Apparatus and method for measuring degradation kinetics of a biomaterial in aqueous solution below the normal freezing temperature

Inventors

SILVESTRE DUARTE, Andreia Filipa • GIL SENA REGO, Pedro • DE BRITO ESTRELA, Rui

Assignees

Smartfreez Ltda

Abstract

This present disclosure describes an apparatus and a method for measuring degradation kinetics of a biomaterial in aqueous solution below the normal freezing temperature. Further includes new systems comprising high-pressure containers with nucleating-locks, a temperature-control holder for the high-pressure containers, a holder for the nucleating-locks and an insulated low-temperature gradient chamber. A method is described comprising several steps such as filling of the high-pressure container with the biomaterial solution to be slightly below freezing temperature, closing of the container with a nucleating-lock, which is below freezing temperature, allowing the container to reach the reaction temperature, removing the container at selected time and bringing it to above freezing temperature and opening the container to recover the biomaterial solution.

Core Innovation

The disclosure relates to an apparatus and method for measuring degradation kinetics of a biomaterial in aqueous solution below the freezing temperature of the biomaterial at atmospheric pressure. A high-pressure container is filled with the biomaterial in aqueous solution and the biomaterial is submitted to a temperature below the freezing temperature at atmospheric pressure, while a nucleating-lock is used to keep the biomaterial solution below freezing without inducing nucleation until the high-pressure container is closed.

The high-pressure container is closed with a nucleating-lock comprising a compressing-lid and a plug with a retainer, where the retainer prevents detachment of the plug and insulates the plug from the compressing-lid below the freezing temperature. The tip of the plug contacts the surface of the biomaterial solution to induce nucleation, whereby an ice layer is formed acting as a natural piston and ensuring isochoric conditions.

After reaching a pre-determined temperature different from the freezing temperature at atmospheric pressure, the high-pressure container is removed at a selected time, placed at a temperature higher than the freezing temperature at atmospheric pressure, opened, and the biomaterial in aqueous solution is recovered for degradation kinetics measurement. The disclosure further describes an insulated low-temperature gradient chamber and a temperature-control holder for controlling the high-pressure container temperature, including a holder for cooling nucleating-locks to a nucleation temperature.

An example apparatus includes a nucleating-lock with a plug comprising a cavity that comprises a nucleating agent, optionally coated with a water-permeable porous material. The disclosure further describes a configuration with multiple cavities, heat exchange on a chamber bottom surface, and a temperature-control holder comprising a heater, and states that this addresses limitations of earlier isochoric methods, including complexity, reproducibility, cleaning/sterilization, air-bubble control, and thermal-history consistency across many small-volume samples.

Claims Coverage

The independent claims are directed to three main inventive areas: a sample preparation method that uses a nucleating-lock to form an ice layer acting as a natural piston for isochoric conditions, an apparatus combining an insulated low-temperature gradient chamber with temperature-controlled operation and nucleation induction, and a nucleating-lock structural configuration with an insulating retainer and a temperature-control holder with a heater. Across the independent claims, the inventive features are concentrated in the nucleating-lock, the thermal control in an insulated gradient chamber, and the resulting formation of an ice layer to ensure isochoric conditions.

Across the independent claims, the coverage centers on preparing biomaterial samples in a high-pressure container operated below atmospheric-freezing temperature, where a nucleating-lock induces nucleation to form an ice layer acting as a natural piston to ensure isochoric conditions. The apparatus coverage further specifies an insulated low-temperature gradient chamber with a temperature-control holder including a heater to control container temperature and cool nucleating-locks to a nucleation temperature, with a plug-cavity nucleating agent and retainer-based insulating nucleating-lock structure.

Stated Advantages

Documented Applications