Integrity Testing of Flexible Containers - Defects as small as 10 μm can be detected without compromising product cleanliness using helium integrity testing. - BioPharm International


Integrity Testing of Flexible Containers
Defects as small as 10 μm can be detected without compromising product cleanliness using helium integrity testing.

BioPharm International
Volume 24, Issue 11, pp. 42-49


Microbiological challenge test

While immersion biotesting has long been used to challenge packages, particularly cans, for pinholes and channel leaks, they are not the real conditions that a package generally encounters during its use. Hence, package integrity-evaluation methods that employ bioaerosols that simulate the conditions that the package will be expected to tolerate during storage and distribution are more relevant and are gaining prominence. Test bags with defects between 2 and 50 μm were exposed to a microbial environment and served as test sample. The test bags not exposed to bacteria served as negative control for the aseptic filling. For each organism, a bag containing no defect was injected with 0.1 mL of a 103 CFU/mL solution of the organism to serve as positive control. The positive controls exhibited growth after 1 d, thus validating the test conditions for detecting microbial ingress.

Table I: Microbial challenge test data.
As expected, under the conditions of the test, microbial ingress into a package took longer as the defect size got smaller. Test bags with defect sizes of 50 and 20 μm took the same time (i.e., 5 d) to allow microbial growth, meaning that a clear channel for microbial ingress was already established at 20 μm. However, when the defect size was 15 μm, it took 14 d to show any microbial growth (see Table I). A significant slowdown in the microbial ingress at 15 μm and complete cessation of microbial ingress at 10 μm or smaller defect sizes are interesting, considering that microbial organisms are much smaller than 10 μm and should infiltrate through 10 μm defects just as easily as they did through 20 μm defects. The logical explanation for this observation lies in the threshold pressure inside the test bag (1). To initiate microbial ingress through a defect, the pressure inside the test bag (i.e., threshold pressure) must overcome the force of the liquid surface tension and initiate liquid flow through the defect, thus providing a channel for microbes to travel into the bag. The magnitude of threshold pressure required to initiate liquid flow depends on the location of the defect due to differences in the static head pressures. As defect size decreases, the threshold pressure for a given liquid increases. Thus, in test bags with 10 μm defects, the threshold pressure is lower than the force of the liquid surface tension, preventing the formation of a channel through which microbes can travel. One other reason offered in the literature for this behavior is the formation of a biofilm on the film surface, which prevents microbial ingress.

Although additional studies may be required to confirm the root cause for the lack of penetration of microbes through a 10 μm defect, these results are in agreement with researchers in the food packaging industry such as Lampi and Chen, who have shown the critical dimension for microbial ingress to be about 10 μm (2, 3). The differences in the critical defect size for microbial ingress between different studies could be due to differences in the test methods employed and the contact materials that affect the surface tension of the liquid. Based on these results, it is clear that defects larger than 10 μm cause sterility breach. Therefore, integrity testing for on-line package testing must detect 10 μm defects to ensure product sterility.

Leak detection data

In an ideal world, the helium in the test background would be nonexistent, and a good bag would not leak helium at all. On the other hand, a defective bag would leak a definite amount helium, that would be detected, thus resulting in distinct separation of helium leak rates for defective bags versus those for good bags. This ideal behavior would result in a high degree of resolution, allowing existing leak-detection methods to be used to detect bag defects.

However, the walls of flexible bags are often made of polymeric materials, which are intrinsically permeable to gases. Helium gas has a smaller molecular size and permeates faster through polymeric materials than air or nitrogen. Thus, even a good test bag can leak a significant amount of helium by diffusion through the bag walls. This diffused helium creates high helium background levels in the test chamber, thus making it difficult to quantify actual helium leaks through the defects in the test bag. The high helium background essentially masks the helium leaking from defects, limiting the lowest leak rate that can be reliably measured. HIT testing ensures that helium flowing through the defects is maximized, while the background helium concentration is minimized. The test time was kept as short as possible to prevent elevation in the helium background as the test progressed.

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