2.2. Generation of Different Kinds of Nanobubbles

Nanobubble water was produced by a ultra-micro bubble generator (XZCP-K-1.1) (100 nm to 10 μm) provided by Xiazhichun Co. Ltd. (Kunming, China). When the generator was turned on, gas from a gas cylinder was pumped into the machine in a negative pressure, and inlet and outlet pipes were immersed in deionized water/electrolyte solutions in the 10 L glass container where there was a circulation between the inlet and outlet, as shown in Figure 1A. The machine mixed the gas and liquid, and then high-density, uniform and “milky” nanobubble water (Figure 1B) was produced through a nozzle by the hydrodynamic cavitation method. The cloudy and milky nanobubble water gradually became clear through the process of the microbubbles rising and collapsing at the air–water interface (Figure 1C). This process took 2 to 3 min (Figure 1D). The machine worked for 15 min to generate high number density nanobubble water of more than 10⁸ bubbles/mL (obtained from NanoSight, NS300, Malvern (Worcestershire, UK) outsourcing).

Procedure of nanobubble generation. (A) Before generation; (B) During generation of micro-nano bubbles; (C) Stop the generation of bubbles; (D) After standing for 2 to 3 min.

After the above-mentioned preparation steps, the nanobubble water had a temperature of about 313 K. It was then left to cool down to room temperature. Next, from the prepared nanobubble water, the large sizes of the bubbles, from 1 to 10 μm, were removed by centrifugal treatment in the following manner. The nanobubble water was stored in a 50 mL centrifuge tube; then, centrifugal treatment was performed in 6000 rpm, i.e., 31.4 m/s peripheral velocity, for 6 min to remove possible impurities and large bubbles. After centrifugation, the size distribution, zeta potential, Eh and pH were measured from one centrifuge tube to record the first day’s data while other sealed centrifuge tubes with nanobubble water were kept at room temperature (298 K) for continuous measurements.