Cryogenic Storage and Shipping

Below, we cover the essentials of cryogenics with particular reference to healthcare and the pharmaceutical industry. Please continue reading for a definition of cryogenics and the reasons why it is crucial in pharma and modern medicine. You will also find an introduction to portable cryogenic containers used for storage and transportation, as well as an introduction to larger freezer units.

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Definition

Information published by the US National Institute of Standards and Technology refers to cryogenics as temperatures of minus 150ºC (-238ºF) or colder. However, in active pharmaceutical ingredients (API) production, temperatures of approximately -80ºC are sufficiently low for most current good manufacturing practice (cGMP) requirements.

Why Are Pharmaceuticals and Samples Deep Frozen?

Cryogenic conditions can limit impurities and facilitate the processing of highly reactive compounds. Usefully, the extreme cold tends to improve reaction selectivity and either eliminate or reduce unwanted side reactions. Other benefits include the limitation of compound volatility, thus improving overall safety until the completion of a reaction. Last but not least, rapid chilling means that ice crystals do not form.

Cryopreservation for biobanking purposes involves preserving tissue or cells in a vitrified state. To achieve this aim, technicians freeze the sample rapidly to bypass the hexagonal (crystal) phase of ice formation. Instead, the freezing process advances directly to the amorphous ice phase, making storage viable over the long term.

As the temperature is so low, all the water content of the cells is either removed by osmosis or vitrified. Thus, metabolic processes cease.

There are various societal implications. Some border on science fiction, whereas others form a part of today's treatments for childless couples. Notably, bio-banks have brought about a change in traditional social boundaries. In her 2018 University of Zurich paper entitled Suitable substances: how bio-banks (re)store biologicals, Sandra Bärnreuther asserts that biobank fertility treatments and IVF techniques have led to situations that differ from natural reproduction*.

On a commercial scale, cryogenic manufacturing processes require equipment and plants that are considerably more substantial than typical laboratory arrangements. To list but a few, heat transfer fluids, jacketed vessels, sophisticated pumps and complex control systems are necessary.

As C. Challener points out in Increasing API Complexity Drives Demand for Cryogenic Capabilities (Pharmaceutical Technology 42 (8), 2018), cryogenics requires specialized analytical tools. For instance, the liquid in standard mercury or alcohol thermometers would freeze, so platinum resistance thermometers and probe sensors are the norms instead.

Cryo Applications

Find out who is using cryogenic containers in the industry.

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Cryo Applications

Cryogenics in the Pharmaceutical Industry

As outlined above, the use of very low temperatures can influence reaction pathways. This point is especially significant in those reactions where desired and undesired products differ only slightly from an energetic standpoint, whether in stereo-, regio-, or chemoselectivity.

Similarly, reactions involving unstable intermediates, notably organometallic reagents, are not feasible at or near room temperature. Conversely, such reactions are often possible at much lower temperatures. In addition, processes involving gaseous reagents are sometimes easier to carry out in sub-zero conditions.

Thus, for all the above reasons, the pharmaceutical sector is developing new cryogenic production methods. The aim is to boost efficiency in the manufacture of existing products.

Next, let us consider cooling methods and storage in cryogenics.

Dry Ice

Relatively inexpensive and globally available, dry ice comprises solidified carbon dioxide. It is suitable for cooling substances and samples down to temperatures of -80°C.

Although personal protective equipment, including gloves, is necessary to handle dry ice safely, it is relatively easy to manage. In industrial processes, it can cool down large volumes of products, such as an entire pallet.

Dry ice is the right choice whenever a temperature of around -80°C is appropriate, as is typical in API or for preserving and transporting human tissue or other samples.

Conveniently, it is acceptable inside aircraft cargo holds. Other advantages include its relatively low cost and non-toxicity.

Characteristically, dry ice comes in large or small blocks or small cylindrical pellets. Standard blocks weigh around 30 kilograms. Wrapped in taped paper, these larger blocks are standard in shipping applications. Crucially, their high density and low surface-area-to-volume ratio mean that sublimation (the equivalent of evaporation) tends to be slow. In contrast, pellets of approximately a centimeter diameter are suitable for small scale use, such as inside insulated laboratory chests.

Looking for a Dry Ice Temperature Monitoring Solution?

ELPRO offers a range of temperature monitoring and data logger solutions for dry ice. All three types are IATA compliant and come with free calibration certificates.

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Looking for a Dry Ice Temperature Monitoring Solution?

Liquid Nitrogen

Costlier than dry-ice and more dangerous to handle, liquid nitrogen is your only choice if you need to keep a sample, substance or intermediate product in the glass phase (i.e., colder than -150°C). Liquid nitrogen boils at -195.8°C and freezes at -210°C.

