Multiparameter Sensor System for Single-UseBioreactorsStrategische Übungen – (Electro)chemical Sensors

ETHZ, 23.10.2018

Caspar DemuthInstitute of Chemistry und BiotechnologyZurich University of Applied Sciences

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C. Demuth | Strategische Übungen | ETHZ | 23.10.2018 2

Top 10 largest selling pharmaceutical products (2015)

https://en.wikipedia.org/wiki/List_of_largest_selling_pharmaceutical_products

Rank Drug Trade name Type Main indications CompanySales (109USD /year)

∆ vs 2014

1Adalimumab Humira Biologic Rheumatoid arthritis AbbVie Inc. 14,012 1,469

2Ledipasvirsofosbuvir Harvoni

Small molecule Hepatitis C Gilead Sciences 13,864 11,737

3Etanercept Enbrel Biologic Rheumatoid arthritis Amgen/Pfizer 8,697 4,0094Infliximab Remicade Biologic Crohn's Disease Johnson & Johnson 8,355 1,487

5RituximabMabtheraRituxan Biologic

Lymphoma/Leukemia/Autommune disorders Roche 7,115 1,456

6Insulin glargine Lantus Biologic Diabetes mellitus Sanofi 7,029 517Bevacizumab Avastin Biologic Metastatic cancers Roche 6,751 2708Trastuzumab Herceptin Biologic Breast cancer Roche 6,603 265

9Lenalidomide RevlimidSmall molecule Multiple myeloma Celgene 5,801 821

10Sofosbuvir SovaldiSmall molecule Hepatitis C Gilead Sciences 5,276 (5,007)

C. Demuth | Strategische Übungen | ETHZ | 23.10.2018 3

Spot the differences!

https://www.teachchemistry.org/content/aact/en/periodical/issues/may-2016/chemistry-fun.html

C. Demuth | Strategische Übungen | ETHZ | 23.10.2018 4

Spot the differences!

Stainless steel bioreactor

ZHAW

Single use bioreactor

ZHAW

C. Demuth | Strategische Übungen | ETHZ | 23.10.2018 5

Task

Propose a multiparameter sensor system for real-time, in situ measurements in single-use bioreactors for critical quantities in bioprocess

control

Include analytical as well as more general aspects

1. Relevance of critical parameters (pH, pO2, glucose, lactate, ammonium)

2. Requirements of a multiparameter sensor system

3. Outline of a sensor system

Measurement principles

Analytical performance vs. requirements

Overall suitability

C. Demuth | Strategische Übungen | ETHZ | 23.10.2018 6

Relevance of critical bioprocess parameters

pHOptimum pH guarantees efficient productivity

and high yield

Dissolved oxygen (pO2)Respiration! Essential in aerobic processes

GlucoseMain substrate in biotechnological cultivations

Lactate and ammoniumMetabolites, harmful or toxic to cells

pH

µ max

Maximum specific growth rate (μmax) ofE. coli as a function of pH

S.J. Pirt, Principles of microbe and cell cultivation, Blackwell, Oxford, 1975

C. Demuth | Strategische Übungen | ETHZ | 23.10.2018 7

Sensor specifications in bioprocesses

Wetted parts

Biocompatibility,leachables & extractablesCompatibility with

process

Process conditions(p, T, t, SIP,…)

Integration

Ports, software, protocols

Quality

Procedures, approvals,

validation, safetyCosts

«Total cost ofownership»

Analytical specifications

Comparability, robustness, accuracy,…

C. Demuth | Strategische Übungen | ETHZ | 23.10.2018 8

Sensor requirements in single-use reactors

Wetted parts

Use of heat labile materials (polymers,

Compatibility withprocess

«mild» conditions(gamma irradiation)

Integration

Sensor ist is part ofreactor; different

ports (no standard)

Quality

Different handlingand procedures

Costs

Single Use

Analytical specifications

Comparability, robustness, accuracy,…

C. Demuth | Strategische Übungen | ETHZ | 23.10.2018 9

Selected requirements in single-use bioreactors

Process temperature Typically 37°CProcess pressure Typically 1 barProcess duration Days to weeksSterilization procedure Gamma irradiation (25 kGy)Shelf life 1 yearCalibration Factory precalibrated, calibration interval > 1 d

pH pO2 Glucose Lactate AmmoniumMeasurement range

4-8 0-1000 mbar

0.1-30 gl-1 0.1-10 gl-1 0.01-0.5 gl-1

Response time Seconds Minutes Minutes Minutes Minutes

Criticalinterferences

Alkali ions Reducingagents

Reducingagents

K+

C. Demuth | Strategische Übungen | ETHZ | 23.10.2018 10

Available measurement techniques

No single working principle for all parameters combination of electrochemical

and/or optical sensors necessary

A: Electrochemical (potentiometric and amperometric)B: Electrochemical and optical

Potentiometricsensors

Glass membrane pHLiquid membrane ISE pH, NH4+

Severinghaus type NH4+

Amperometricsensors

Clark electrode pO2Enzymatic biosensor Glucose, lactate

Optical sensors Fluorescence/absorbance ofpH sensitive dye

pH

Luminescence quenching pO2Severinghaus optical NH4+

Ion exchange optode NH4+

C. Demuth | Strategische Übungen | ETHZ | 23.10.2018 11

Critical aspects

Potentiometric sensors: reference electrode (e.g.

