DOI:10.1039/B802693A

Vis: 660 nm: Zarah Walsh, Silvija Abele, Brian Lawless, Dominik Heger,

Petr Klán, Michael C. Breadmore, Brett Paull, Mirek Macka, Photo-initiated

Polymerisation of Monolithic Stationary Phases Using Visible Region LEDs,

Chem. Commun., (48), 6504 – 6506, 2008, DOI:10.1039/B816958F

Vis: 470 nm: Zarah Walsh, Pavel A. Levkin, Brett Paull, Frantisek Svec and

Mirek Macka, Visible light initiated polymerisation of styrenic monolithic

stationary phases using 470 nm light emitting diodes, J.Sep.Sci., 33(1),

61-66, 2010, DOI:10.1002/jssc.200900624

UV: 365 nm: Silvija Abele, Smejkal Petr, Yavorska Oksana, Frantisek Foret,

Mirek Macka, Evanescent wave photoinitiated polymerization of open-

tubular capillary monolithic columns, Analyst, 135 (3), 477-481, 2010,

DOI:10.1039/b920789a

Monolith reviews:

Knob, R., Kulsing, C., Boysen, R.I., Macka, M., Hearn,

M.T.W., Surface-area expansion with monolithic open

tubular columns, TrAC, 67, 16-25, 2015

Zarah Walsh, Brett Paull, Mirek Macka, Inorganic

Monoliths in Separation Science: A Review, Anal. Chim.

Acta, 750, 28-47, 2012


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- 67 -





2008, DOI:10.1039/B802693A








61-66, 2010, DOI:10.1002/jssc.200900624




DOI:10.1039/b920789a

Monolith reviews:






Acta, 750, 28-47, 2012


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- 68 -

Abele S., Nie F.-Q., Foret F., Paull B., Macka M., Analyst, 132, 864 - 866, 2008

Monoliths: Synthesis with UV LEDs

Microfluidic chips – easily made in situ where needed

http://www.rsc.org/publishing/journals/AN/article.asp?doi=b802693a

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- 69 -





2008, DOI:10.1039/B802693A








61-66, 2010, DOI:10.1002/jssc.200900624




DOI:10.1039/b920789a

Monolith reviews:






Acta, 750, 28-47, 2012


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- 70 -

Red-LED photoinitiation in polyimide..

“Photoinitiated polymerisation of monolithic stationary

phases in polyimide coated capillaries using visible

region LEDs”

Zarah Walsh, Silvija Abele, Brian Lawless, Dominik Heger, Petr Klán, Michael C. Breadmore, Brett Paull, Mirek Macka, Photo-initiated Polymerisation of Monolithic Stationary Phases Using Visible Region LEDs, Chem. Commun., (48), 6504 – 6506, 2008, DOI: 10.1039/B816958F

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- 71 -


Modelling/simulating light penetration

Counter-intuitive patterns

10% 1% 0.1%

85% 75% 65%

Piasecki T., Brabazon D., Macka M., ISCC 2010, poster A.08., Thu-Fri 3-4 June

Piasecki, T., Macka, M., Paull, B., Brabazon, D, Numerical model for light propagation and light intensity distribution inside coated fused silica capillaries, Optics and Lasers in Engineering, 49 (7), 924-931, 2011

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http://www-scopus-com.ezproxy.utas.edu.au/record/display.uri?eid=2-s2.0-79957719046&origin=resultslist&sort=plf-f&src=s&sid=063B2EDA51157700B992F27F8010C0E9.mw4ft95QGjz1tIFG9A1uw:130&sot=autdocs&sdt=autdocs&sl=17&s=AU-ID(7004295072)&relpos=24&citeCnt=2&searchTerm=

http://www-scopus-com.ezproxy.utas.edu.au/record/display.uri?eid=2-s2.0-79957719046&origin=resultslist&sort=plf-f&src=s&sid=063B2EDA51157700B992F27F8010C0E9.mw4ft95QGjz1tIFG9A1uw:130&sot=autdocs&sdt=autdocs&sl=17&s=AU-ID(7004295072)&relpos=24&citeCnt=2&searchTerm=

- 72 -





2008, DOI:10.1039/B802693A








61-66, 2010, DOI:10.1002/jssc.200900624




DOI:10.1039/b920789a

Monolith reviews:






Acta, 750, 28-47, 2012


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- 73 -

Evanescent wave

photoinitiation (EWP)

EWP in transparent

PTFE-coated

fused silica

capillaries

Conditions

• initiator - DAP

• LED - 365 nm

• polymerisation time 15 min

Results

5 cm of capillary filled with polymer, but

top end is whiter, bottom end – paler

(polymer only around the wall proved by SEM)

Spectra not matched

Evanescent wave photoinitiation Pi

ttcon

201

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- 74 -

EWP

365 nm

LED

20 mm 0 mm 4.5

mm

8.5

mm

11.5

mm

15.5

mm

Evanescent wave photoinitiation

Silvija Abele, Smejkal Petr, Yavorska Oksana, Frantisek Foret, Mirek Macka, Evanescent wave photoinitiated polymerization of open-tubular capillary monolithic columns, Analyst, 135 (3), 477-481, 2010, DOI:10.1039/b920789a

