In the present work, a comprehensive methodology to carry out the screening for novel natural functional compounds is presented. To do that, a new strategy has been developed including the use of unexplored natural sources (i.e., algae and microalgae) together with environmentally clean extraction techniques and advanced analytical tools. The developed procedure allows also estimating the functional activities of the different extracts obtained and even more important, to correlate these activities with their particular chemical composition. By applying this methodology it has been possible to carry out the screening for bioactive compounds in the algae Himanthalia elongata and the microalgae Synechocystis sp. Both algae produced active extracts in terms of both antioxidant and antimicrobial activity. The obtained pressurized liquid extracts were chemically characterized by GC-MS and HPLC-DAD. Different fatty acids and volatile compounds with antimicrobial activity were identified, such as phytol, fucosterol, neophytadiene or palmitic, palmitoleic and oleic acids. Based on the results obtained, ethanol was selected as the most appropriate solvent to extract this kind of compounds from the natural sources studied.

HIGHLIGHTS  Review of pressurized hot water extraction of bioactive compounds, 2009-14  Chemical and physical properties of pressurized hot liquid water  Equipment, method optimization, applications, coupling and future prospects  Challenges with degradation and other chemical reactions during extraction ABSTRACT The purpose of this review is to give the reader a thorough background to the fundamentals and applications of pressurized hot water extraction (PHWE) for the analysis of bioactive compounds. We summarize the field in the period 2009-14, and include fundamentals of water as a solvent: equipment; method optimization; applications; coupling; and, future prospects. We highlight that solvent properties of water are tunable by changing the temperature, particularly self-ionization, dielectric constant, viscosity, diffusivity, density and surface tension. Furthermore, important aspects to consider are the risk of degradation of the analytes and other potential reactions, such as hydrolysis, caramelization and Maillard reactions that may lead to erroneous results. For the extraction of bioactive compounds, we report PHWE methods based on using water of 80-175°C and short extraction times. In conclusion, PHWE provides advantages over conventional extraction methods, such as being "greener", faster and more efficient. Why another review article on pressurized hot water extraction?Pressurized hot water extraction (PHWE) is an extraction technique that uses liquid water as extractant (extraction solvent) at temperatures above the atmospheric boiling point of water (100°C/273 K, 0.1 MPa), but below the critical point of water (374°C/647 K, 22.1 MPa) (Fig. 1). The use of PHWE in analytical chemistry started with the work in environmental analysis by Hawthorne and colleagues in the mid-1990s [1,2], and can also be referred to as subcritical water extraction (SWE), superheated water extraction and pressurized liquid extraction or accelerated solvent extraction with water as a solvent. There are a few relatively recent review articles on analytical PHWE, which the reader is recommended to read [3][4][5][6][7].The aim of this review article is to give a thorough background on the fundamental properties of water -an aspect that has been virtually overlooked in most review articles written so far about analytical PHWE. Hence, the first part of this review article concerns the fundamentals of chemical/physical properties of water and how these change with the increase in temperature, as well as how these affect the extraction performance both positively and negatively in different analytical applications.The second part deals with technical solutions of PHWE and how to conduct the experiment in practice. This technical part includes discussions on using commercially available and home-built equipment. The third part includes aspects on method optimization in PHWE. The fourth part summarizes some of the key applications and related publications mainly in the field of extraction of bioactive compounds from plants, food, ...

Nowadays, a wide variety of compounds such as polyphenols, polyunsaturated fatty acids (PUFA), or phytosterols obtained, for example, from wine, fish byproducts, or plants are employed to prepare new functional foods. However, unexplored natural sources of bioactive ingredients are gaining much attention since they can lead to the discovery of new compounds or bioactivities. Microalgae have been proposed as an interesting, almost unlimited, natural source in the search for novel natural functional ingredients, and several works have shown the possibility to find bioactive compounds in these organisms. Some advantages can be associated with the study of microalgae such as their huge diversity, the possibility of being used as natural reactors at controlled conditions, and their ability to produce active secondary metabolites to defend themselves from adverse or extreme conditions. In this contribution, an exhaustive revision is presented involving the research for innovative functional food ingredients from microalgae. The most interesting results in this promising field are discussed including new species composition and bioactivity and new processing and extraction methods. Moreover, the future research trends are critically commented.

The well-known correlation between diet and health demonstrates the great possibilities of food to maintain or even improve our health. This fact has brought about a great interest for seeking new products that can contribute to improve our health and wellbeing. This type of foods able to promote our health has generically been defined as functional foods. Nowadays, one of the main areas of research in Food Science and Technology is the extraction and characterization of new natural ingredients with biological activity (e.g., antioxidant, antiviral, antihypertensive, etc) that can contribute to consumer's well-being as part of new functional foods. The present work shows the results of a bibliographic revision done on the chemical composition of different macroalgae together with a critical discussion about their potential as natural sources of new functional ingredients.

In this contribution, pressurized liquid extraction (PLE) has been employed to isolate bioactive compounds from three native Romanian plants, oregano (Origanum vulgare), tarragon (Artemisia dracunculus) and wild thyme (Thymus serpyllum). Different PLE conditions have been tested including extraction with water, ethanol and their mixtures in a wide range of extraction temperatures (50-200°C), and the antioxidant capacity of the extracts was measured using different assays (DPPH radical scavenging, TEAC assay and Folin-Ciocalteau assay to measure total phenols). Moreover, a complete chemical characterization by using LC-MS/MS was carried out to be able to correlate the bioactivity with the particular chemical composition of each extract and plant. The use of PLE with water as a solvent at the highest temperature tested (200°C) always provided the highest extraction yields for the three studied plants, being maximum for oregano (>60%). Besides, oregano's pressurized water extracts at lower temperatures (50°C) presented the highest content on total phenols (184.9 mg gallic acid/g extract) and the best antioxidant activities (EC(50) 6.98 μg/ml). In general, oregano extracts were the most active, followed by wild thyme extracts. The antioxidant capacity measured by DPPH assay was highly correlated with the amount of total phenols. Moreover, the use of a LC-MS/MS method allowed the identification of 30 different phenolic compounds in the different extracts, including phenolic acids, flavones, flavanones and flavonols, which have an important influence on the total antioxidant capacity of the different extracts.