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Merck

Sustainable Detergents for Virus Inactivation and Cell Lysis

Our new offer of Emprove®detergents for a sustainable Triton™ X-100 detergent replacement for biomanufacturing

*Triton™ is a trademark of The Dow Chemical Company

Detergents Use In Bioprocessing

Detergents, such as zwitterionic non-ionic surfactants, are used in biomanufacturing processes for a variety of applications including viral inactivation and cell lysis.

Deviron® C16
Deviron® 13-S9

Looking For A Triton™ X-100 Alternative?

Triton™ X-100 (4-tert-octylphenol polyethoxylate), a non-ionic detergent, is commonly used for virus inactivation and cell lysis as it disrupts the lipid membrane of cells and enveloped viruses.

Triton™ X-100 is the main detergent templated in biomanufacturing processes, but as of January 2021, its unauthorized use has been prohibited in the European Union by the European Commission, due to its listing to REACH (Registration, Evaluation, Authorization and Restriction of Chemicals).

In response to the global implications of restrictions on Triton™ X-100 detergent, we have developed the Deviron® portfolio of sustainable detergents.

The Deviron® Detergent Portfolio as a Sustainable Triton™ X-100 Alternative

Our Deviron® portfolio of sustainable detergents should be considered as a primary alternative to Triton™ X-100 by all biologics manufacturers.

To address the need for alternatives to Triton™ X-100 that do not require a REACH authorization, we have launched the Deviron® C16 detergent and the Deviron® 13-S9 detergent.

 

Deviron® C16 Emprove® Evolve Detergent

Deviron® 13-S9 Emprove® Expert Detergent

Triton™ X-100 Emprove® Expert Detergent

Product code

108693

108694

108643

Surfactant type

Zwitterionic (pI 8.9)​

Non-ionic​

Non-ionic​

Chemical name

N,N-Dimethyltetradecylamin-N-oxid

Alcohols, C11-15-secondary, ethoxylated

4-(1,1,3,3-tetramethylbutyl)phenol ethoxylated

CAS number

3332-27-2

68131-40-8

9036-19-5

Critical Micelle Concentration (CMC)

0.002-0.003 % wt​ (24 °C)

0.005 %​ wt (24 °C)

0.013 % wt (24 °C)

Biodegradibility (Readily biodegradable: >60 % within 28 days) OECD 301B

Readily biodegradable
88 % degradation

Readily biodegradable
74 % degradation

Substance of Very High Concern as defined by the REACH1

Virus inactivation efficiency (LRV ≥ 4).
(Detergent ≥0.5 % wt
T=15-25 °C)

>5 LRV

>5 LRV

Benchmark

Cell lysis efficiency for viral vector processing

Better or comparable to benchmark

Better or comparable to benchmark

Benchmark

Endotoxins removal for plasmid manufacturing:
Natrix® Q chromatography membrane/ detergents added to wash buffer supplement

99.997 % removal

99.969 % removal

Benchmark

Emprove® & MQ level

Emprove® Evolve, MQ400, (ISO9001 or equivalent)

Emprove® Expert, MQ500, (IPEC-PQG GMP)​

Emprove® Expert, MQ500, (IPEC-PQG GMP)​

Composition

30 % wt. water solution

Pure substance (100 %)

Pure substance (100 %)

Available formats

1L, 2.5 L, 25 L

1 L, 2.5 L, 25 L

1 L, 2.5 L, 25 L, 190 L

Customized Certificate of Analysis options

Available

Available

Available

Detection method

HPLC-ELSD method available

HPLC-ELSD method available

 

*Benchmark: Triton™ X-100 detergent

KEY CONSIDERATIONS FOR DETERGENT SELECTION IN BIOPROCESSING

Several factors should be considered when evaluating detergents in biomanufacturing.

These include:

  • Biodegradability (REACH* Compliance) and lack of harmful degradation products
  • Viral inactivation (5 Log Reduction Value)
  • Cell lysis efficiency
  • Minimal impact on product functionality or quality
  • Lack of process interference
  • Effective removal by downstream operations
  • Availability of sensitive detection method
  • Manufacturability of high volumes as IPEC-PQG GMP quality material
  • Impurity profile compatible with parenteral usage

*Registration, Evaluation, Authorisation and Restriction of Chemicals

Virus Inactivation for CHO Cell Harvest and Human Plasma Protein Purification

Viral safety is a major concern for biotherapeutic manufacturers.

