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Constantly EvolvingLeon Blumberg reveals his role in the
development of separation science
2 Constantly Evolving Leon Blumberg has been at the cutting edge of chromatography since
the late 1980s. The Column spoke to him about his contribution to the past, present, and future of chromatography.
Cover Story
Features
21 Preview of Topics at HPLC 2016, Part 4: State-of-the-Art MS Methods for Structural Assessment of mAbs and ADCs: From the Research Lab to Routine Characterization Alain Beck1 and Arnaud Delobel2, 1Centre d’Immunologie Pierre Fabre (CIPF),
2Quality Assistance SA This is the fi nal instalment in a series of four articles exploring topics that will be
addressed at the HPLC 2016 conference in San Francisco, USA, from 19–24 June 2016.
18 Making Method Development Faster for the Analysis of Natural and Artificial Flavourings Philipp Jochems and Gesa Schad, Shimadzu Europa A simple, rapid, and robust ultrahigh-performance liquid chromatography (UHPLC)
method for the simultaneous determination of natural and artifi cial vanilla fl avouring substances as well as some precursors has been developed using an automated method scouting or method optimization workfl ow.
Regulars9 News The latest company news, peaks of the week, and news in brief are featured in this issue.
12 Tips & Tricks GPC/SEC: Branching Analysis Daniela Held, Peter Montag, and Wolfgang Radke, PSS Polymer Standards
Service GmbH Branching is one of the parameters chemists can adjust to produce polymer
materials with optimized physical properties. Chromatography and advanced detection can help to characterize branched molecules. This instalment of Tips & Tricks explains more.
25 The 31st International Symposium on Chromatography (ISC 2016) A preview of the upcoming 31st International Symposium on Chromatography
(ISC 2016), which is due to be held 28 August–1 September at University College
Cork, Ireland.
27 Training Courses and Events
28 Staff
7 June 2016 Volume 12 Issue 10
Constantly EvolvingLeon Blumberg reveals his role in the
development of separation science
2 Constantly Evolving Leon Blumberg has been at the cutting edge of chromatography since
the late 1980s. The Column spoke to him about his contribution to the past, present, and future of chromatography.
Cover Story
Features
21 Preview of Topics at HPLC 2016, Part 4: State-of-the-Art MS Methods for Structural Assessment of mAbs and ADCs: From the Research Lab to Routine Characterization Alain Beck1 and Arnaud Delobel2, 1Centre d’Immunologie Pierre Fabre (CIPF),
2Quality Assistance SA This is the fi nal instalment in a series of four articles exploring topics that will be
addressed at the HPLC 2016 conference in San Francisco, USA, from 19–24 June 2016.
18 Making Method Development Faster for the Analysis of Natural and Artificial Flavourings Philipp Jochems and Gesa Schad, Shimadzu Europa A simple, rapid, and robust ultrahigh-performance liquid chromatography (UHPLC)
method for the simultaneous determination of natural and artifi cial vanilla fl avouring substances as well as some precursors has been developed using an automated method scouting or method optimization workfl ow.
Regulars9 News The latest company news, peaks of the week, and news in brief are featured in this issue.
12 Tips & Tricks GPC/SEC: Branching Analysis Daniela Held, Peter Montag, and Wolfgang Radke, PSS Polymer Standards
Service GmbH Branching is one of the parameters chemists can adjust to produce polymer
materials with optimized physical properties. Chromatography and advanced detection can help to characterize branched molecules. This instalment of Tips & Tricks explains more.
25 The 31st International Symposium on Chromatography (ISC 2016) A preview of the upcoming 31st International Symposium on Chromatography
(ISC 2016), which is due to be held 28 August–1 September at University College
Cork, Ireland.
27 Training Courses and Events
28 Staff
7 June 2016 Volume 12 Issue 10
Constantly EvolvingLeon Blumberg has been at the cutting edge of chromatography since the late 1980s. The Column spoke with him about his contribution to the past, present, and future of chromatography.
— Interview by Lewis Botcherby
Q. You began your career as an
electrical engineer. How did you come
to work within the fi eld of analytical
chemistry and more specifi cally gas
chromatography?
A: I joined the Avondale Division of
Hewlett-Packard Co, which is now
Agilent Technologies, in 1977. HP
Avondale designed and manufactured gas
chromatography (GC) instrumentation. My
initial responsibilities were to design the
electronics and the software for digitizing
and analyzing the chromatograms. From that
grew the need for a better understanding
of the chromatographic limits to separation
performance and speed of analysis. I was
also asked to fi gure out the effect of peak
focusing by the velocity gradients within a
column on the column performance — a
controversial issue at the time.
Q. Why was this?
A: It is a fascinating story, but, first,
I would like to clarify that, in all
forthcoming answers, I distinguish the
peaks in chromatograms (their widths
are measured in the time units) from the
bands of analyte within a column (their
widths are measured in the distance
units). In the late 1980s, a group of
inventors approached HP with a proposal
to substantially improve the speed of GC
analysis by using the dynamic focusing
(negative temperature gradients moving
from the column inlet to the outlet). An
enormous increase in the speed of analysis
without losing resolution was expected
(120 min PONA analysis would be reduced
to a few minutes). The idea was that the
negative temperature gradients along
the column create the negative velocity
gradients causing the front of an analyte
band migrating along a column to move
slower than its tail. This compresses the
bands making them narrower in distance
along the column, and thus presumably
improves the separation-speed trade-offs.
Ray Dandeneau, a co-inventor of fused
silica capillary columns and R&D manager
at the time, asked me to contact the
inventors for a detailed exploration of the
potentials. After preliminary discussions, Ph
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it was decided that HP would help the
inventors to construct the experiments,
and I began, with help from Terry Berger,
to search for theoretical answers. Soon
I found that the dynamic focusing was
an old idea proposed by Russian scientist
Zhukhovitskii1 in 1951 before the invention
of conventional temperature-programmed
GC.2 Since then, high expectations from
dynamic focusing had been periodically
flaring up and dying down again from time
to time with no definitive experimental
improvement and no theoretical solutions.
As others before them, our experiments
did not show any definitive improvement.
The explanation given by the inventors
was that the experiments were not good
enough and Terry and I continued the
search for definitive theoretical answers.
In 1992, we published a theoretical paper3
on the fundamentals of the effects of
the velocity gradients. I was lucky to have
the opportunity to have an encouraging
discussion of the paper with the late Prof.
Giddings during the 1992 International
Conference on Chromatography in
Aix-en-Province, France. Later, I published
a more general theory4,5 and then two
theoretical papers6,7 demonstrating that,
ideally, the velocity gradients can only
harm the separation-speed trade-offs in
chromatography, although the gradients
can reduce the harm from some non-ideal
conditions like poor sample introduction.
These general conclusions can be explained
this way. The same velocity gradients that
compress the analyte bands (make them
narrower in distance along the column)
also reduce the distance between the
bands and slow down the speed of their
migration and elution. The latter two
factors reduce the separation of the bands
and broaden the peaks in chromatograms.
Together, the three conflicting factors
(band compression, worsening of their
separation, and their slower elution) will at
best cancel each other out; at worst they
will reduce the separation-speed trade-offs.
After my involvement in the focusing study,
my expertise in GC theory became one of
my main responsibilities at HP.
Q. Could you briefl y talk about the
work you performed and what you
helped develop while you were at HP?
A: My first job assignment was to
develop an analog-to-digital converter
(ADC) for the first stand-alone digital
integrator in the industry, which
was under development at the time.
This was followed by development
of data-processing and presentation
software for another integrator that
the company introduced later, and then
Q&A: Blumberg
3
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by defining the speed-related aspects
for the next-generation high speed GC
instrument — the first in the industry with
fully integrated electronic pressure and
flow control. As part of that assignment,
I co-invented ADC for GC8 — the fastest
at the time for the needed resolution —
and a GC method translation concept.9,10
Later, I developed a GC method translation
software that became a popular method
development tool. I also co-invented the
retention time locking (RTL) concept11,12,13
— the basis for another useful software
tool introduced by HP. The tool enabled
an order of magnitude improvement in GC
retention time reproducibility.
Q. You continued to work on
the development of fast and
multi-dimensional GC. What do you
feel were some of your most important
publications on these topics?
A: 1D GC (one-dimensional GC): I
completed the publication of a four-part
series on the theory of fast GC,14–17 and
co-authored several papers on heating rate
optimization in GC.18,19
GC×GC (comprehensive two-dimensional
GC): In my first GC×GC lecture (25th
International Symposium on Capillary
Chromatography, Riva del Garda, Italy,
2002, published in 200320), I outlined
the first (as far as I know) theory of
optimization of GC×GC, and theoretically
demonstrated that the separation
performance of GC×GC can potentially be
orders of magnitude higher than that of
1D GC. However, I also demonstrated that,
because of insufficiently sharp modulation,
the GC×GC systems available at the time
were not better than their 1D counterparts.
This caused heated debates,21,22 lasting for
many years and summarized in Pat Sandra’s
2007 lecture23 at the 4th International
GC×GC Symposium (Dalian, China,
2007), published in 200824 (with several
co-authors including myself). Years later,
better modulators became available, and
the key problems of GC×GC configuration
were fixed (all as I was preaching for years).
Several years ago, Jack Cochran described
(International GC×GC Symposium, Riva
del Garda) the first GC×GC system known
to me at the time that performed near its
theoretical potential. The system evaluation
was presented by Matthew Klee at the
11th International GC×GC Symposium (Riva
del Garda, 2014) and published in a paper
that I initiated and co-authored.25 I am
also thankful to Luigi Mondello for inviting
me to contribute a chapter on theory and
optimization of GC×GC coupled to mass
spectrometry (MS) for his volume on the
same topic published in 2011.26
Q&A: Blumberg
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Q. In 2010 you published
Temperature-Programmed Gas
Chromatography, which is a very
comprehensive publication covering
many aspects of GC theory.27 How did
this book come about?
