# Rapid Green Extractions of C-Phycocyanin from _Arthrospira maxima_ for Functional Applications
## 3 Results and Discussion
### Optimization of HPH Extraction Conditions for C-PC of Arthospira Maxima
Data are shown as mean \(\pm\) SD, n = 3.  
cellular rupture of _Chorella vulgaris_ also were capable of inducing the complete disruption of the cells when they worked with higher pressures (to 1500 bar)  
The supernatants obtained from HPH treated samples were characterized by a strong green color being the most intensive in all extraction processes similar to the results expose by [PERSON], [PERSON], [PERSON], [PERSON], [PERSON], [PERSON] and [PERSON] [34]. The major advantage in HPH is that this process can be scaled up easily with large volumes of biomass and with a wide range of disruption of cells (algae, bacteria and yeast). This technique is also non-selective with the product's recovery and with temperature increasing but it could be improved with the solvent selection and with the optimization of HPH passes, respectively [34; 37; 38]. In particular, chlorophyll extraction from _Spirulina platensis_ using the HPH process was described by [PERSON] and [PERSON] [39]. Unlike our results, its study greatly increased the extraction yield of chlorophyll at less pressure than our data (improved at 690 bar and decreased at 1370 bar). Finally, the good extraction yield obtained in HPH allowed a relevant comparison of the processes' performances with other environmentally friendly techniques such as ultrasound, supercritical fluids, pressurized liquid extraction etc. Comparison of C-PC Content of Arthospira Maxima between Conventional and Green Extraction Techniques  
Two conventional methods were also carried out to extract C-PC from _A. maxima._, Maceration (M) and freeze-thawing (FT) extraction, these being the conventional methods that were compared with green extraction techniques. Figure 3 shows C-PC content with the conventional extraction method using three independent solvents previously described. In this case, M extraction was the worst method to extract C-PC from _A. maxima_ (\(\sim\)151.8 mg/g as average with any solvent). However, the FT method was better and the results were similar to that of MW extraction (Na-buffer phosphate \(\sim\) 232.8 mg/g). According to the literature, the FT method is commonly used to extract phycocyanin from wet biomass, due to the fact that it improves the extraction efficiency [40; 41]. Although [PERSON], et al. [42] reported that the extraction of phycocyanin with water did not differ much from that phosphate buffer, we found relevant significances in all methods except the M process. It could be because the buffer used in that study was 10 mM sodium acetate buffer (pH 5.0) different from our buffer used. The isoelectric point is essential to proteins interacting with the solvent, so pH is an important factor to improve the extraction. \begin{table}
\begin{tabular}{l c c} \hline \hline  & \multicolumn{2}{c}{**C-PC Content (mg/g)**} \\ \hline
**Terms of the Model** & **Estimated** & \(p\)**-Value** \\ \hline constant & 211.26 & \\ A Pressure & 10.3651 & 0.0001 * \\ B Solvent & 10.9577 & 0.0000 * \\ AA Pressure \(\times\) Pressure & \(-\)29.2046 & 0.0000 * \\ AB Pressure \(\times\) solvent & 8.37285 & 0.0061 * \\ BB Solvent \(\times\) solvent & 47.2129 & 0.0000 * \\ Statistics for goodness of fit of the model & & \\ R2 & 0.881058 & \\ RSD & 11.1752 & \\ P & 0.4979 & \\ RRSD (\%) & \(-\)7.51706 & \\ \hline \hline \end{tabular} R2-determination coefficient, RSD-residual standard deviation, \(p\)-value of the lack-of-fit test for the model, RRSD-residual standard deviation expressed as a percentage of the mean value of the response; * significant coefficients of the model.  
\end{table} 
Table 4: Regression coefficients (values of variables are specified in their original units) and statistics for the fit obtained by multiple linear regression. [PERSON], et al. [43] has described in their research four different methods for the extraction of pycobiliproteins from _Arthrospira maxima_, recording that the highest content of phycocyanin recovery was 78.5 mg/L (equivalent to 8.7%, respect to dry biomass) under the phosphate regulator method with dry ice.