Bromelain is a mixture of proteolytic enzymes that is extracted primarily from pineapples. It is present in almost all the aerial parts of the plant—the peels, leaves, stems and fruit—although only the fruit and stems contain significant amounts. The market-demand for bromelain is quickly increasing, especially in the health sector as it can be used as a drug and/or nutraceutical. Although the complete molecular mechanism has not been fully identified, bromelain possesses several properties, including anti-inflammatory, anti-oedema, antithrombotic and fibrinolytic, mucolytic, anticancer and cicatrizing effects that have been evaluated in several clinical trials. However, one of the main limitations for the clinical use of this supplement is the varying composition of extracts, which leads to heterogeneity in results and, therefore, difficulty in making evidence-based prescriptions. In fact, different geographic locations, land and modes of cultivation, as well as extraction methods, can all give extracts with different activities, depending on the plant part used. Additionally, although novel extraction techniques have been developed to improve bromelain purification and extraction and give higher yields without loss in enzymatic activity, these methods are still expensive and challenging. This review will describe the state of the art in the main conventional and unconventional extraction and purification methods of bromelain and discuss the advantages and limitations of these strategies. Pharmacokinetic and pharmacodynamic profiles and the clinical applications that have arisen from randomized controlled clinical trials are also discussed. Finally, future perspectives for bromelain extracts will be presented.

Bromelain is a mixture of cysteine endopeptidase usually extracted from pineapple juice and is used for the treatment of various human diseases and in industrial applications. Bromelain demand is quickly increasing, and its recovery from pineapple residues appears to be a sustainable waste management strategy. Pineapple core is among the most significant waste materials in the production of canned pineapple and is richer in bromelain than other pineapple residues. In this project, we compared the enzymatic properties and composition of bromelain extracts from either pineapple core or pulp to address the recovery of bioactive bromelain from pineapple core, thus contributing to the valorization of this waste material. Although significant differences were detected in the protein content of the two preparations, no differences could be detected for their proteolytic activity and for the effect of pH on their enzymatic activity. Mass spectrometry (MS) approaches identified the same peptidases in the fruit and in the core. This confirmed the possibility of using pineapple core to obtain relevant amounts of bioactive bromelain by applying a relatively simple procedure, thus paving the way to implementing a circular economy in this specific industrial sector.

Bevacizumab (BVZ) is a monoclonal antibody that binds to human vascular endothelial growth factor A (VEGF-A) and inhibits the interaction between VEGF-A and VEGF receptors, thus blocking the angiogenesis. Repeated intravitreal injections of BVZ for the treatment of ocular pathologies that present an excessive proliferation results in a low patience compliance. BVZ is specially indicated for the treatment of diabetic and degenerative retinopathy. In the present study, we designed lipid nanoparticles (NPs) as a BVZ sustained drug delivery system for reducing the frequency of administration. We used a simple and highly efficient procedure, “Cold dilution of microemulsions”, to obtain spherical NPs with mean diameters of 280–430 nm, Zeta potentials between −17 and −31 mV, and drug entrapment efficiencies between 50 to 90%. This study focused on the biochemical and biophysical stabilities of BVZ after entrapment in NPs. SDS-PAGE electrophoretic analysis and circular dichroism, dynamic light scattering, and scanning electron microscopy were used to characterize BVZ-loaded NPs. The biocompatibility was assessed by in vitro cell compatibility studies using the ARPE-19 cell line. Thus, in this work, a stable BVZ-loaded system was obtained. In addition, several studies have shown that BVZ is released slowly from the lipid matrix and that this system is biocompatible. The results are promising and the developed NPs could be exploited to create a new, potentially effective and minimally invasive treatment of intraocular diseases.

α-amylases are ubiquitous enzymes belonging to the glycosyl hydrolase (GH13) family, whose members share a high degree of sequence identity, even between distant organisms. To understand the determinants of catalytic activity of α-amylases throughout evolution, and to investigate the use of homologous enzymes as a model for the human one, we compared human salivary α-amylase, Aspergillus oryzae α-amylase and pancreatic porcine α-amylase, using a combination of in vitro and in silico approaches. Enzyme sequences were aligned, and structures superposed, whereas kinetics were spectroscopically studied by using commercial synthetic substrates. These three enzymes show strikingly different activities, specifically mediated by different ions, despite relevant structural homology. Our study confirms that the function of α-amylases throughout evolution has considerably diverged, although key structural determinants, such as the catalytic triad and the calcium-binding pocket, have been retained. These functional differences need to be carefully considered when α-amylases, from different organisms, are used as a model for the human enzymes. In this frame, particular focus is needed for the setup of proper experimental conditions.

The high-fat content of hazelnuts, mainly triglycerides, makes them prone to lipid oxidation during storage, which has a big impact on their sensory and nutritional quality. The chemical pathways leading to hazelnut oxidative rancidity have been well characterized and are faster on free fatty acids. Lipase(s) enzymes are required in oilseeds to hydrolyse the ester bonds to free the single molecule of fatty acids. This step, necessary for germination, is the first event that triggers rancidity. Identifying the lipase(s) in hazelnuts and the biochemical pathways involved in rancidity would lead to an effective strategy to prevent fat deterioration. Different proteins have been characterized in hazelnut seed and great interest has been risen towards the non-specific lipid transfer protein (nsLTP) family because they were identified as human allergens. Here we show that Cor a 8 - a member of nsLTPs - is a novel non-regiospecific lipase able to bind to oil-water interfaces and hydrolyse the triacylglycerol (TAG) ester bonds by a non-canonical active site (non-serine dependent). Molecular modelling and molecular dynamics suggest that Cor a 8 is not only able to catalyse the hydrolysis of TAGs but also to stabilize the resulting free fatty acids within the active site. Cor a 8 homologues are present in all land plants, but the specific catalytic amino acids are found only in angiosperms, suggesting an evolutionary adaptation for lipid metabolism unique to flowering plants. This study sets the foundation for understanding this new lipid metabolism in plants and its role in rancidity development.