How Shock Waves Revolutionize Biotechnology and Pharmaceuticals

At UNAM's Center for Applied Physics and Advanced Technology (CFATA), a group of scientists, advised by Miriam Rocío Estévez González, was able to extract phytochemicals with antioxidant, antibacterial, and anticancer properties from endemic plants using shock waves in a faster, more efficient, and environmentally friendly way than with conventional methods because no solvents were used.

Similarly, the Center's Shock Wave Laboratory in Juriquilla, Querétaro, changed the genes of fungi and "shaved" their spores. This is important to the pharmaceutical industry because spores are used to make enzymes, antibiotics, molecules for vaccines, and anticoagulants, among other things, according to the head of the lab, Achim Max Loske Mehling.

With this change to filamentous fungi, "we can increase the production of the enzymes they secrete," and their genetic transformation with shock waves, which can also affect the food and textile industries, got a national patent.

A shock wave, the scientist explained, is generated when enough energy is deposited in a small space. It is a sudden "rise" and "fall" of pressure, up to 1,500 times the atmospheric pressure at sea level, and in an extremely short time (which can be millionths of a second).

In nature, for example, they are generated by lightning during a thunderstorm. They have been used for decades, for example, in medicine to pulverize kidney stones and in orthopedics, cardiology, traumatology, and dermatology. However, we did not expect that they could be useful in pharmaceuticals and biotechnology, as the expert pointed out.

The equipment used for these applications is a bathtub with water and a shock wave generator, which we can imagine as a kind of audio speaker that has a series of piezoelectric crystals mounted on it; the waves are concentrated in a region called the focus. "There we place a small container called a vial, which contains what we want to treat."

In the case of filamentous fungi, which are microscopic, genetic material (DNA) or other macromolecules of interest are inserted to make them more efficient at secreting substances that are important for the pharmaceutical field; "we force" them to produce the compounds we want or to generate a much greater quantity of those they create.

The phenomenon through which this genetic transformation occurs is called acoustic cavitation. The suspension containing the fungi and the genetic material to be introduced is placed inside the vial, and a shock wave is passed through it, compressing the microbubbles present in the liquid. When they collapse, they emit microjets or high-velocity fluid sparks that perforate the fungus and function as "microsyringes" that allow the DNA to enter. Tens or hundreds of shock waves are usually applied.

Achim Loske said that Francisco Fernández Escobar experiments with tandem shock waves in the lab. These waves help boost the flow of these tiny jets of fluid because they are double: we send a second wave just as the microbubble is bursting, which gives it a lot more power.

By studying the processes of genetic transformation and trying to improve them by changing the parameters of the shock wave generator (so that the microjets pass through the fungus but don't kill it), the university student, Blanca Edith Millán Chiu, and their team found a new method: shaving the three-micron-long conidia.

These, he said, are a type of fungal spore; the cells that allow them to remain dormant in adverse situations, such as drought or a lack of nutrients until suitable conditions for development exist. "We wanted to see what happened to the membrane and outer wall of the conidia, which is relatively resistant."

In this way, they discovered spore "shaving." The microjets produced by the aforementioned collapse of the bubbles work like microsyringes, and because they travel at extremely fast speeds, 700 to 900 meters per second (equivalent to a bullet), they generate turbulence, shear, and intense stresses over the vicinity. "It is as if a projectile were passing close to an object, but without hitting it, only by glancing at it, and even if it does not touch it, it generates effects."

The turbulence and stresses generated "shave" the so-called ornamentations, which are protective structures present on the surface, as Achim Loske described. This procedure could replace more laborious processes used in pharmaceuticals.

What's important here is that there are ways to get them, such as destroying the conidia with ultrasound: "They are pulverized, and then, through laborious procedures, certain molecules of the ornamentation are extracted."

Thanks to the effects of microjets, they can be obtained relatively quickly and easily. This recent parallel result, we think, has great potential and a bright future because the equipment we use can be scaled up to industrial dimensions. The finding was published in the International Journal of Fungi, through the article Effect of Shock Waves on the Growth of Aspergillus niger Conidia: Evaluation of Germination and Preliminary Study on Gene Expression, on October 24.

In another novel application of acoustic cavitation, scientists discovered that it also works for the extraction of compounds. In this context, the expert said that algae and native plants are good sources of important phytochemicals like flavonoids and fucoxanthins.

So far, they are obtained through a variety of simple methods, such as maceration, which means "squeezing" the important parts out of plants like palo azul (Eysenhardtia polystachya), which has important properties like compounds that fight cancer.

We have discovered that if we grind the bark of this tree into fine particles, make a suspension, place it in a vial, and pass shock waves through it, we can extract phytochemicals without solvents, without heating, or using ultrasound; in other words, in a faster, more efficient, and environmentally friendly way.

The research team also tested marine algae, such as sargassum and cuachalalate, because they are high in phytochemicals, with the same encouraging results.

With this discovery, we hope to replace conventional techniques. "By adjusting the time between the tandem waves and, with it, the diameter and size of the microjets, we can have some control over the extraction process of the compounds; it can even be selective, which makes it more interesting."

Although this research is still basic science, it holds promise for future applications. The industry may be interested in optimizing its processes and replacing conventional systems with cheaper and simpler ones, says Achim Loske.

Miguel Angel Martinez Maldonado, a postdoctoral researcher at CFATA, and Miguel Angel Gómez Lim, an academic at CINVESTAV, Irapuato, as well as academics and students from UNAM's postgraduate programs in materials science and engineering and the Autonomous University of Querétaro faculty of chemistry, all play important roles in the projects mentioned above.