Ag/AgCl)!

pH glass electrode: shelf life, hydration of gel layer

Optical pH sensors: drift, long term stability, bleaching,

dependence on ionic strength

Ammonium sensors: measurement range and

selectivity (nonactin-based sensors)

Enyzmatic biosensors: drift and long term stability

Biocompatibility of selected materials

InSUS 307, Mettler Toledo AG / Thermo Scientifc

Nonactin

C. Demuth | Strategische Übungen | ETHZ | 23.10.2018 12

Proposed sensor system

Other solution are feasible as well!

pH pO2 Glucose Lactate Ammonium

Measurementprinciple

optical optical enzymaticbiosensor, amperometric

enzymaticbiosensor, amperometric

optical

Process temperature Process pressure Process duration () () ()Sterilizationprocedure

Shelf life Calibration Measurement range ()Response time Interferences

C. Demuth | Strategische Übungen | ETHZ | 23.10.2018 13

Optical sensors for pH and pO2

X.D. Wang et al, Chem Soc Rev 2014, 43, 3666–3761.

B.J. Müller et al., Sensors and Actuators B 216 (2015) 527–534

S.M. Borisov et al, Analyst 2010, 135, 1711-1717.

C. Demuth | Strategische Übungen | ETHZ | 23.10.2018 14

Outlook: Parameters in bioprocess control

«relevance»

«rea

dyne

ss»

physical

chemical

biological

pT pH

conductivity pO2pCO2

sustratesmetabolites

biomassproductscell size &

morphology

C. Demuth | Strategische Übungen | ETHZ | 23.10.2018 15

Process Analytical Technology

https://www.fda.gov/downloads/drugs/guidances/ucm070305.pdf

C. Demuth | Strategische Übungen | ETHZ | 23.10.2018 16

Voltammetric pH measurement

Immobilization of pH sensitive and pH insensitive

redox active components

“calibration free” non glass all solid-state sensor

V.G.H. Lafitte et al, Electrochem Commun 2008, 10,1831–1834

C. Demuth, Chem. Unserer Zeit, 2014, 48, 60 – 67

ww

w.s

enov

asys

tem

s.co

m

C. Demuth | Strategische Übungen | ETHZ | 23.10.2018 17

Optical measurements in single-use bioreactors

C. Busse et al., Eng. Life Sci. 2017, 17, 940-952

Combination of different

measuring modes

- Optodes- Fluorescence, Raman- Absorption (UV/VIS)- ATR (IR)

Standardized in SU

bioreactors needed

C. Demuth | Strategische Übungen | ETHZ | 23.10.2018 18

Sensors for online measurement of biomass concentration

Dielectric or impedance spectroscopy

(commercially available)

www.aber-instruments.co.uk

J. Carvell at al, BioProcess International 14(3) 2016, 40-48

Online measurement in root cultures

C. Demuth et al., Heiligenstädter Kolloquium 2016

C. Demuth | Strategische Übungen | ETHZ | 23.10.2018 19

RFID sensors for SU bioreactors

passive radio frequency identification (RFID) tags on single-use

bioreactors

Sensor antenna changes characteristic resonance, dependent on

physical/chemical properties of process medium

Parameters: pH, conductivity, temperature, biomass,…

R.A. Potyrailo et al, Meas. Sci. Technol. 2011, 22, 082001.

R.A. Potyrailo et al., 2011 IEEE International Conference on RFID-Technologies and Applications

C. Demuth | Strategische Übungen | ETHZ | 23.10.2018 20

Sensors for single-use bioreaktors –Challenges

Lifetime Critical for optical pH sensors and biosensors

Recalibration during process (risk of contamination, expenses)

Comparability Comparison of sensors based on different principles

Comparison of online and offline methods is problematic!

Traceable procedures and standards are necessary

Standardization Conventional bioreactors: Standard ports and connectors available

Will allow users to choose best suited sensors for their appilcation

C. Demuth | Strategische Übungen | ETHZ | 23.10.2018 21

Finally…

Single Use: Perfect situation for sensor development growing market

“Disruptive” production technology, new perspectives

Potential for new measurement techniques is not fully exploited yet

Sensor specifications and requirements Not only technical / research viewpoint!

Trends in the process industry favour application of sensors Quality (PAT, QbD)

Automation

Parallelization, Miniaturization

Foliennummer 1Top 10 largest selling pharmaceutical products (2015)Spot the differences!Spot the differences!TaskRelevance of critical bioprocess parametersSensor specifications in bioprocessesSensor requirements in single-use reactorsSelected requirements in single-use bioreactorsAvailable measurement techniquesCritical aspectsProposed sensor system Optical sensors for pH and pO2Outlook: Parameters in bioprocess controlProcess Analytical TechnologyVoltammetric pH measurementOptical measurements in single-use bioreactorsSensors for online measurement of biomass concentrationRFID sensors for SU bioreactorsSensors for single-use bioreaktors – ChallengesFinally…Thank you!Backup SlidesSensoren für Single-Use-BioreaktorenMehrwegsensoren in Einweg-Reaktoren?