Published December 5, 2014 in BioPharma Reporter.

Chemists at IBBR — a joint institute of the National Institute of Standards and Technology (NIST) and the University of Maryland – found traces of the plant enzymes in batches of supposedly pure, commercially sold human blood protein that was genetically manufactured from plant seeds.

“Because they are active agents that promote biochemical reactions, enzyme contamination at even low levels could have an outsized effect on measurement reproducibility, and quality control in biomanufacturing,” NIST says. “And in a clinical context, if the enzymes end up in blood products given to patients, they might cause toxic or immune reactions.”

The contamination discovery came about almost randomly while IBBR chemists Robert Brinson, John Marino and colleagues were studying how different molecules bind to HSA, which can be used as a component of media for growing cell cultures to manufacture proteins.

The scientists discovered that the phytases eluded manufacturers’ quality control tests, such as enzyme-linked immunosorbent assay (ELISA), as well as liquid chromatography and mass spectrometry.  But when tested with NMR, the scientists noticed that bonds between phosphate groups and carbon atoms were breaking in one of the tested molecules, known as adenosine triphosphate.

The scientists realized that these phytase enzymes can “have an outsized effect” even at very low concentrations, Marino said, because, like chemical catalysts, they accelerate biochemical reactions without being consumed in the process.  “Because their background effect might raise or lower concentrations of key biomolecules, phytase contamination could cause researchers to misinterpret experimental results based on cell cultures grown in HSA-based medium,” Brinson and Marino said.

After their initial discovery, the team tested seven HSA products from four different vendors and found varying levels of phytase activity in each batch. They also tested human serum-derived and yeast-derived HSA products but did not detect carbon-phosphate bonds breaking, which seemed to be further proof that the contamination was from the plant enzymes.

Brinson and Marino said that NMR screening could provide an important quality control test for the increasing number of plant-produced biotech products.  Brinson also told BioPharma-Reporter.com that the cost of the NMR screening would not be prohibitively expensive. “For companies or other institutes with their own NMR, the cost is negligible. For companies with access to an NMR, the going industrial rate tends to be $100-$150” per hour, he said.

The researchers also emphasized that NMR should be used in addition to, not as a replacement for, traditional screening methods.