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Insulin Adsorbent
High performance, alkali resistant and robustsynthetic ligand affinity chromatographyadsorbent designed for…..
…..capture and purification ofnaturally occurring forms of insulin andinsulin analogues from a variety ofsources
WW
W.P
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ETIC
BIO
SCIE
NCE
S.CO
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Non-bound
Elution
Strip
INSULIN ADSORBENT - Technology
A range of insulin analogues have been commercially developed byvarying the amino acid sequences of natural insulin, hence alteringits ADME (adsorption, distribution, metabolism and excretion)characteristics in glycaemic control. Insulin analogues are generallyeither faster-acting (i.e. offering a better post-prandial profile) orlonger-acting (i.e. to supply the basal level during the day orproviding overnight control) than natural insulin, offering advantagesfor diabetes mellitus management.
Insulin analogues are recombinant proteins, often produced inmicrobial expression systems and are commonly purified using acombination of downstream process steps such as reverse-phaseHPLC, ion exchange or size exclusion chromatography. Due to thegrowing demand to reduce the number of process steps and overallcost of purifying insulin analogues, ProMetic BioSciences Ltd (PBL)has developed a high performance synthetic ligand affinitychromatography adsorbent which is compatible with large-scaleprocess applications.
PBL’s Insulin Adsorbent shares the same chemistry as PBL’s provenMimetic Ligand™ adsorbents and provides high yields and purities fornaturally occurring forms of insulin and insulin analogues from avariety of sources and is alkali resistant, robust, non-toxic and isdesigned for use in cGMP manufacturing of biological molecules.
EASY TO PACK
PBL’s Insulin Adsorbent comprises a novel small synthetic(chemical) ligand coupled to PuraBead® 6HF near mono-disperse beaded agarose which provides an easy to packchromatography adsorbent with asymmetry results in the rangeof 0.8 to 1.2 as shown in the chromatogram below.
Volume (mL)
HETP - 0.041 cm
Plates/m - 2435
Asymmetry - 0.98
EXCELLENT FLOW PROPERTIES
PBL’s Insulin Adsorbent near mono-disperse agarose beadsprovide excellent flow properties and can be processed atoperational flow rates of up to 600 cm/hr with low column backpressures, as shown in the graph below.
Flow
rat
e (c
m/h
r)
Column Parameters: 10 cm diameter column, 17.6 cm bed heightPacking Buffer: 0.2 M sodium acetate, pH 5.5
Pressure(psi)
ADVANTAGES
5000 1000 1500 2000
Column Parameters: 10 cm diameter column, 17.6 cm bed height
700
500
400
100
200
300
600
800
02 10 140 4 8 12 166
KEY FEATURES
UNIFORM PARTICLE SIZE�PURABEAD® 6HF�
EASY TO PACK
EXCELLENT FLOW PROPERTIES
SELECTIVE FOR MOST FORMSOF INSULIN INCLUDING �SINGLE�CHAIN & TWO�CHAININSULIN
HIGH PURITIES & YIELDS
COST EFFECTIVE & RELIABLE
EASY TO CLEAN �ROBUST,LONG LIFE ADSORBENT�
Abs
orba
nce
(mA
u) 80
60
40
20
0
100
1&2
ADSORPTION
WASH
DESORPTION
Ligand: Synthetic derivative structure
Mean particle size: 90 ± 10 µm
Base matrix: Highly cross-linked 6% near mono-disperse beaded agarose (PuraBead® 6HF)
Dynamic binding capacity: Up to 25 g of insulin/L of adsorbent (dependent of feedstock)
Packing conditions: Equilibration buffer at a constant pressure of 1.5 bar (~22 psi)
Operational flow rate: Up to 600 cm/hr
Chemical stability: Stable in all commonly used buffers and solutions
Sanitization: 0.5 M NaOH
Storage: 2-30°C, 20% ethanol (v/v)
PROPERTIES
HIGH PERFORMANCE
The chromatogram and SDS-PAGE below show the capture and purification of recombinant single-chain insulin using PBL’s InsulinAdsorbent. Pre-conditioned yeast cell culture supernatant, containing recombinant single-chain insulin, was loaded onto PBL’sInsulin Adsorbent (5 mL column volume, 1.6 cm diameter, 2.5 cm bed height) and provided 95% purity and high yield in a singlecapture step. PBL’s Insulin Adsorbent was then followed by a polishing step using PBL’s SP PuraBead® HF which gave 99% purity.
Volume (mL)
Abs
orba
nce
mA
U)
150 2000 50 100
2000
1500
1000
500
0
2500
3000
3500
4000Non-bound
Elution
Strip
Equilibration Buffer: 0.1 M sodium acetate, pH 5.4Elution Buffer: 0.3 M acetic acid Strip: 1 M acetic acid, 20% ethanol
Clean in Place (CIP): 0.5 M NaOH
1 2 3 4 5 6 7
191 kDa
97
64
51
39
28
1914
6 Recombinant insulin
Non-reduced SDS-PAGE of the processfractions for the purification ofrecombinant single-chain insulin usingPBL's Insulin Adsorbent. Lane 1 – mwmarker; Lane 2 – load; Lane 3 – flowthrough; Lane 4 – post load wash; Lane5 – elution (dilution 1 in 3); Lane 6 –strip; Lane 7 – CIP
95% PURITY
INSU
LIN
AD
SORB
ENT
BULK ADSORBENT SLURRY -
For column packing, from BENCH SCALE (25 mL, 100 mL and500 mL pack sizes) to PROCESS SCALE (single batches ofadsorbent of up to 275 L).
PURAPLATE™ -
96 column block format for high throughput processoptimization and rapid method development (0.25 mLcolumn volume).
PRE-PACKED COLUMNS -
PBL’s Insulin Adsorbent is available in a range ofpre-packed column sizes from 1 mL to >10 L, for attachmentto chromatography workstations.
PBL supply bulk adsorbent gel slurry for both developmental and regulated cGMP manufacturing
applications in volumes sufficient to meet the requirements of the largest biomanufacturing
processes. PBL’s Insulin Adsorbent is supported by a comprehensive regulatory support file and
is manufactured in our controlled environment manufacturing facility under an accredited
ISO 9001 quality management system.
SALES & TECHNICAL SUPPORT
Please contact us for further sales or technical support information
regarding PBL’s Insulin Adsorbent or any of our other innovative
biochromatography products and services – let us show you how we
can provide you with solutions to your biomolecular purification
needs today:
PuraPlate™, Mimetic Ligand™, ProMetic BioSciences Ltd and the PBL Logo are trademarks of ProMetic BioSciences Ltd. PuraBead® is registered with the U.S. Patent& Trademark Office. 5 mL column image was kindly provided by Atoll GmbH. 50 mL and 500 mL column images were kindly provided by Repligen Bioprocessing.Large stainless steel process column images were kindly provided by Proxcys.
Copyright © 2013 ProMetic BioSciences Ltd. All rights reserved. Issue - 101013
ORDERING
CONTACT US
For sales: [email protected]
For tech support: [email protected]
Website: www.prometicbiosciences.com
NORTH AMERICA
1330 Piccard DriveSuite 201Rockville, Maryland 20850, USA
Tel: +1 301 251 8821 Fax: +1 301 251 8826
The Freeport,Ballasalla, Isle of Man,IM9 2AP, British Isles
Tel: +44 (0) 1624 821450 Fax: +44 (0) 1624 821451
EUROPE/RoW
YourPurificationSolutions
¹ http://en.wikipedia.org/wiki/Insulin_analog ² http://www.ncbi.nlm.nih.gov/pubmed/11853368