A: Although temperature programming
is a key factor of a column’s performance
in GC, no books on this topic had been
published since 1966.28 I had been thinking
about publishing such a book since the
mid-1990s. I began the writing in 2003. Up
until the book was published,27 I did not
work on much else. Unfortunately, I did not
include in the book everything I wanted
(performance metrics, optimization), but the
time to stop came. I believe, however, that
what’s covered is done reasonably well. In
addition to good published reviews, several
colleagues verbally expressed pleasant
comments. Later, Colin Poole gave me an
opportunity to publish a summary of the
performance metrics and the optimization in
a theoretical chapter of the volume on GC
that he edited.29
Q. Your most recent publication focused
on optimizing the mixing rate in
linear solvent strength gradient liquid
chromatography (LC),30 could you
briefl y explain what the mixing rate is
and its importance?
A: The mixing rate is the rate of the
temporal increase (the increase with time)
of the volume fraction of stronger solvent
in LC mobile phase. The mixer is the device
changing the solvent composition. From
that perspective, gradient LC can be viewed
as LC with programmed solvent mixing. It
is well known that the analysis time of this
technique can be substantially shorter than
that of isocratic LC (where the mobile phase
composition does not change during the
analysis). The term mixing rate, which was
recently introduced by Gert Desmet and
myself,30 is a synonym of the wider know
terms gradient slope and time steepness
of the gradient. Why the new term? The
solvent strength programming causes two
types of solvent strength change within
an LC column — the temporal change
and the spatial one (the change along the
column). It is important to recognize the
difference between these two types of
changes because they affect the column
performance differently (see below). Using
the term rate for the temporal changes,
and the term gradient only for the spatial
ones, helps to reinforce the distinction,
and to separately evaluate the effects of
each phenomenon. Using this distinction,
I was able to demonstrate theoretically31
that sharpness of chromatographic peaks
and the resulting speed improvement in
Q&A: Blumberg
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gradient LC compared to isocratic conditions
comes from the temporal changes while
the effects of the gradients are practically
insignifi cant under typical conditions. It is
important for the clarity of terminology that
the term mixing rate that represents only
the temporal changes does not include the
words like “gradient” and “slope”, which
have spatial implications. From a broader
perspective, gradient LC is a special case
of chromatography with dynamic focusing,
and all conclusions regarding the general
case6,7 apply to LC. Interestingly, in line with
predictions of general theory of dynamic
focusing,6,7 the gradient band compression
typically broadens the peaks slightly30 rather
than focusing (narrowing) them as frequently
expected.
Q. When you optimize anything there is
a trade-off between parameters. These
changes are based on the optimization
goal. What exactly was the optimization
goal within this research?
A: The optimal mixing rate in LC (as
well as the optimal heating rate in GC)
is defi ned19,29,32 as the one at which a
required separation performance is obtained
in the shortest time.
Q. What practical recommendations
have come out of this research?
A: In essence, the optimal mixing rate
(OMR) mostly depends only on the void
time and on the molecular weights of the
sample components. What’s nice is that the
OMR does not directly depend on column
dimensions, its solid support structure, or the
solvent type, but only through their effect
on the void time. This makes it possible to
reduce the optimization results down to
specifi c numerical recommendations. Thus,
for small-molecule samples, the solvent
strength increase of about 5% per time
increment equal to the void time is optimal
or close to optimal for all LC analyses that
can be approximated by the LSS (linear
solvent strength model). The per-void-time
increment for proteins should be about 10
times smaller. This is similar to the optimal
heating rate of 10 ºC per void time in
GC.19,29
Q. What are you currently researching?
A: I am grateful for having the opportunity
to work with Gert Desmet — a leading
expert in theory of gradient LC. We are both
interested in optimization of the technique.
A key to the optimization is the simple and
transparent performance metrics. Together,
we published a paper on the metrics
of separation performance in gradient
LC30 and on the optimal mixing rate32
in a simple single-ramp solvent strength
Q&A: Blumberg
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program. Currently, we are working on
the optimization of more complex mixing
programs.
Q. What do you regard as the most
exciting areas of chromatography at the
moment?
A: In my view, comprehensive
multidimensional techniques are the
most promising developments in column
chromatography (LC, GC, etc.). The
GC×GC “is probably the most promising
invention in GC since discovery of capillary
columns.”24 The overall performance of
a chromatographic analysis is a trade-off
between three factors — the separation
performance, the time, and the detection
limit (DL). Here’s an example from GC with
which I am more familiar: Each twofold peak
capacity increase in 1D GC without changing
the DL costs eightfold longer analysis time
(1 h analysis becomes 8 h analysis). On the
other hand, adding the second dimension
can increase the peak capacity by more than
an order of magnitude without changing
the analysis time and DL. Currently, not all
potentials of GC×GC are exploited. Thus,
peak deconvolution substantially increases
the separation performance of 1D GC and
GC×GC. However, only the deconvolution
along the second dimension is currently used
in GC×GC. Adding the deconvolution to
the fi rst dimension of GC×GC can further
increase its overall separation performance
several times.20,26 Unfortunately this
approach remains mostly unknown.
Q. Will the evolution of mass
spectrometry eventually lead to the
extinction of chromatography?
A: I am not an expert in mass spectrometry
(MS) and in the multi-stage MS techniques.
However, the way I see this world, there
will always be the need for the separation
of more and more complex mixtures. The
improvement in the separation-time-DL
trade-offs in multi-dimensional chromatography
and in multi-stage MS will always complement
each other, but more separation power will
always be needed. Could you imagine a
day when there will be no practical need
for further substantial improvement in
computer speed or memory, or in the speed
of the Internet? I think the same is true for
the separation-time-DL performance of
analytical tools.
References
1. A.A. Zhukhovitskii, O.V. Zolotareva, V.A.
Sokolov, and N.M. Turkel’taub, Doklady
Akademii Nauk S.S.S.R. 77, 435–438
(1951).
2. J. Griffiths, D. James, and C.S.G. Phillips, The
Analyst 77, 897–904 (1952).
Q&A: Blumberg
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19. L.M. Blumberg and M.S. Klee, J. Microcolumn
Sep. 12, 508–514 (2000).
20. L.M. Blumberg, J. Chromatogr. A 985, 29–38
(2003).
21. C.M. Harris, Anal. Chem. 74, 410A (2002).
22. L.M. Blumberg, Anal. Chem. 74, 503A (2002).
23. P. Sandra, F. David, M.S. Klee, and L.M.
Blumberg, “Comparison of one-dimensional
and comprehensive two dimensional capillary
GC separations,” Dalian, China, 4–7 June
2007, (CD ROM).
24. L.M. Blumberg, F. David, M.S. Klee, and P.
Sandra, J. Chromatogr. A 1188, 2–16 (2008).
25. M.S. Klee, J.W. Cochran, M. Merrick, and L.M.
Blumberg, J. Chromatogr. A 1383, 151–159 (2015).
26. L.M. Blumberg, in Comprehensive
Chromatography in Combination with Mass
Spectrometry, L. Mondello, Ed. (Wiley,
Hoboken, NJ, USA, 2011), pp. 13–63.
27. L.M. Blumberg, Temperature-Programmed
Gas Chromatography (Wiley-VCH, Weinheim,
Germany, 2010).
28. W.E. Harris and H.W. Habgood, Programmed
Temperature Gas Chromatography (John Wiley
& Sons, Inc., New York, USA, 1966).
29. L.M. Blumberg, in Gas Chromatography, C.F.
Poole, Ed. (Elsevier, Amsterdam, 2012) pp.
19–78.
30. L.M. Blumberg and G. Desmet, J. Chromatogr.
A 1413, 9–21 (2015).
31. L.M. Blumberg, Chromatographia 77, 189–197
(2014).
32. L.M. Blumberg and G. Desmet, Anal. Chem.
88, 2281–2288 (2016).
Leon Blumberg
graduated from the
Leningrad Electrotechnical
Institute (Leningrad, USSR;
currently St Peterburg,
Russian Federation) in
1960 with a diploma
in electrical engineering and went on
to join a computer design company in
Leningrad. In 1961–1965, Leon completed
a full math course for professional
mathematicians from Leningrad University.
He obtained a PhD equivalent in electrical
engineering from Leningrad Electrotechnical
Institute of Telecommunications. In
1977, Leon immigrated to the US with
his family, joining the Avondale Division
of Hewlett-Packard Co. (now Agilent
Technologies). Currently, as part of his
consulting company Advachrom, Leon
provides consulting services mostly in
GC×GC. His main scientifi c interest
now is to develop a unifi ed theory of
temperature-programme GC and
gradient LC.
E-mail: leon@advachrom.com
3. L.M. Blumberg and T.A. Berger, J. Chromatogr.
596, 1–13 (1992).
4. L.M. Blumberg, J. Chromatogr. 637, 119–128
(1993).
5. L.M. Blumberg, J. High. Resolut. Chromatogr.
16, 31–38 (1993).
6. L.M. Blumberg, Anal. Chem. 64, 2459–2460
(1992).
7. L.M. Blumberg, Chromatographia 39, 719–728
(1994).
8. L.M. Blumberg, J. Bush, and R.P. Rhodes, U.S.
patent 5,448,239, 1995.
9. W.D. Snyder and L.M. Blumberg, U.S. patent
5,405,432, 1995.
10. L.M. Blumberg and M.S. Klee, Anal. Chem. 70,
3828–3839 (1998).