  • Cell-based processes may produce endogenous retroviral particles, and adventitious viruses can be introduced from contaminated source materials or during the manufacturing process.
  • Human plasma-derived products are at risk of containing viruses, despite extensive screening of donation material.

Detergent-mediated viral inactivation is widely used in multiple biotherapeutic production processes as part of an overall virus safety strategy.

Our Deviron® portfolio shows efficient viral inactivation for both matrices with different model viruses and operating conditions.

Virus inactivation was assessed in mAb-containing CHO clarified harvest and human plasma matrices with XMuLV (Xenotropic Murine Leukemia Virus).

  • The standard viral inactivation practice is described in the ASTM E3042-16.
  • mAb processes >0.5%, no solvent and incubation time >60 min.
  • Plasma processes typically utilize 1% detergent /0.3% solvent TnBP and longer incubation time (4-6 h).

Deviron® C16 and Deviron® 13-S9 demonstrate effective viral inactivation (LRV >5) in all conditions tested. Additional data with PRV and BVDV viruses are available. 

Viral inactivation (XMuLV) with Deviron® C16 in CHO clarified harvest at 22°C compared to Triton™ X-100

Deviron® C16 in CHO clarified harvest at 22°C

Viral inactivation (XMuLV) with Deviron® 13-S9 in CHO clarified harvest at 15° compared to Triton™ X-100

Deviron ® 13-S9 in CHO clarified harvest at 15°C

Viral inactivation (XMuLV) with Deviron® C16 in neat plasma with 0.3% TnBP at 22°C compared to Triton™ X-100

Deviron ® C16 in neat plasma with 0.3% TnBP at 22°C

Viral inactivation (XMuLV) with Deviron® 13-S9 in cryo-poor plasma with 0.3% TnBO at 22° compared to Triton™ X-100

Deviron ® 13-S9 in cyro-poor plasma with 0.3% TnBP at 22°C compared to Triton™ X-100

Viral inactivation (XMuLV) with Deviron® C16 in CHO clarified harvest at pH 5.5 and pH 8.5 at 22°

Deviron ® C16 in CHO clarified harvest at pH 5.5 and pH 8.5 at 22°C

Viral inactivation (XMuLV) with Deviron® 13-S9 in IgG at pH 6 and pH 8 at 15°

Deviron ® 13-S9 in IgG at pH 6 and pH 8 at 15°C

Cell Lysis for AAV Manufacturing

For biomanufacturing production of viral vectors used as therapeutics, a lysis step is needed to release the viral particles of interest. For Adeno Associated Virus (AAV) vectors used for gene therapies, host cells (HEK293 or Sf9 cells) used for production of viral vectors are often lysed using detergents like Triton™ X-100 or Polysorbates 20/80.

We introduce two new cell lysis options with our Deviron® detergent portfolio.

Cell lysis efficiency was assessed with HEK293 cells. After one day of cultivation, the cells were transfected with the selected plasmid polyethylenimine (PEl) complexation. After 72 hours of cultivation, the cells were harvested and lysate prepared according to the following procedure:

  • Detergent concentration: 0.5% wt/vol
  • Detergent: Tween™ 20 and Triton™ X-100 (benchmarks), Deviron® C16 and Deviron® 13-S9
  • Nuclease: 25 U/mL Benzonase® with 2 mM MgCl2
  • Lysis time: 2 hr
  • Temperature: 37 °C

Following incubation, each sample was counted with ViCell and total cell count and viability determined.

Cell viability for HEK293 Adherent cells in AAV2 production

HEK293 Adherent cells – AAV2

Cell viability for HEK293 suspension cells in AAV5 production

HEK293 suspension cells – AAV5


THE EMPROVE® PROGRAM FOR DETERGENTS

Roadmap for Choosing Fit-For-Purpose Raw Materials

As a supplier of thousands of raw materials for biologics manufacturing, we know the importance of quality and consistency. Our Emprove® program is designed to provide customers with the transparency and information needed to select fit-for-purpose raw materials and access to detailed documentation to meet regulatory requirements. 

DEVIRON® C16 Emprove® Evolve 

Provides detailed and transparent supply chain information and documentation to support risk assessments for critical raw materials used in manufacturing processes. Non-GMP but Manufactured utilizing GMP concepts and elements; ISO9001 or equivalent.

DEVIRON® 13-S9 Emprove® Expert

Addresses higher risk applications, where the lowest microbiological and endotoxin levels are of utmost importance. Released with IPEC-PQG GMP.


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