However, this popular coolant boils immediately in contact with warmer (or less cold) surfaces, thus limiting its efficiency. Furthermore, as this sub-zero boiling occurs, the chilled object acquires an insulating envelope of nitrogen bubbles: the Leidenfrost effect. To minimize this effect, you plunge objects into a slush of solid and liquid nitrogen instead of LN2 alone to achieve faster cooling.

At these ultra-low temperature levels, dedicated cryo compatible containers are necessary. They have a vacuum chamber and a relatively small payload, sometimes of only a few cubic centimeters.

LN2 or liquid nitrogen's ability to maintain extremely cold temperatures means that this refrigerant is useful for the:

  • Storage of cells for laboratory work.

  • Cryopreservation of blood, reproductive cells, other biological material and samples.

  • Preservation of tissue from surgical excisions for medical studies.

  • Cryoconservation of animal genetic resources. Apart from R&D and life science uses, cryogenically preserved specimens can revive breeds that are endangered or extinct.

  • Very low temperature reactions and the controlled evaporation typical of advanced processes.

  • Cooling baths and vacuum pump traps typical of chemistry and laboratory research.

Cryogenic Containers

Also known as cryo-cans, the holding time of vacuum flasks varies depending on their design and size. Their construction reduces heat transfer and vaporization Typically, their inner lining comprises borosilicate or stainless steel. For durability, the outer linings are usually carbon steel or stainless steel.

Research dewars typically have a capacity of between 0.5 and 6 liters. In contrast, the MVE Lab Series of dewars have internal volumes of between 4 and 50 liters.

Storage and Transport Dewars

Today's portable cryogenic containers feature spill-free designs. By following the recommended usage instructions and correct filling technique, the outer shell of these dewars remains dry. Consequently, they attract a non-hazardous classification, thus simplifying courier handling and reducing transport costs. Moreover, the method favors the maintenance of sample viability.

Crucially, the filling process must be slow. Depending on the type of container, two to four cycles may be needed as the absorbent layer soaks up the liquid nitrogen. However, for obvious safety reasons, you should avoid overfilling.

While older dewars might have maintained temperatures low enough for a few hours, today's modern equivalents can perform well for weeks. Over time, though, shippers tend to accumulate moisture which decreases their performance.

Inside a typical liquid nitrogen dewar container, the dry nitrogen vapor chills the contents to -150°C or colder. Internal capacity is usually sufficient for approximately twenty-two 10 milliliter vials or seventy-five smaller 2 milliliter vials.

The maximum holding time for most portable dewars is around 10-12 days. Handling and transportation are safer, thanks to the latest protective shipping container mechanisms that prevent tilting and keep the shipper upright.

Freezers

The latest pressurized super-insulated vacuum vessels enable the storage of samples in liquid nitrogen for longer. Significantly, losses are as low as 2 percent per day. However, for larger volumes and long-term storage, liquid nitrogen freezers are more appropriate.

Cryogenic equipment manufacturers produce a choice of easy access wide neck freezers and high-efficiency versions designed for low LN2 consumption. The less voluminous models feature castors for occasional changes of position. Usually, these stainless steel units measure more than a meter in height.

Wiring and plumbing are usually fully enclosed, conferring safety and aesthetic advantages. Some of these cylindrical freezers have a standard single access step, while some taller variants have double-tier folding steps. On top, a robust metal handle operates the seal.

The latest product catalogs list cryogenic containers and freezers with vapor phase or liquid phase chilling options. Apart from accurate measurement, some advanced features include touchscreen controls, password security, user-defined alarms, and remote monitoring. In addition, their firmware permits communication with multiple devices and network connections. With ample data storage, the inbuilt memory can log thousands of events.

How to ship products using Cryogenic Containers

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How to ship products using Cryogenic Containers

Monitoring a Cryo-Container Comes With Challenges

In recent years, the development of cell and gene therapy (CGT), also called vein-to-vein treatments, has called for closed-loop supply chain and transport requirements. Time is often of the essence in this specialized arena, thus presenting unique challenges for CGT logistics. Specifically, excessive temperature variations or delivery delays could lead to clinical risk and financial loss.

Specifying Requirements

More recently, the Coronavirus pandemic and the intense initiative to research and produce effective vaccines has underlined the need for adequate monitoring of cryogenic containers and their contents. You may already be familiar with user requirements specifications (URS) as part of consignment documentation and control.

You use a URS to identify devices, list their contents, and specify other parameters such as the maximum acceptable deviation from recommended temperature. The document also details where to register any such temperature excursions.