11. M.S. Klee, P.L. Wylie, B.D. Quimby, and L.M.
Blumberg, U.S. patent 5,827,946, 1998.
12. M.S. Klee, B.D. Quimby, and L.M. Blumberg,
U.S. patent 5,987,959, 1999.
13. L.M. Blumberg, B.D. Quimby, and M.S. Klee,
U.S. patent 6,153,438, 2000.
14. L.M. Blumberg, J. High. Resolut. Chromatogr.
20, 597–604 (1997).
15. L.M. Blumberg, J. High. Resolut. Chromatogr.
20, 679–687 (1997).
16. L.M. Blumberg, J. High. Resolut. Chromatogr.
22, 403–413 (1999).
17. L.M. Blumberg, J. High. Resolut. Chromatogr.
22, 501–508 (1999).
18. L.M. Blumberg and M.S. Klee, Anal. Chem. 72,
4080–4089 (2000).
Q&A: Blumberg
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Changing the Landscape of Mass Detection in the
Chromatography Lab
EVENT OVERVIEW
Traditionally mass detection instruments have been for
the mass spec experts and not played a major role in
the majority of chromatography labs. With the advent of
smaller, more accessible mass detectors, the potential of
mass data is coming more and more within the reach of
the chromatographer. With this webcast we look to see
how the landscape of the chromatography lab is chang-
ing and how the value of mass data can be realized by
the chromatographer.
Key Learning Objectives
■ How the landscape of mass detection in the chromatog-raphy lab is changing
■ Value of mass data to a chromatographer
■ How a chromatography data system (CDS) should utilize the power of mass detection and mass data
Who Should Attend
■ Method development chemists
■ Development lab managers
■ QC lab managers
Sponsored by Presented by
All attendees will receive a FREE Executive Summary! For questions, contact Kristen Moore at kmoore@advanstar.com
ON-DEMAND WEBCASTOriginally aired 04/28/16
Presenter
David Wayland
Empower product
owner
Waters Informatics
Moderator
Laura Bush
Editorial Director
LCGC
Register free at www.chromatographyonline.com/lcgc/
landscape
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Waters Announces Additions to Leadership Team
Waters Corporation (Milford, Massachusetts, USA) has announced
the addition of Richard Chang and Xiao Ran Yu to its Asia Pacifi c
leadership team. Chang has been appointed as Vice President of
Waters’ Asia Pacifi c Operations and Yu has been appointed as
General Manager of Waters China.
“Mr. Chang’s and Mr. Yu’s impressive proven track records have
demonstrated an intimate understanding of the marketplaces in
the region,” said Dr. Mike Harrington, Senior Vice President, Global
Markets, Waters Corporation. “With their extensive experience and
strong commitment, our China business has become the single biggest
market for Waters outside the US. We strongly believe that they will
continue to drive growth in the Asia Pacifi c region,” he continued.
Beginning his career with Waters in Taiwan, Chang moved to
China following four successful years as General Manager to help
develop the Chinese market. Appointed as President of Greater
China operations in 2012, he has led Waters’ signifi cant growth
within the Asia Pacifi c region.
“It’s my great honour to lead the Asia Pacifi c team and I look
forward to working closely with our customers and partners in
meeting their needs, and bringing Waters innovation to them in the
most accessible way,” said Chang.
“I’m excited to join the leadership team in Asia Pacifi c and apply
my wealth of experience and passion for science and technology
to support customers in China,” enthused Yu, who has more than
22 years of experience in the analytical science industry. The former
operations manager for South China had led his team to signifi cant
sales revenues during his tenure.
Waters are hopeful these two new appointments can drive
growth within the region and capitalize on the emerging strength
of the Asia Pacifi c markets.
For more information please visit www.waters.com.
Thermo Fisher Partner with University of Birmingham’s New Phenome CentreThermo Fisher Scientifi c (San Jose, California, USA) has extended a technology collaboration with the University of
Birmingham in the UK. Thermo Fisher will supply a range of mass spectrometry instruments to be used at the University’s
new Phenome Centre.
The £8 million facility provides University of Birmingham scientists with the tools needed to conduct large-scale metabolic
phenotyping research, advancing the understanding of biochemical mechanisms, targets, and biomarkers associated with
ageing and disease.
“We are pleased to continue our long-standing collaboration with Thermo Fisher in these important areas of
metabolome research,” said Professor Mark Viant, Director of Phenome Centre Birmingham.
“Combined with our strong impact in technology and method development, we look forward to the benefi ts
we will translate to the human population through stratifi ed medicine approaches,” continued Viant.
Stratifi ed medicine, a method of predicting which treatments cancers are likely to respond to by
studying the cells and genetic make up of large groups of cancer patients, is part of the larger
strategy employed at the facility.
Iain Mylchreest, vice president of R&D for chromatography and mass spectrometry at Thermo
Fisher, congratulated the university on the opening of the Phenome Centre, adding, “These
scientifi c relationships are mutually benefi cial in helping us to advance our analytical
tools and technologies that, in turn, enable our customers to make the world healthier,
cleaner, and safer.”
This recent collaboration is just one of many between the University of Birmingham
and Thermo Fisher across 10 years. Most recently, the organizations have collaborated
to accelerate research in high-resolution accurate mass (HRAM) and triple quadrupole
liquid chromatography-mass spectrometry (LC–MS) for life sciences applications.— L.B.
For more information about Thermo Scientifi c,
please visit www.thermofi sher.com
For further information about the Phenome Centre Birmingham, please visit
http://www.birmingham.ac.uk/research/activity/phenome-centre
9
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The Column www.chromatographyonline.com
Shimadzu Lab4You AcceptingApplications
Ph
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Following a successful launch last year,
Shimadzu’s Lab4You program is once again
accepting applications from young and
enthusiastic researchers hoping to advance
their research.
Shimadzu is offering a few promising
scientists the chance to work at “Shimadzu
Laboratory World”. The winners will be
able to advance their research utilizing the
state-of-the-art equipment at the facility
including well-serviced analytical instruments
and on-hand product specialists.
“The work was very interesting and I
highly recommend other students apply for
the Lab4You student program,” said Carola
Schultz, University of Münster.
Schultz was one of two winners from last
year’s lab4you competition. Her research
investigated organic lithium-ion battery
electrolytes using liquid chromatography–
mass spectrometry (LC–MS).
“They [lithium-ion batteries] contain organic
carbonates and the lithium-ion conducting
salt LiPF6. But during cycling of the cell, the
electrolyte ages and the cell loses capacity.
The generated ageing products are built
up out of reactions from the salt with the
organic carbonates,” explained Schultz.
“So my aim was to do a full characterization
of the electrolyte with LC–MS, to investigate the
main components as well as ageing products
in order to facilitate the elucidation of ageing
mechanisms occurring inside the cell into the
electrolyte,” continued Schultz, who is currently
working towards publishing her results.
The Lab4You program emerged from
Shimadzu’s 2013 Laboratory World opening
in Duisburg, Germany. The 1500 m2
testing facility contains Shimadzu’s entire
product range and was used for customer
demonstrations and application training.
“We felt that there was still some more
capacity for the instruments to be used.
So we came up with the idea to invite students
with interesting research projects to come to
Duisburg and use the high-end equipment
that is not available at their University, in our
Laboratory Worlds,” said Dr. Gesa J. Schad,
HPLC Product Manager, Shimadzu.
Minimum requirements for candidates
include an undergraduate degree with a
science background, an approved topic of
research (MSc or PhD thesis or postdoc
research), and some practical experience in
analytical chemistry.
“We are also looking for good
interpersonal and communication skills.
The successful candidate will spend a
considerable amount of time working with
us, so they should fi t in well with the team.
Shimadzu offers to sponsor registration to
relevant conferences, where the obtained
data can be presented,” explained Schad.
Interested students can apply by submitting
a short abstract of their research at www.
shimadzu.eu/lab4you. Deadline for submission
is 31st October 2016. — L.B.
Carola Schultz during her time at Shimadzu’s Laboratory World.
From left to right: Anja Grüning, Product
Specialist LC–MS, Dr. Julia Sander, Product
Specialist Life Sciences; Carola Schultz,
lab4you participant; Uta Steeger, Manager
Marketing; Dr. Gesa Schad, Product Manager
HPLC; Robert Ludwig, Product Specialist HPLC;
and Philipp Jochems, Product Specialist HPLC.
News
10
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News In Brief
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LCGC magazine is pleased to announce the addition of Deirdre Cabooter and Debbie Mangelings to the editorial advisory boards of LCGC North America and LCGC Europe. Cabooter is an assistant professor in the Department of Pharmaceutical and Pharmacological Sciences at the University of Leuven in Belgium. Mangelings currently works as an associate professor in the Department of Analytical Chemistry and Pharmaceutical Technology.
