# Recent Advances and Insights into Bromelain Processing, Pharmacokinetics and Therapeutic Uses
## 3 Extraction and Purification Methods
### Comparison of Bromelain Extraction Techniques  
The direct comparison of the published data on bromelain extraction techniques is a difficult task because several types of bromelain extracts and test conditions were used. Purification level and activity are the two main parameters used to assess the differences between extraction techniques (Table 3). The aqueous two-phase extraction technique showed the highest purification factor even if it involved a strong reduction in enzymatic activity [62]. Moreover, despite this strategy being considered a rapid and scalable technique, one of the main disadvantages of this process is the high salt concentration and the critical recovery that hamper recycling. Precipitation has a high activity recovery, second only to ultrafiltration, in addition to being a low-cost technique and, for this reason, the most used for commercial bromelain extracts. Nevertheless, the purification fold with this method is generally low and it presents the same problem as the aqueous two-phase extraction technique of high precipitant content (waste environmental impact) [20]. Ion exchange chromatography is the most expensive technique for several reasons: the low recovery and separation efficiency, and the small sample loading capacity [35]. However, it can produce satisfactory results especially when combined with other techniques such as gel filtration. In this regard, one of the challenges for the coming years is the study of new  
\begin{table}
\begin{tabular}{c c c c c} \hline \hline  & \begin{tabular}{c} **Ctivity** \\ **Recovery (\%)** \\ \end{tabular} & \begin{tabular}{c} **Purification** \\ **Fold/Factor** \\ \end{tabular} & **Reference** \\ \hline \multirow{4}{*}{\
\begin{tabular}{c} Aqueous two-phase \\ extraction \\ \end{tabular} } & PEG/K\({}_{2}\)SO\({}_{4}\) & 228 & 4.0 & [27] \\ \cline{2-5}  & PEG/polyacrylic acid & 335.27 & 25.78 & [55] \\ \cline{2-5}  & PEG/(NH\({}_{4}\))\({}_{2}\)SO\({}_{4}\) & - & 11.80 & [38] \\ \cline{2-5}  & PEG/MgSO\({}_{4}\) & 113.54–206 & 2.23–62 & [10; 39; 56; 57] \\ \cline{2-5}  & Block copolymers & 79.5 & 1.25 & [36] \\ \hline \multirow{4}{*}{Filtration} & Microfiltration and ultrafiltration & 85–100 & 10 & [32] \\ \cline{2-5}  & Microfiltration, ammonium sulphate & 50 & - & [30] \\ \cline{2-5}  & precipitation, ultracentrifugation & 64.75 & 5.3 & [31] \\ \cline{2-5}  & Nano-TiO\({}_{2}\), ultrafiltration & 64.75 & 5.3 & [31] \\ \hline \multirow{2}{*}{
\begin{tabular}{c} Reverse micellar \\ systems \\ \end{tabular} } & Reverse micelle systems & 85–97.56 & 4–5.2 & [45; 46; 48; 58] \\ \cline{2-5}  & Affinity-based reverse micelle system & 185.6 & 12.32 & [59] \\ \hline \multirow{4}{*}{Chromatopraphy} & High speed counter-current & - & - & [45] \\ \cline{2-5}  & chromatography, reverse micelle system & 94.6 & 15.4 & [60] \\ \cline{1-1} \cline{2-5}  & Precipitation, ion exchange chromatography & - & 3.3 & [35] \\ \cline{1-1} \cline{2-5}  & Cation exchange chromatography & - & 10 & [61] \\ \hline \hline \end{tabular}
\end{table}
Table 2: Unconventional techniques for the extraction and purification of bromelain (adapted from [PERSON] et al., 2016 [54]).  
hybrid extraction techniques to optimize costs, reduce environmental impact and increase extract yield and purity. Limited information is available for the use of affinity membranes, despite the fact that the potentially high selectivity could represent an important advantage for bromelain extraction [63]. Lastly, the ultrafiltration technique seems to be the most promising for bromelain large-scale extraction operation because it is easily scalable to high product throughput and environmentally friendly [32]. To date, the most promising and economically viable results have been obtained by combining different processes such as the reverse micellar systems with ultrafiltration [33] or microfiltration prior to ultrafiltration [64].