Lyophilization of Cryogenized Cellular Compositions Under Pressure

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Solution Overview

Problem

Existing methods for freezing and lyophilizing cells, particularly lactic acid bacteria, result in significant viability loss due to intracellular crystal formation and lack of control over water and oxygen levels, leading to poor long-term stability and viability of lyophilized products.

Innovation Solution

A method involving cryogenic freezing under pressure, which dissolves gases in the cellular matrix before lyophilization, allowing for better preservation of cell integrity and viability by maintaining anaerobiosis and controlling critical parameters like cell concentration and water content.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Reliability

If conventional freezing methods are used, then equipment requirements are simple and operation is easy, but significant cell viability loss occurs due to intracellular crystal formation

Engineering Contradiction:

Improvecell viabilityVSAvoidintracellular crystal formation

Core Design Contradiction:

ReliabilityVSObject-affected harmful factors

Solution Approach 1:

The invention changes the physical parameters of freezing by applying high pressure (10-1000 bar) during cryogenic freezing, which fundamentally alters the freezing behavior to prevent intracellular crystal formation while maintaining simplicity of operation

Inventive Principle:

Principle #35Parameter changes

Solution Approach 2:

The invention utilizes phase transition of gases to liquids under pressure during the freezing process, where gas is dissolved in the cellular matrix under high pressure before freezing, preventing harmful crystal formation while preserving cell viability

Inventive Principle:

Principle #36Phase transitions

2Reliability

If lyophilization is used to store cells, then storage stability is improved and ambient temperature storage is enabled, but the process is long, expensive and very energy-consuming

Engineering Contradiction:

Improvestorage stabilityVSAvoidenergy consumption

Core Design Contradiction:

ReliabilityVSUse of energy by stationary object

Solution Approach 1:

The invention performs preliminary action by dissolving gas in the cellular matrix under high pressure before the freezing and lyophilization steps, which prevents water and oxygen from compromising cell quality during subsequent processing, thereby reducing energy consumption and processing time while maintaining storage stability

Inventive Principle:

Principle #10Preliminary action

3Reliability

If pressure is lowered during freezing, then some improvement in cell survival is achieved, but long-term stability at ambient temperature is not obtained

Engineering Contradiction:

Improvecell survival rateVSAvoidlong-term stability

Core Design Contradiction:

ReliabilityVSDuration of action of stationary object

Solution Approach 1:

The invention changes the pressure parameter from low pressure to high pressure (10-1000 bar) during the freezing step, which fundamentally improves both cell survival rate and long-term stability at ambient temperature by controlling water and oxygen levels in the product

Inventive Principle:

Principle #35Parameter changes

Applied Scientific Principles

This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.

Function Achieved in This Case

This method significantly improves the long-term viability and stability of lyophilized cells, enabling them to be stored at ambient temperature for extended periods with enhanced production yields and reduced damage, while eliminating the need for cryoprotectants and allowing for quicker and more efficient processing.

Implementation Method 1

the freezing step is carried out by cryogenics, and which comprises: a) providing a cellular composition comprising cells in an aqueous medium; b) dissolving a gas in said composition by passage through a dense zone of gas molecules

Methodology Applied

Scientific EffectCryogenics: Cryogenics

Implementation Method 2

dissolving a gas in said composition by passage through a dense zone of gas molecules, such a density being obtained (i) either owing to the flow of gas generated by the evaporation of a cryogenic fluid, (ii) or by raising the pressure

Methodology Applied

Scientific EffectDissolution: Solvation

Implementation Method 3

c) cryogenizing said gas-rich composition obtained in step b) at a pressure that makes it possible to keep said gas dissolved for obtaining frozen granules, particles or beads; d) lyophilization of said frozen granules, particles or beads to obtain a lyophilized cellular preparation

Methodology Applied

Scientific EffectLyophilization: Freeze Drying

Data Source

PatentUS20230017228A1Method of lyophilization of a cryogenized cellular composition containing dissolved gas

Publication Date: 2023.01.19 GENIALIS

AI summary

The invention relates to the field of lyophilizates of biological materials. More particularly, the invention relates to a new method for preparing a lyophilizate of cells comprising a step of cryogenics “under pressure”. In a preferred embodiment, this method is applied to lactic acid bacteria.