W.R. Grace & Co. has agreed to sell product lines associated with its chromatography instruments, columns, and related laboratory products businesses to four separate buyers: Buchi, Dr. Maisch, Hichrom, and S*Pure. Grace will retain three growing service lines for the pharmaceutical and nutraceutical industries that are aligned with the company’s growth plans, while concentrating on its core materials science and manufacturing capabilities.https://grace.com/en-us/newsroom/Pages/
news-item.aspx?ItemID=498
A study has been conducted to evaluate the migration of monomers and plastic additives in microwaved solid or liquid packed and retailed food. Using GC–MS and QuEChERS recoveries ranged from 49 ± 16% (OP) to 130 ± 16% (BPA) and from 63 ± 22% (OP) to 127 ± 29% (NP) in solid and liquid foods. The QuEChERS method was able to determine the presence of these chemicals in packed food, thereby allowing the evaluation of compounds that can affect food quality.doi:10.1016/j.lwt.2015.06.066
LCGC TV HighlightsLCGC TV: Barbara Larsen on Replacing a
Mass Spectrometer: What to ConsiderReplacing an older mass spectrometer involves a reevaluation of the way the instrument is being used. Barbara Larsen from DuPont Central Research and Development discusses the checklist
you should follow.Watch Here>>
LCGC TV: The Potential for Ionic Liquid-Based Coatings in SPMEMore and more analytes are now being found in complicated matrices. Jared Anderson from the Iowa State University recently designed new SPME coatings based on polymeric ionic liquids suitable
for these matrices. He discusses the potential for these coatings to improve on SPME. Watch Here>>
Peaks of the WeekThe LCGC Blog: The Middle Ground on Unconventional Oil and Gas Development (aka
“Fracking”) is a Lonely Place — Kevin A. Schug discusses his research consortium’s work on the
quality of groundwater in close proximity to fracking sites, the disparity in the media’s interpretation of
the data, and addresses valid queries made by critics. Read Here>>
The Modulator in Comprehensive Two-Dimensional Liquid Chromatography —
This article illustrates the variety of commonly used modulators, paying particular attention to
focusing modulators.
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The SFC Renaissance? — Jean-Luc Veuthey and Alexandre Grand-Guillaume-Perrenoud reveal the
latest developments in supercritical fluid chromatography (SFC) that are bringing the technique
back to the limelight.
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The Column www.chromatographyonline.comPh
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pm
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News
11
Q&A: Blumberg2 News9 Tips and Tricks12 Jochems and Schad1899 122Beck and Delobel21 ISC Event Preview25 Training and Events27 Staff28252 272
Tips & Tricks GPC/SEC: Branching Analysis
Branching is one of the parameters chemists can adjust to produce polymer materials with optimized physical properties. Chromatography and advanced detection can help to characterize branched molecules. This instalment of Tips & Tricks explains more.
Daniela Held, Peter Montag, and Wolfgang Radke, PSS Polymer Standards Service GmbH, Mainz, Germany.
An advantage of polymer materials is that
their physical properties can be tailored
for a specific application by adjusting
many parameters. Besides composition,
average molar mass, and the width of
the molar mass distribution, another key
parameter to control application properties
is branching.
Branching requires at least a single
branch point where three or more chains
are connected. Branching can occur as an
undesired side reaction during synthesis
or it can be introduced deliberately to
optimize the physical properties of the
material. Different routes exist to synthesize
defined structures, such as star-shaped or
comb-shaped molecules.
The properties of branched polymers
differ significantly from linear ones with
respect to (melt) viscosity, glass transition
temperature, the coefficient of bulk
thermal expansion, solubility, and others.
The property change depends on the
parameters such as type of branching,
length of the branches, and branching
density.
The characterization of complex
mixtures comprising not only a molar mass
distribution but branching distribution
as well, represents a real challenge.
Depending on the type of branching
there are various detection and separation
options that provide deeper insight.
Gel permeation chromatography/
size-exclusion chromatography (GPC/SEC)
hyphenated with on-line viscometry1 (or
less accurate multi-angle light scattering)
can be used to characterize defined
structures, such as star or comb-shaped
polymers, or to investigate long chain
branching. High temperature GPC (HT-GPC)
with infrared (IR) detection can be used
to investigate short-chain branching in
polyolefins.2
For samples exhibiting broad molar mass
distributions for branches and backbone Ph
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: Ju
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Eye
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/Ge
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12
Q&A: Blumberg2 News9 Tips and Tricks12 Jochems and Schad1899 122Beck and Delobel21 ISC Event Preview25 Training and Events27 Staff28252 272
The Column www.chromatographyonline.com
and variations in branching density, the
resolution of the size-based separation
in GPC/SEC might not be sufficient to
fully resolve the structures. Therefore
alternative separation methods such as
interaction chromatography (for example
gradient polymer high performance liquid
chromatography [HPLC] or temperature
gradient interaction chromatography
[TGIC]) or two-dimensional (2D)
chromatography should be applied.3
GPC/SEC-Viscometry
An on-line viscometer is classifi ed as a
molar mass sensitive detector, however, the
signal intensity is dependent on the viscosity
rather than on molar mass. Viscometers
provide direct access to the density of the
molecules in solution. While the setup of
such instrumentation, in most cases in
combination with a light scattering detector,
is very common, understanding the results
and limitations requires more experience.
The Mark–Houwink plot is important for
branching analysis; the logarithm of the
intrinsic viscosity (obtained using on-line
viscometry) is plotted versus the logarithm
of the molar mass (obtained using universal
calibration or light scattering detection).
The slope of the Mark-Houwink plot
(Mark-Houwink coeffi cient, α) is dependent on
the shape of the molecule in solution. If the
intrinsic viscosity (solid sphere) has no molar
mass dependence a slope of 0 is expected,
however, the Mark–Houwink exponent of rigid
rods is 2. Typical random coil polymers exhibit
Mark-Houwink exponents in the range of 0.5
to 0.8, depending on solvent quality.
Branching analysis can be
straightforward, assuming that the data
can be compared to a linear chain of
identical chemical structure and molar
mass. Figure 1 shows Mark–Houwink
plots for different polyethylene samples.
A HT-GPC equipped with an on-line
viscometer has been used to generate this
plot. While the Mark–Houwink plot of the
linear low-density polyethylene (LLDPE),
which has only short chain branching,
nearly superimposes with the linear sample,
10 2
Vis
cosi
ty
PS
SW
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PC
Un
ity,
Bu
ld 6
08
2,
LA
B_A
ll 4
, In
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z#
1
Mark-Houwink-Plot
NBS-1475: lin. PE, pink, α = 0,68LLDPE-C6: black, α = 0,70
LDPE: blue, α = 0,42
10 1
1×10 4
Universal calibration (Da)
1×10 5 1×10 6
Figure 1: Mark-Houwink plot overlay of three different polyethylene samples. NBS-1475 (pink) is linear, linear low density polyethylene (LLDPE, black) has short-chain branching that does not infl uence the viscosity much, and low density polyethylene (LDPE, blue) exhibits long-chain branching, which leads to a reduction of viscosity.
Tips and Tricks
13
Q&A: Blumberg2 News9 Tips and Tricks12 Jochems and Schad1899 122Beck and Delobel21 ISC Event Preview25 Training and Events27 Staff28252 272
Proteins of biopharmaceutical interest are generally hetero-geneous mixtures of proteoforms comprised of modifications. Accurate knowledge of the proteoform profile is critical for assessing the safety and stability of the drug. Current methods of analyzing post-translational modifications (PTMs) and glycan structures require proteolysis or glycan profiling that can result in lost information on correlated PTMs. Sample handling can also introduce artifacts and therefore, minimal sample preparation is desirable. Top-down mass spectrometry (MS) analysis of highly heterogeneous biopharmaceuticals is challenging due to the limited ability of current separation techniques in resolving proteoforms with small structural changes. In this work, we describe the top-down analysis of interferon-β1 (Avonex) in detail and preliminary data on middle-down (reduced) and intact mAbs.
Key Learning Objectives:
■ Find out how CESI-MS enables high resolution intact separation and on-line top-down MS identification of PTMs with minimal sample preparation
■ Learn how glycan isomers (such as the two isomers G2F and 1 NANA) and deamidations can be resolved by intact analysis
■ Learn how a similar approach was applied for intact and reduced analysis of mAbs
Who Should Attend
■ Principal Investigators, department chairs, senior scientists, R&D directors, post-doctoral fellows, post-graduate researchers, and medical researchers.
Sponsored by
Presented by
For questions, contact Kristen Moore at kmoore@advanstar.com
High-Resolution Quantitative Characterization of Intact Biopharmaceuticals and Their ProteoformsON-DEMAND WEBCAST Aired May 25, 2016
Register for free at www.chromatographyonline.com/lcgc/sciex_series2
Presenter:
David R. Bush, Ph.D.Scientific ManagerGenedata, Inc.
Moderator:
Laura Bush
Editorial Director LCGC
30 MinuteFormat
The Column www.chromatographyonline.com
the low-density polyethylene (LDPE), which
comprises long-chain branching, deviates
significantly. At the same molecular weight
the LDPE chains reveal a significantly lower
intrinsic viscosity compared to the linear
sample. This is a consequence of branches
being present. The deviation increases
with increasing branching density. By
extrapolating the Mark–Houwink plots
of the branched and linear polymer to
a common intercept, it is possible to
detect the molar mass at which branching
first occurs. By taking the ratio of the
intrinsic viscosity of the branched and
linear polymer at the same molar mass, it
is possible to determine the contraction
factor, g’, from which conclusions on the
number of branches can be deduced.
Figure 2 shows the results of GPC/
SEC-viscometry of a poly(tert-butyl acrylate),
PtBuA, star polymer. Star polymers are
relatively simple branched polymers because
they consist of several arms (linear chains)
connected to a central core.
The star polymer was synthesized using
the arm-first approach. PtBuA arms of
narrow molar mass distribution have
been coupled using a small amount of
a bifunctional cross-linker to form the
core. This means that the molar mass
increase of the star was obtained by
coupling an increasing number of arms
of approximately the same length to the
core. The Mark–Houwink plot reveals a
maximum for the intrinsic viscosity, which
has also been observed for dendrimers.
Starting with a linear precursor, the
coupling of two linear chains forms a still
linear molecule, the dimer. Further reaction
of precursor molecules with the core leads
to three-arm and higher arm star polymers.
Here the increase of the intrinsic viscosity
with molar mass is counterbalanced by
the decreasing intrinsic viscosity resulting
from an increasing segment density
with increasing number of arms for the
branched structures.