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LEDs in photochemistry

LEDs in the photoswitching of photochromic molecules

MC-Co Complex Absorbance Data

0

0.2

0.4

0.6

0.8

1

1.2

1.4

1.6

1.8

2

300 400 500 600 700 800

Wavelength (nm)

Ab

so

rba

nce

0.00005M

0.0001M

0.0005M

0.001M

0.005M

0.01M

0.05M

0.1M

MC - no Co present

370nm

525nm

Walsh, Z., Scarmagnani, S., Benito-López, F., Abele, S., Nie, F.-Q., Slater, C., Byrne, R., Diamond, D., Paull, B., Macka, M., Photochromic spiropyran monolithic polymers: Molecular photo-controllable electroosmotic pumps for micro-fluidic devices, Sensors and Actuators, B: Chemical, 148 (2), 569-576, 2010

Scarmagnani, S., Walsh, Z., Slater, C., Alhashimy, N., Paull, B., Macka, M., Diamond, D., Polystyrene bead-based system for optical sensing using spiropyran photoswitches, J.Mater.Chem., 18 (42), 5063-5071, 2008

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Future?

Every

perspective

is personal

and

relative…

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LEDs: Deep-UV?

From ~0.01 mW to 1-3 mW

HIC

SU

NT

LE

ON

ES

0.01

0.1

1

10

100

1000

200 300 400 500 600 700

Wavelenght [nm]

Ra

dio

me

tric

po

we

r [m

W]

€0.00

€100.00

€200.00

€300.00

€400.00

€500.00

€600.00

€700.00

€800.00

Pri

ce

[€

]

Single chip LED power emitters

Multichip LED high power emitters

Single chip LED price

Multichip LED price

Mirek Macka, Tomasz Piasecki Parmendu K Dasgupta, Light Emitting Diodes (LEDs) for Analytical Chemistry, Annual Review of Analytical Chemistry, 7, 183-207, 2014

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Future of SSLSs in science

SSLSs: Deep-UV?

The “Alloy Road” to deep-UV-LEDs: AlN

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Future of SSLSs in science

LEDs: Spectra and prices now and into the future

Emission spectra for LEDs and LDs

0

0.5

1

200 300 400 500 600 700 800

Wavelength (nm)

Re

lati

ve

In

ten

sit

y

256 nm

281 nm

370nm

401 nm

473 nm

523 nm

Laser 532 nm

634 nm

660 nm

$

101

102

100

$

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SSLSs: New ‘cool gadgets’…

High power low cost lasers

http://www.wickedlasers.com/

UV-LED torches

395 nm http://au.element14.com/night-searcher/nsnuvled395/torch-led-uv/dp/1823957?Ntt=1823957&CMP=i-55c5-00001402

365 nm, 395 nm

5W (electrical input)

http://www.farnell.com/datasheets/606066.pdf

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http://www.wickedlasers.com/

http://au.element14.com/night-searcher/nsnuvled395/torch-led-uv/dp/1823957?Ntt=1823957&CMP=i-55c5-00001402
























- 81 -

SSLSs: New ‘cool gadgets’…

Applications: photosynthesis – plant growth

Spectrum needed:

Chlorophyl spectrum

http://en.wikipedia.org/wiki/Chlorophyl

Red + blue

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http://en.wikipedia.org/wiki/Chlorophyl

- 82 -

Future?

Light sources SSLSs

LEDs

Deep-UV (<300 nm)

Higher power! (W) + ‘clean’ spectra

VUV ? (<180 nm) ???

MIR (2-7 um)

LDs

Parallel the success of LEDs

‘Specialties’ e.g. QCLs (IR)

New areas, strongest growth

Vacuum UV (VUV), deep-UV, IR light sources

Portability

Imaging – all wavelengths

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Acknowledgements

SPECIAL THANKS TO:

$ Australian Research Council (ARC):

Future Fellowship Level 3 (Professorial)

$ Australian Endeavour Fellowship

$ University of Tasmania, Hobart, Australia,

ACROSS & School of Physical Sciences

$ Agilent, Waldbronn, Germany

University Relations Grant

$ Royal Jubilee Scholarship,

Mahidol University, Thailand

$ Grant Agency of the Czech Republic

OTHER COLLABORATORS :

Companies: LabSmith, USA; Knauer AG, Germany

Academic: Prof Leena Suntornsuk, Mahidol University, Bangkok, Thailand

…….

AU$

€

CZK

THB

AU$

AU$

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UTAS: PhD scholarship available

Tasmania: Beautiful & mild climate

Follow Charles Darwin

Numerical modelling

Thank you!

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Thank you!

The presentation available at

https://www.researchgate.net/profile/Mirek_Macka

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https://www.researchgate.net/profile/Mirek_Macka

macka talk pittcon 2016-v2016-03-06-given-watermarked

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