## 4 Discussion  
The HPLC peak intensity and areas clearly showed that the high-pressure homogenization process at 690 bar yielded substantial amounts of intact chlorophyll \(a\), which was identified by its retention time compared to a standard, while the ethanol extract had very small peak areas. The concentration of chlorophyll \(a\) achieved by HP was very high compared to the results from other works, which ranged from 1.2 mg/g to 9.3 mg/g chlorophyll \(a\) under various extraction conditions [4; 7; 25] and, in particular, was much higher than the 5.27 mg/g achieved by conventional 70% EE as a control. These results suggest that there must be an optimal operating pressure for obtaining the maximal amount of chlorophyll \(a\) from high-pressure homogenization, since some of the chlorophyll \(a\) within _Spirulina_ could be destroyed at the extremely high shear stress of approximately 50,000 (1/h) generated by 1370 bar of input pressure. This potential degradation of chlorophylls explains why the chlorophyll \(a\) content of HP-3 was lower than that of HP-2 but was higher than that of EE or HP-1. This result is also supported by other research showing that the cell walls of _Spirulina_ are not as rigid as those of plants [26]. In addition to this result, only a small yield of chlorophyll \(a\) was achieved at a low pressure of 350 bar (HP-1), possibly because the cell walls of _Spirulina_ were not completely broken at this low shear stress and consequently did not allow large amounts of chlorophyll to diffuse into the solvent in the relatively short extraction time. Therefore, a simple high-pressure homogenization process can greatly increase the extraction yield of chlorophyll \(a\) by efficiently disrupting the structure of _Spirulina_ under an optimal input pressure. When processed at 350 bar, many of the cells remained intact, which explains why the yield of chlorophyll \(a\) was lower at that pressure than at 690 bar. Therefore, the results shown in these figures confirmed the results shown in Table 1, which indicated that there must be an optimal input pressure when employing high-pressure homogenization to obtain chlorophyll \(a\) from _Spirulina_. In this work, 690 bar emerged as a preliminary optimal pressure, which easily released chlorophyll \(a\) from the biomass within a very short time even at room temperature, when the intact chlorophyll \(a\) content and neuroprotective effects of extracts were compared across three different levels of input pressures such as 350, 690, and 1370 bar. However, more exact optimal conditions should be determined with greater accuracy by optimizing many effective parameters of high-pressure homogenization in processing _Spirulina_ sp. Other works observed similar effects of high-pressure homogenization, finding that it easily broke down the structures of the biomass [16; 17]. These results were very interesting and constituted a starting point for applying this unique process to extract other components from natural resources. Our results concerning antioxidant activity strongly indicated that the HP extract had higher antioxidant activity than chlorophyll \(a\) alone even at the same dosage, possibly due to additive effects of other components in the HP extract. By contrast, the EE extract showed lower antioxidant effects than 10 mg/mL of chlorophyll \(a\) alone because the EE yield was not high enough for there to be 10 mg/mL of chlorophyll \(a\) in the extract and there were not enough other bioactive components. These results demonstrated that the extract from a simple high-pressure homogenization had higher antioxidant activity than the EE extract or even chlorophyll \(a\) alone, which meant that the HP extract was expected to show strong neuroprotective effects as well. The neuroprotection results also strongly indicated that chlorophyll \(a\) must play an important role in protecting cell growth against damage by glutamate, since the HP extract contained higher amounts of chlorophyll \(a\) than the ethanol extract did.