This variation in molecular structure
can be nicely monitored from the viscosity
measured using an on-line viscometer.
0.010
102
0.009
0.008
PSS W
inG
PC
Un
ity
Sp
eci
fic
vis
cosi
ty
Intr
inis
c V
isco
sity
Number of ArmsDegree of Branching
Intr. Visc.Linear Counterpart
Star Polymer withaverage arm number 5
4.5% Dimer
12.1% Residual Arm
Molar mass from universal calibration curve (D)
4.0% CoupledStar Polymer(H-shaped)
0.007
0.006
0.005
0.004
0.003
0.002
0.001
5*10 4
1*10 5
5*10 5
1*10 6
(still linear)
Figure 2: Mark-Houwink for a star polymer using the arm-fi rst approach. Arms with a narrow molar mass distribution are coupled to a core; the molar mass increase is a result of the addition of more arms to the core. The structure change (linear coil to a more and more dense sphere) is refl ected by a maximum in intrinsic viscosity.
Tips and Tricks
14
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LIVE WEBCAST: Wednesday, June 15, 20168 am PDT | 10 am CDT | 11 am EDT | 4 pm BST | 5 pm CESTRegister free at: www.chromatographyonline.com/lcgc/quantitation
EVENT OVERVIEW
The screening and routine quantitation of pesticide residues
in food products is one of the most important and demanding
applications in food safety. Despite the recent technological
advancements in LC-MS, it is still challenging to quantify
hundreds of LC-amenable pesticides with a robust, sensitive
workflow solution.
This presentation describes the development and implemen-
tation of complete workflow solutions based on LC-MS/MS
and LC-HRAM-MS/MS. These ready-to-go solutions have been
validated in three matrices across four different laboratories.
In addition, customized software used for data acquisition
and processing allows the users to rapidly implement these
methods and enhance productivity.
For questions, contact Kristen Moore at kmoore@advanstar.com
Key Learning Objectives
■ Address critical challenges in targeted or untargeted quanti-
tation of pesticides in food laboratories using either triple
quadrupole MS or high-resolution accurate mass (HRAM)
MS instrumentation
■ Learn about robust, routine workflows that can increase
laboratory and organizational productivity
Who Should Attend
■ Researchers and analysts in need of fast and cost-effective
solutions for the analysis of pesticides in food
Sponsored by
Presented by
Presenters
Ed George
Senior Applications Scientist,
Environmental and Food Safety,
Chromatography and Mass Spectrometry
Thermo Fisher Scientific, Inc.
Debadeep Bhattacharyya
Senior Marketing Manager,
Triple Quadrupole MS
Thermo Fisher Scientific, Inc.
Moderator:
Laura Bush, Editorial Director, LCGC
Pesticide Residues Analysis Webinar Comprehensive Pesticide Quantitation Workflow with LC-MS
The Column www.chromatographyonline.com
A very interesting observation is the
sudden change of intrinsic viscosity
occurring at approximately twice the
molecular weight of the peak maximum.
The drastic increase in viscosity is most
probably a result of the formation of
H-shaped molecules as coupling of two
stars occurs via an arm.
In contrast to the application above,
star polymers can also be synthesized
using the core first approach, for
example, using a multifunctional initiator.
In this case the observations and results
would be different. There would be no
structural change because the molar mass
increase of the star would result from the
3,0
2,5
2,0
1,5
1,0
0,5
0,0
-0,5
-1,03 4 5
Linear
3 arm star
4 arm star
7 arm star
log M
log
(η
)
6
Figure 3: Schematic Mark-Houwink plots for three star polymers synthesized via core-fi rst approach compared to a linear molecule. Arms are started from a multi-functional initiator core — molar mass increase is a result of the addition of monomer units to the arms. The Mark-Houwink plots are parallel to one of the linear polymers but shifted to lower intrinsic viscosities.
Tips and Tricks
15
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Bioanalysis: LC–MS-MS, Sample Prep, and Dried Blood Spot Analysis Bioanalysis uses a variety of separation techniques to analyze samples ranging from plasma and urine to dried blood spots. Participants in this Technology Forum are Ling Bei, Patrik Appelblad, and Dave Lentz of EMD Millipore; Nadine Boudreau of PharmaNet Canada; Diab Elmashni, Jeff Zonderman, and Simon Szwandt of Thermo Fisher Scientific; and Debadeep Bhattacharya of Waters Corporation. More...
Name your budget or application—the Agilent 1200 Infinity Series has you covered. From our affordable 1220 Infinity LC starting at just $15,000 to our cutting-edge 1290 Infinity LC, we have a solution that’s right for you. Plus, our most popular 1200 Infinity Series LC configurations are now available with a 3-5 year up-and-running guarantee. Read more.
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Health Sciences Unit LaunchedLGC has launched a new business unit, Health Sciences, which combines the group's sport, food, consumer safety and pharmaceutical testing activities within a single entity. More...
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Your brilliance. Our know-how. Collaborative Life Science. It all joins forces at EMD Millipore. Now your organization can leverage the combined synergies of two leading Life Science companies – for deep insight and know-how along every step of the biotherapeutic value chain. Find out how at www.emdmillipore.com
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Featured Application Note:Screening and Quantification of Multiple Drugs in Urine Using Automated Online Sample Preparation
and Tandem Mass Spectrometry Barbora Brazdova and Marta Kozak, Thermo Fisher Scientific
Learn about a 9-min, sensitive (LOQ 1—50 ng/mL) method to quantitate 30 immunosuppressant
drugs using TurboFlow technology and LC–MS-MS.
Evaluation of the Ultra Inert Liner Deactivation for Active Compounds Analysis by GCLimian Zhao, David Mao, and Allen Vickers, Agilent Technologies
Endrin and DDT breakdown and active semivolatiles tests were used for the Ultra Inert liner deactivation performance evaluation. The results indicate that the Ultra Inert deactivated liners provide superior inertness for analysis of active compounds.
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Food Analysis of PAHs Using GCxGC-TOFMS and QuEChERSLECO CorporationThe combination of QuEChERS extraction and GCxGC-TOFMS is a fast and accurate method for
detecting and identifying PAH contaminants in complicated foodstuff matrices such as liquid infant formula and blended blueberries.
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Screening and Identification with High Confidence Based on High Resolution and Accurate Mass LC–MS-MSAndre Schreiber and David Cox, AB SciexThis note describes a workflow and tools to identify targeted and nontargeted pesticides in fruits and vegetables. High resolution, accurate mass LC–MS-MS data is mined using advanced
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� Highly Sensitive UV Analysis with the Agilent 1290 Infinity LC System for Fast and Reliable Cleaning Validation – Part 1Edgar Naegele and Katja Kornetzky, Agilent TechnologiesThis application note demonstrates high sensitivity measurement of pharmaceutical compounds
with the Agilent 1290 Infinity LC. It also demonstrates a performance comparison of different flow cells with the Agilent 1290 Infinity LC diode array detector (DAD) for highly sensitive UV measurement including calibration, validation, and determination of LOD and LOQ.
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Chloramphenicol in Shrimp and Other Marine Food ProductsPhilip J. Koerner, Matthew Trass, Liming Peng, and Jeff Layne, PhenomenexA method for the analysis of chloramphenicol in shrimp has been developed with a limit of quantitation (LOQ) of 0.001 ng/g in shrimp (0.001 ppb) based on the calibration standards. This is 300 times lower than the current U. S. Food and Drug Administration (USFDA) method. The method described uses Strata-X solid phase extraction (SPE) cartridges for sample cleanup and
concentration, followed by ultra-fast LC–MS-MS analysis (<5 min) using a Kinetex core-shell column.
��
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www.chromatographyonline.com/enews
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growth of the arms. For such star
polymers the Mark–Houwink plots
are expected to be as shown in Figure
3. For each star polymer the Mark-
Houwink plot will be shifted in parallel
to lower viscosities at the same molar
mass. Analysis would yield the same
Mark-Houwink, α, but a reduced
Mark-Houwink, K (intercept). The shift to
lower intrinsic viscosities increases with
increasing number of arms.
Advanced Separation Techniques
One limitation of GPC/SEC is that it
separates only based on the size of the
molecule in solution. This has the following
consequences for branched samples:
t��$POWFOUJPOBM�DBMJCSBUJPO�XJUI�SFGFSFODF�
materials will underestimate the molar
mass of the branched samples. A
solution here is the use of molar mass
sensitive detectors, such as on-line
viscometers or light scattering detectors.
100
90
80
70
60
50
40
30
20
10
0
0 5 10 15
Elution volume (mL)
No
rmalized
co
nce
ntr
ati
on
Star with 3 Arms
2 arms/Dimer
Arm/Precursor
4 5 6 7 8 910
20 25 30 35 40
PSS W
inG
PC
sci
en
tifi
c V
& 1
0
Figure 4: Separation of a star branched polymer by interaction chromatography. The resolution of GPC/SEC for such samples is lower, because GPC/SEC separates based on size, which for this application increases only slightly for stars with more arms.
The viscometer can then be used for
structure analysis and for molar mass
determination based on universal
calibration.
t��*O�DBTFT�XIFSF�UIF�IZESPEZOBNJD�WPMVNF�
increases only slightly with molar mass,
such as arm-first star polymers, the
resolution of size-based separations
is limited. In this case interaction
chromatography can be used as a
complementary technique. Figure 4
shows the chromatogram of a gradient
separation of an arm-first star polymer
with a very high resolution even for stars
with higher arm numbers.
t��*G�TBNQMFT�FYIJCJU�CSPBE�NPMBS�NBTT�
distributions besides structural
heterogeneity (for example, branched
and linear chains present) the risk
of co-elution increases. In that case
branched molecules of higher molar
mass with the same hydrodynamic size
as linear chains of lower molar mass
elute at the same retention volume.