Notably, a URS describes what the equipment involved does and the criteria for safe storage and transportation. Descriptions of how equipment works are not necessary.

For clarity, you should not list multiple requirements in a single section. Instead, to simplify checking and validating each condition, number the items and list them individually. As staff carry out checks, it is usual to record details such as names, locations, dates, times, and weight of the container, both when empty and full.

Data Loggers and Sensors

To monitor the level of chilling inside the container, you need an external sensor with a probe. Therefore, when choosing a logger, it is essential to check how the sensor fits into the interior chamber of the cryogenic container. Ideally, your data logger will also have an internal sensor to record ambient conditions, i.e., inside the logger's casing.

Other features to look for include customizable conditions and regular monitoring of the container contents during loading, preparing for shipment, and shipping. Tilt detection is valuable to prevent losses and safety incidents caused by mishaps with the container(s) under monitoring. If your company or organization operates a rental fleet, you will need the logger's battery to last for at least one year.

The LIBERO CD PDF data logger has an internal sensor and Bluetooth® connectivity but is suitable only for temperatures down to -95ºC. To work with temperatures as low as -200ºC, the CE model and real-time web-browser compatible LIBERO GE has an external sensor and an option to upgrade to ISO17025 standards.

ISO/IEC 17025 is a popular benchmark that lists the general requirements for testing and calibration. It enables laboratories to demonstrate competence and confirm their test results and measurements, thereby promoting confidence. The standard also facilitates cooperation between laboratories and national or international bodies. Thus, adherence to its terms is advantageous in commerce.

Why is Monitoring a Cryogenic Container Difficult?

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Why is Monitoring a Cryogenic Container Difficult?

Qualification of Cryogenic Containers

Typically, pharmaceutical products, CGT vials, and vaccine consignments ship from a laboratory or manufacturing plant to their ultimate destination, usually a hospital, clinic, or pharmacy. As pharma companies entrust their cargo to others, there is little or no room for error in each stage of the journey through storage depots, trucks, aircraft, or ships.

During the winter of 2020 and until recently, COVID-19 vaccine development and transportation featured in the news headlines. Nonetheless, the ongoing need to safeguard CGTs and tissue-engineered products is testing the industry's cold chain and logistical arrangements.

As one would expect, the transport of pharmaceutical products is subject to strict regulations. Therefore, simple or fleet qualification of each shipping lane or route is mandatory. In the final sections below, you will find the key recommendations and regulations to follow.

US Regulations

In the US, cryogenic material transport is subject to Department of Transportation (DOT) regulations that govern motor vehicle movement of hazardous materials. Nonetheless, these regulations have certain exceptions, one of which is Materials of Trade (MOT).

The MOT exception applies if employees receive awareness training and follow set requirements. Specifically, cryogenic materials, including liquid nitrogen and helium, fall under DOT hazard class 2.2. To transport them under the MOT exemption, containers should not weigh more than 220 pounds (100 kilograms) or be pressurized higher than 25.3 PSI (pounds per square inch), or approximately 1.7 bar. Also, closures should be secure in design and held firmly in place.

Outer containers are to be secure inside vehicles and not capable of shifting during the journey. Notably, the maximum load weight is 440 pounds (200 kilograms) under the MOT exception.

Single LN2 dewars of less than one-liter capacity are exempt from DOT regulations. However, they should have double glass wall insulation and robust outer packaging, such as boxes with internal cushioning and clear labeling.

European Regulations

In 2013, the EC published guidelines on good practice in the distribution of medicinal products for human use. Containers should bear labels showing the contents and their source, along with handling requirements and safety precautions.

Products subject to particular conditions or medicinal products comprising highly active materials require dedicated containers and secure vehicles for safety. Additionally, the relevant safety precautions and security measures must comply with international agreements and national legislation.

The transport of temperature-sensitive products requires qualified equipment such as thermal packaging and temperature-controlled containers. Similarly, vehicles should have adequate air conditioning or the refrigeration necessary to ensure the right ambient conditions during the movement of consignments between the manufacturer, wholesale distributor, and customer.

Regular maintenance and calibration are essential for the monitoring equipment installed in transport vehicle fleets. Moreover, the recommendations state that temperature mapping should be carried out under representative conditions and take seasonal variations into account.

Fleet Qualification of Cryogenic Containers

Why is it recommended to perform a fleet qualification on your cryogenic containers?

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Fleet Qualification of Cryogenic Containers

Finally, customers are entitled to request information to demonstrate that pharmaceuticals and medical samples have been subject to the required physical conditions during transit, as is the case for other perishable goods. Such assurance forms part of their quality checks and guarantees compliance with regulations.