Consequently, the GPC/SEC fractions
eluting from the column cannot be
regarded to be monodisperse any longer.
Comprehensive characterization of
branched polymers might be possible when
2D separations are applied. Two-dimensional
combines two independent separation
Tips and Tricks
16
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ULTRA-FAST GCUseful Tool or Just Another Gimmick?
EVENT OVERVIEW
The fundamental principle of ultra-fast gas chromatogra-
phy is based on rapid temperature programming of the
GC analytical column at rates generally between 60 and
200 °C per minute. At these ramp rates, the dynamic tem-
perature range of most columns is used up in less than 2
minutes, and this is insufficient time to fully elute high-
boiling-point compounds, so the technique lends itself
to short columns. Shorter columns have less resolving
power, therefore it’s important to know the techniques to
optimize the parameters that give speed and resolution
without sacrificing column capacity.
Key Learning Objectives
■ Where to use ultra-fast GC
■ How to optimize parameters to get the best resolution without sacrificing column capacity
■ Types of ultra-fast GC systems
Who Should Attend
■ GC Users with large numbers of samples looking to increase sample throughput
■ GC Users with long analysis cycle times looking for faster cycle times
■ GC Users looking to reduce energy usage and environmental impact of GC analysis
ON-DEMAND WEBCAST Aired May 31, 2016
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For questions, contact Kristen Moore at kmoore@advanstar.com
Presenter
Phillip JamesManaging DirectorEllutia Ltd
Moderator
Meg L’HeureuxManaging EditorLCGC
The Column www.chromatographyonline.com
two-dimensional chromatography can be
applied.
References
1. D. Held, The Column 8(2), 12–16 (2012).
2. P. Montag, The Column 12, 14–17 (2008).
3. J. Gerber and W. Radke, Polymer 46,
9224–9229 (2005) doi: 10.1016/j.
polymer.2005.07.038.
Daniela Held studied polymer chemistry
in Mainz, Germany. She currently works
at the PSS software and instrument
department and is responsible for
education and customer training.
Peter Montag studied chemistry at
the University of Duesseldorf, Germany,
achieving his PhD at the Max Planck
Institute for Coal Research. He is the head
of the PSS contract analysis department
and responsible for hyphenated techniques.
Wolfgang Radke studied polymer
chemistry in Mainz, Germany, and
Amherst, Massachusetts, USA. He is
head of the PSS application development
department and is also responsible for
instrument evaluation and for customized
training courses.
techniques to generate contour plots, which
can also be used to quantify the different
species. Figure 5 shows an example where
linear and comb shaped molecules of
different composition have been successfully
separated.
Summary
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mass.
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can be analyzed by on-line viscometry,
while information on short-chain branching
can be gained by Fourier-transform infrared
spectroscopy (FT-IR) detection.
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molecules can be a problem. In such
cases advanced separation techniques or
E-mail: DHeld@pss-polymer.comWebsite: www.pss-polymer.com
8
7
6
5
4
3
2
1
5
1
2
3
4
5
6
7
8
6 7 8 9
SEC elution volume (mL)
Gra
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mL)
10 11 12
20
40
60
80
100
Figure 5: Separation of linear and comb shaped copolymers, which co-elute in GPC/SEC alone.
Tips and Tricks
17
Q&A: Blumberg2 News9 Tips and Tricks12 Jochems and Schad1899 122Beck and Delobel21 ISC Event Preview25 Training and Events27 Staff28252 272
Making Method Development Faster for the Analysis of Natural and Artificial FlavouringsPhilipp Jochems and Gesa Schad, Shimadzu Europa, Duisburg, Germany.
Vanilla is one of the most important
flavours worldwide and is widely used
in foods, beverages, and perfumes.
Natural vanilla extract contains up to
several hundred substances with vanillin,
vanillic acid, 4-hydroxybenzoic acid,
and 4-hydroxybenzaldehyde the major
components. As a result of continuously
increasing demand and the resulting high
costs of natural vanilla extracts, artificial
flavourings are often used instead.
Vanillin can be obtained through various
methods such as chemical synthesis,
biotransformation, or degradation of
waste sulphite liquors, as well as extraction
of natural vanilla pods. These artificial
flavours can contain synthetic vanillin,
ethyl vanillin, eugenol, guaiacol, vanillin
mandelic acid, and others.
As authenticity criteria for vanilla, the
ratios of the major components vanillin,
4-hydroxybenzaldehyde, vanillic acid,
and 4-hydroxybenzoic acid are frequently
used. In order to monitor the composition
and therefore quality of vanilla flavours
contained in food, an analytical method
needs to enable individual quantification of
any of these ingredients as well as possible
ingredients like the precursors from the
synthesis of vanillin.
A simple, rapid, and robust ultrahigh-performance liquid chromatography (UHPLC) method for the simultaneous determination of natural and artifi cial vanilla fl avouring substances as well as some precursors has been developed using an automated method scouting or method optimization workfl ow. The most suitable mobile phase and stationary phase combination was identifi ed in a scouting run. These conditions were used to create a two-dimensional model in computer simulation software. Temperature and gradient time were varied to establish the optimum fast and robust separation conditions. This approach resulted in a 5.5 min gradient method that allowed for fast screening of 11 compounds of interest.
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The article describes the computer-assisted development and optimization of a rapid ultrahigh-performance liquid chromatography (UHPLC) screening test for the separation and quanti� cation of natural and arti� cial vanilla � avouring substances as well as some precursors for the quality control
of vanilla products. This article illustrates the effectiveness and speed of software-assisted method development tools.
ExperimentalEquipment and Chromatographic Methods: For UHPLC method scouting,
a Nexera X2 Method Scouting System (Shimadzu) was used, consisting of two quaternary solvent pumps, autosampler, and column oven including a six-column switching valve. The system was also equipped with a high resolution photo diode array detector. Acetonitrile and water + 10 mM HCOONH4 (pH 2.8) were used as mobile phase. The different stationary phases applied for method scouting for the separation of 11 analytes relevant to vanilla products are displayed in Table 1.
Method scouting was performed in a sequence using 7.5 min gradient runs at 40 °C with combinations of organic and aqueous mobile phases (acetonitrile/H2O + 10 mM HCOONH4; gradient: 5–95%) on the six different columns, which cover a wide spectra of diverse stationary phase materials for reversed phase chromatography.
The phenyl-hexyl-column showed the most promising results with all analytes of interest at least partly separated using a mobile phase consisting of A: 10 mM ammonium formate in H2O (pH 2.8) and B: acetonitrile. These conditions were used to create a two-dimensional DryLab model (Molnár Institute) using 2 min and 6 min gradient runs at 25 °C and 50 °C as input data. These experiments resulted in
Table 1: Stationary phases used in method scouting.
Column
10 × 2.1 mm, 3-µm C18 column
10 × 2.1 mm, 2.5-µm phenyl-hexyl column
10 × 2.1 mm, 3-µm C18-column with aromatic selectivity
10 × 2.1 mm, 3-µm RP-cyano column
10 × 2.1 mm, 3-µm RP-amide column
10 × 2.1 mm, 3-µm pentafluorophenyl bonded column
6.50
T (o C
)
50
45
40
35
30
25
5 10
6.005.505.004.504.003.503.002.502.001.501.000.500.00
tG (min)
Figure 1: Colour-coded resolution map for UHPLC method development.
Jochems and Schad
1919
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OCTOBER 5 - 7, 2016
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DEADLINES
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The software predicted an optimum
separation with a minimum resolution of
the critical peak pair of 2.1 in a gradient run
from 5% to 70% B in 5.5 min at a fl ow rate
of 0.5 mL/min at 50 °C. A comparison of
the predicted chromatogram with an actual
sample run is displayed in Figure 2.
The advantage of a computer-assisted
method development approach compared
to a more traditional one lies in the time
savings because four gradient runs (see
above) are enough input for the software
to calculate the optimal conditions
(gradient, oven temperature) for the
method. This can mean lower costs
because of fewer working hours and
smaller solvent consumption. However,
there are some limitations such as when
developing a method with a two- (or
more) step gradient. In such cases the
simulations are not that precise as those
with a linear gradient. This is because the
data for the calculation are from a linear
gradient run and a computer simulation
is more precise the more similar the
conditions from the input data and the
simulation are.
Conclusion
A robust, fast, and sensitive UHPLC method
for the simultaneous separation of natural
and artifi cial vanilla fl avouring substances as
well as some precursors has been developed.
The method scouting experiment and further
optimization using computer simulation
software was able to save time and offered
visualization of the design space in a
resolution map to establish the most robust
separation method.
E-mail: shimadzu@shimadzu.euWebsite: www.shimadzu.eu
Philipp Jochems graduated in 2013 with
a master of science in applied chemistry
from the University of Applied Science in
Krefeld, Germany. In 2014 he was a scientifi c
assistant at the University Medical Center of
the Johannes Gutenberg University Mainz,
Germany, and since 2016 has worked as
a HPLC product specialist in the analytical
business unit of Shimadzu Europa in
Duisburg, Germany.
Gesa Schad graduated with a diploma in
chemical engineering from the Technical
University, NTA in Isny, Germany, in 2004
and as a master of science in pharmaceutical
analysis from the University of Strathclyde
in Glasgow, UK, in 2005. Until 2006 she
worked as a consultant in HPLC method
development and validation in an analytical
laboratory of the FAO/IAEA in Vienna,
Austria. She gained her doctorate for
research in pharmaceutical sciences at the
University of Strathclyde in 2010 and was
employed as an HPLC specialist in the R&D
department at Hichrom Ltd. in Reading, UK,
from 2009. Since 2013, she has worked as
a HPLC product specialist in the analytical
business unit of Shimadzu Europa in
Duisburg, Germany.
a colour-coded resolution map for simple
identification of the optimum separation
conditions (Figure 1). The figure shows
the calculated resolution (colour-coded)
for the different simulated combinations
of retention time (x-axis) and oven
temperature (y-axis).
1.2
1.1
1.0
0.9
0.8
0.7
0.6
0.5
0.4
0.3
0.2
0.1
0.0
1.2
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Van
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Van
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0.7
0.6
0.5
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0.3
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0.0
0.00 0.25 0.50 0.75 1.00 1.25 1.50 1.75 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00
DryLab Model
Actual RunTime (min)
0.00 0.25 0.50 0.75 1.00 1.25 1.50 1.75 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00
Time (min)
Figure 2: Comparison of predicted and actual chromatogram of the UHPLC analysis of natural and artifi cial vanilla fl avourings.
Jochems and Schad
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Preview of Topics at HPLC 2016, Part 4: State-of-the-Art MS Methods for Structural Assessment of mAbs and ADCs: From the Research Lab to Routine Characterization
This is the fi nal instalment in a series of four articles exploring topics that will be addressed at the HPLC 2016 conference in San Francisco, USA, from 19–24 June 2016.
Alain Beck1 and Arnaud Delobel2, 1Centre d’Immunologie Pierre Fabre (CIPF), Saint-Julien-en-Genevois, France, 2Quality Assistance SA, Donstiennes, Belgium.
Monoclonal antibodies (mAbs) are
highly complex tetrameric glycoproteins
that require extensive analytical and
structural characterization to become
drug candidates. This is also true for
antibody–drug conjugates (ADCs). These
immunoconjugates are based on highly
cytotoxic small-molecule drugs covalently
attached via conditionally stable linkers to
mAbs and are among the most promising
next-generation empowered biologics
for cancer treatment. ADCs are more
complex than naked mAbs, because the
heterogeneity of the conjugates adds
to the inherent microvariability of the
biomolecules. The development and
optimization of ADCs rely on improving
their analytical and bioanalytical
characterization by assessing several
critical quality attributes, namely the
distribution and position of the drug, the
amount of naked antibody, the average
drug-to-antibody ratio (DAR), and the
proportions of residual small-molecule
drug and drug-linker).1
As a result of advances in multilevel
(top, middle, bottom) state-of-the-art
mass spectrometry (MS) methods,
including native MS, ion mobility MS,2
capillary electrophoresis–electrospray
ionization–MS,3,4 two-dimensional
liquid chromatography–MS (2D LC–
MS),5 extended bottom-up,6 and
top-down sequencing,7 combined with
chromatographic and electrophoretic
techniques8 very precise characterization
21
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of biotherapeutics is now possible. Until
recently, however, these techniques
were considered suitable only for research
use. With the advent of robust and
user-friendly solutions (both hardware
and software), these techniques are
now amenable for routine use.
Examples of their application to the
characterization of mAbs and ADCs
are discussed below.
Middle-Level Characterization of mAbs
During their biosynthesis or during their
shelf life, mAbs can undergo many
modifications, such as glycosylation,
oxidation, deamidation, and C-terminal
lysine clipping, to name a few. In most
cases, these variants cannot be easily
identified at the intact antibody level,
because of limitations in chromatographic
separation and MS resolution.
A middle-up approach using IdeS
digestion (to yield Fab, Fc/2, and
light-chain fragments) combined with
a super macroporous reversed-phase
column enables quick and efficient
characterization of mAb variants.9 The
relatively low molecular mass of the
subunits (~25 kDa) allows accurate mass
determination by high-resolution MS as
well as top-down sequencing by electron
transfer dissociation (ETD). Using the
same sample preparation, the glycoforms
can also be separated and characterized
using an approach such as hydrophilic
interaction chromatography (HILIC) with
MS detection.10 An example of this
approach is presented in Figure 1 for
adalimumab.
Determination of Drug-to-Antibody
Ratio on Intact ADCs
The number of cytotoxic molecules
attached to an antibody (the
drug-to-antibody ratio, or DAR) is a critical
quality attribute of an ADC. It can be
determined by UV spectrophotometry or
hydrophobic interaction chromatography
(HIC) with UV detection,11–13 but these
methods are not universal and have some
drawbacks. Mass spectrometry can be a
universal tool to determine the DAR value,
whatever the coupling chemistry or the
Fc/2
Fd’
Fc/2
LC
Fd’0.008
(a)
(b)
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0
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0
9.43
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8 9 10
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11 12 13 14 15 16 17 18 19 20 21
Time (min)
Time (min)
Time (min)
Inte
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cou
nts
)Sig
nal (E
U)
Sig
nal (E
U)
LC
Oxidized species Pyroglutamic acid
Figure 1: HPLC–UV spectra obtained for adalimumab sample digested with IdeS and analyzed on (a) a reversed-phase or (b) a HILIC column.
Beck and Delobel
22
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Data integrity problems in pharmaceutical quality control
laboratories are driving more regulatory action than ever
before. It is obvious that something has changed to drive
all this activity. There is plenty of information available,
but much of it seems to confuse or frustrate rather than
clarify or help. In this webinar, we will provide clarity,
dispelling confusion by looking at the facts, based on a
study of available resources and direct interactions with
FDA staff and their consultants.
Key questions that indicate you should tune in to this web seminar
■ Do I understand what the new wave of data integrity enforcement means?
■ Are my laboratory software and processes ready for the increased scrutiny?
■ Do I understand what my responsibilities are for ensuring that both my vendor’s software and my organization’s processes will ensure data integrity?
After this webinar, you should be able to■ Articulate the drivers behind the “new” data integrity
regulatory enforcement actions
■ Communicate the current interpretations of existing regulations
■ Understand a methodology to evaluate your laboratory software
Who should attend■ Quality control laboratory managers, compliance and quality
managers, IT managers
For questions, contact Kristen Moore
at kmoore@advanstar.com
Presenter:
LOREN SMITH
Software Compliance
Program Manager
Agilent Technologies
Moderator:
LAURA BUSH
Editorial Director
LCGC
Back
by P
opul
ar
Deman
d!
Sponsored by
Presented by
Data Integrity in Pharma QC LabsWhat You Need to KnowON-DEMAND WEBCAST Aired June 9, 2016
Register for free at:
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size-exclusion chromatography (SEC)14
and detected by high-resolution
electrospray ionization MS. The DAR
can be automatically calculated by
software after deconvolution of the
multicharged electrospray spectrum.
When working with cysteine-linked
ADCs, native conditions must be used
to avoid disruption of noncovalent
interactions. In these methods, native
SEC–MS is commonly used, which requires
optimization of mobile phases and MS
conditions.
Characterization of mAbs and
ADCs by UHPLC–MS and HPLC–MS
Peptide Mapping
Peptide mapping with MS detection
is a common methodology for protein
characterization. It can be used for the
confirmation of the primary sequence,
the quantification of post-translational
modifications (PTMs), and the study of
disulphide-bond scrambling. Thanks to
commercially available software packages
dedicated to biopharmaceutical analysis,
the analysis of these MS data can be fully
automated.
Applied to ADC characterization,
peptide mapping is also a valuable tool
to localize conjugation sites and
determine site occupancy. ETD
fragmentation can be used to localize
conjugation sites for lysine-conjugated
ADCs on peptides containing several
lysine residues.
nature of the drugs.
After N-deglycosylation, the ADC is
desalted online using reversed-phase
high performance LC (HPLC) or
0 1 2 4 6 8
(5.1%) (0.8%) (1.9%) (38.4%-45.2%)(15.4%) (15.8%) (12.4%)
2e5
2.5e7
1.5e8
1e8
5e7
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)
Inte
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)In
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(co
un
ts)
Inte
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)
Retention time (min)
1412108642
25041.00
25017.00
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76400 76600 76800 77000 77200 77400 77600
24500 25000 25100 25200 25300 25400
25095.00 25139.00
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x2
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103277.00
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102756.00103104.00
104063.00
1.03e51.03e5 1.03e5 1.04e5 1.04e51.03e5
Mass (Da)
Mass (Da)
Figure 2: (a) HIC–UV chromatogram obtained for brentuximab vedotin and (b) deconvoluted mass spectra obtained by HIC–reversed-phase LC–MS for the two isomers of DAR4.
Beck and Delobel
23
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9. S. Sjögren, F. Olsson, and A. Beck, unpublished
data.
10. A. Peria, S. Fekete, A. Cusumano, J.L. Veuthey,
A. Beck, M. Lauber, and D. Guillarme,
unpublished data.
11. M. Rodriguez-Aller, D. Guillarme, A. Beck,
and S. Fekete, J. Pharm. Biomed. Bioanal. 118,
393–403 (2016).
12. A. Cusumano, D. Guillarme, A. Beck, and S. Fekete,
J. Pharm. Biomed. Bioanal. 121, 161–173 (2016).
13. S Fekete, J.L. Veuthey, A Beck, and D. Guillarme,
J Chromatogr. B, in press.
14. M. Rodriguez-Aller, A. Cusumano, A Beck, D.
Guillarme, and S. Fekete, J Chromatogr. B, in press.
15. M. Sarrut, A. Corgier, S. Fekete, D. Guillarme,
D. Lascoux, M.C. Janin-Bussat, A. Beck, and H.
Heinisch, unpublished data.
16. M. Sarru, M.C. Janin-Bussat, S. Fekete,
D. Guillarme, A. Beck, and H. Heinisch,
unpublished data.
17. R.E. Birdsall, S.M. McCarthy, M.C. Janin-Bussat,
M. Perez, J.F. Haeuw, C. Weibin, and A. Beck,
mAbs 8(2), 306–317 (2016).
Alain Beck is the Senior Director of the
Physico-Chemistry Department at the
Centre d’Immunologie Pierre Fabre (CIPF)
in Saint-Julien-en-Genevois, France.
Arnaud Delobel is the scientific manager
for biologics and the R&D team manager
at Quality Assistance SA, in Donstiennes,
Belgium.
E-mail: alain.beck@pierre-fabre.com / arnaud.delobel@quality-assistance.beWebsite: www.cipf.com/en / www.quality-assistance.com
quantification of small-molecule drugs.17
The application of this methodology
to brentuximab vedotin is presented in
Figure 2.
All these new techniques and case
studies will be discussed at the HPLC 2016
meeting in San Francisco in June.
References
1. A. Beck, G. Terral, F. Debaene, E. Wagner-
Rousset, J. Marcoux, MC Janin-Bussat, O. Colas,
A. Van Dorsselaer, and S. Cianferani, Exp. Rev.
Proteomics 13(2), 157 –183 (2016).
2. G. Terral, A. Beck, and S. Cianferani, J
Chromatogr. B, in press.
3. R. Gahoual, A. Beck, Y.N. François, and E.
Leize-Wagner, J. Mass Spectrom. 51(2), 150–158
(2016).
4. Y.N. François, M. Biacchi, N.Said, C. Renard, A.
Beck, R. Gahoual, and E. Leize-Wagner, Anal.
Chim. Acta 908, 168–176 (2016).
5. D. Stoll, J. Danforth, K. Zhang, and A. Beck,
unpublished data.
6. N. Gasilova, K. Srzentic, L. Qiao, B. Liu, A. Beck,
Y.O. Tsybin, and H.H. Girault, Anal. Chem. 88,
1775–1784 (2016).
7. R Gahoual, A. Beck, E. Leize-Wagner, and Y.N.
François, unpublished data.
8. A. Resenman, W. Jabs, A. Wiechmann, E
Wagner-Rouset, O. Colas, W. Evers, E. Belau,
L. Vorwerg, C. Evans, A. Beck, and D. Suckau,
mAbs 8(2), 318–330 (2016).
2D LC–MS Analysis of mAbs and
ADCs
Two-dimensional LC with MS detection
is widely used for the identification of
proteins from complex proteome samples
in many laboratories. Robust 2D–HPLC and
2D–UHPLC systems are now commercially
available, thus enabling the routine analysis
of biopharmaceuticals with this technology.
Two-dimensional LC can be used
to hyphenate MS-incompatible
chromatographic separations to mass
spectrometry detection: After a first
dimension using mobile phases containing
nonvolatile salts, the peak of interest is
sent to a second dimension consisting of
a reversed-phase column to desalt the
sample and separate potentially coeluted
species. This methodology (heart-cutting
2D LC–MS) can be routinely applied to
identify the different species detected
in size-exclusion and ion-exchange
chromatography of monoclonal
antibodies.
Another main application of 2D LC–MS
is the rapid on-line structural elucidation
of species observed in HIC distribution
profiles of cysteine-conjugated ADCs.15,16
The identification of the different species
is required to be able to determine the
DAR value based on the HIC–UV profile
as well as for the detection and the
Beck and Delobel
24
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The 31st International Symposium on Chromatography (ISC 2016)A preview of the upcoming 31st International Symposium on Chromatography (ISC 2016), which is due to be held 28 August–1 September at University College Cork, Ireland.
The 31st International Symposium on
Chromatography (ISC 2016) will be held from
28 August–1 September 2016 at University
College Cork, Cork, Ireland. This major
chromatography conference will be hosted
in the Emerald Isle for the fi rst time in its
history and will attract chromatographers
from around the world. The greater Cork
area is home to nine of the top 10 global
pharmaceutical companies in the world, and
seven out of 10 of the world’s best-selling
drugs are produced there. As a major
European centre for the life science industry,
Cork is an ideal choice to host ISC 2016.
Furthering the international appeal
of the ISC series, The Chromatographic
Society (ChromSoc) will be sponsoring key
presentations at ISC 2016 in celebration of
its 60th anniversary. ISC 2016 will also host
the prestigious “Award for Outstanding
Achievements in Separation Science”,
which is awarded to a preeminent
separation scientist by the California
Separation Science Society (CaSSS).
ISC 2016 provides the perfect forum for
scientific exchange between attendees
from academia, industry, and government
research institutions, as well as excellent
networking opportunities with up to 800
international delegates expected to attend.
The major focus of the symposium will be
on the impact and continuing contribution
25
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The Column www.chromatographyonline.com
of chromatography and separation science
to the pharmaceutical industry, food,
health, science, and medicine.
The major theme of ISC 2016 will be
the Innovation and Impact of
Chromatographic Separations on Science,
Industry, and Life. The symposium
programme reflects these themes, and
aims to highlight new challenges and
emerging opportunities in separation
science detection systems, methods,
and marketing solutions. The scientific
programme is set to be wide ranging and
diverse with topics including:
t� Pharmaceutical
t� Biomedical
t� Forensics
t� Environmental analysis
t� Process chromatography
t� PAT
t� Biomarkers
t� Diagnostics and clinical analysis
t� “Omics” technologies
t� New material science
t� Characterization, miniaturization, and
on-chip devices
t� Mass spectrometry
t� Food and health
t� Separation and sensing
t� Trace elements speciation
t� Supercritical fluid applications
t� Biopharmaceutical
International leaders in each of
these areas will provide inspiring and
thought-provoking presentations
to stimulate researchers. While an
international exhibition and vendor lecture
series on instrumentation and services
for chromatography, separation science,
and mass spectrometry will add another
integral part to the scientific programme.
ISC organizers look forward to
introducing attendees to the famous Irish
hospitality and the charms of Cork with
the nearby coastlines, beaches, hiking
routes, cycling routes, and world-class golf
courses offering exceptional scenery. While
the city’s wide array of hotels, restaurants,
traditional music and dancing, and, of
course, exceptional scientific conferences
complete the package.
On-line registration closes 25 August
2016.
Co-Chairs : Apryll Stalcup and Jeremy D. GlennonTel. : +353 1 280 2641E-mail : ISC2016@mci-group.comWebsite : http://www.isc2016.ie/
ISC Event Preview
26
Q&A: Blumberg2 News9 Tips and Tricks12 Jochems and Schad1899 122Beck and Delobel21 ISC Event Preview25 Training and Events27 Staff28252 272
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@6<9�)9(5+�0:�,=,9@;/05.�ThermoFisher.com/BeverageTesting
Training CoursesGC
The Theory of GC
Website: http://www.chromacademy.
com/gc-training.html
Complete GC and GC–MS
13–17 June 2016
The Open University, Milton Keynes, UK
Website: http://www.anthias.co.uk/
training-courses/completeGC
The Technique of GC in Three
Parts — Fundamentals/
Troubleshooting/Method
Development
14 July 2016
Reading, UK
Website: www.hichrom.co.uk
HPLC/LC–MS
The Theory of HPLC
On-line training from CHROMacademy
Website: http://www.chromacademy.
com/hplc-training.html
HPLC Troubleshooting and
Maintenance
6 July 2016
Kings College London, London, UK
Website: http://www.crawfordscientific.
com/training-online-calendar.asp
HPLC Masterclass2–24 August 2016
Laserchrom HPLC Laboratories Ltd,
Rochester, Kent, UK
Website: http://www.hplccourses.com/
index.htm (please mention The Column
when you make an enquiry)
SAMPLE PREPARATIONSampling Techniques for GC & GC–MS12 October 2016
The Open University, Milton Keynes, UK
Website: http://www.anthias.co.uk/
training-courses/sampling
GPC/SECLight Scattering and Viscometry Hands-On Training23–24 June 2016
Mainz, Germany
Website: www.pss-polymer.com
Please send your event and training course information to Kate Mosford kmosford@advanstar.com
27–30 June 2016
International Network of Environental Forensics Conference
Örebro Castle, Örebro, Sweden
Tel: +49 19 301 209
E-mail: ingrid.ericson@oru.se
Website: www.inef2016.com
26–29 September 2016
International Symposium on GPC/SEC and Related Techniques
Novotel, Amsterdam, Netherlands
Tel: +1 508 482 3129
E-mail: joy_longa@waters.com
Website: www.gpcevent.com
13–16 November 2016
27th International Symposium on Pharmaceutical and Biomedical Analysis
Guangzhou, P.R. China
E-mail: pba2016@hotmail.com
Website: www.PBA2016.org
30 November–2 December 2016
2nd ACROSS International Symposium on Advances in Separation
Science (ASASS 2)
Hobart, Tasmania, Australia
Chairman: Brett Paull
E-mail: brett.paull@utas.edu.au
Website: http://www.utas.edu.au/across
Event News
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Mission StatementThe Column (ISSN 2050-280X) is the analytical chemist’s companion within the dynamic world of chromatography. Interactive and accessible, it provides a broad understanding of technical applications and products while engaging, stimulating and challenging the global community with thought-provoking commentary that connects its members to each other and the industries they serve.Whilst every effort is made to ensure the accuracy of the information supplied, UBM EMEA accepts no responsibility for the opinions and statements expressed.Custom Reprints: Contact Brian Kolb at Wright’s Media, 2407 Timberloch Place, The Woodlands, TX 77380. Telephone: 877-652-5295 ext. 121. Email: bkolb@wrightsmedia.com.
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