applied sciences Review Biohydrogen from Microalgae: Production and Applications Antonina Rita Limongi 1,2 , Emanuele Viviano 1,3,*, Maria De Luca 1,4, Rosa Paola Radice 1,2 , Giuliana Bianco 1 and Giuseppe Martelli 1 1 Department of Science, University of Basilicata, 10, 85100 Potenza, Italy; [email protected] (A.R.L.); [email protected] (M.D.L.); [email protected] (R.P.R.); [email protected] (G.B.); [email protected] (G.M.) 2 Bioinnova s.r.l.s., Via Ponte 9 luci, 22, 85100 Potenza, Italy 3 Thema Informatik s.r.l., Via Ressel 2/F, 39100 Bolzano, Italy 4 Almacabio s.r.l., Via Ressel 2/F, 39100 Bolzano, Italy * Correspondence: [email protected] Abstract: The need to safeguard our planet by reducing carbon dioxide emissions has led to a significant development of research in the field of alternative energy sources. Hydrogen has proved to be the most promising molecule, as a fuel, due to its low environmental impact. Even if various methods already exist for producing hydrogen, most of them are not sustainable. Thus, research focuses on the biological sector, studying microalgae, and other microorganisms’ ability to produce this precious molecule in a natural way. In this review, we provide a description of the biochemical and molecular processes for the production of biohydrogen and give a general overview of one of the most interesting technologies in which hydrogen finds application for electricity production: fuel cells. Keywords: green fuel; biohydrogen; microalgae; Chlamydomonas reinhardtii; hydrogenase; fuel cell Citation: Limongi, A.R.; Viviano, E.; De Luca, M.; Radice, R.P.; Bianco, G.; Martelli, G. Biohydrogen from 1. Introduction Microalgae: Production and Nowadays, searching for renewable energy represents one of the major challenges Applications. Appl. Sci. 2021, 11, for the global scientific community. Population growth and industrial activities, with con- 1616. https://doi.org/10.3390/ sequential high-energy demand, require solutions and proposals in the short-term. It is app11041616 assumed that the world population could reach 8–10 billion in 2040, while the increase in global energy demand, within the same year, is estimated between 100 and 170 qBtu. Academic Editor: Dino Musmarra However, models cannot accurately predict sudden changes in the global energy situa- Received: 29 December 2020 tion, as different socio-economic scenarios and political choices modulate each country’s Accepted: 7 February 2021 Published: 10 February 2021 response [1,2]. Considering the slow oil, coal, natural gas formation processes, the exces- sive exploitation of fossil fuels has triggered a drastic and irreversible reserve reduction. Publisher’s Note: MDPI stays neutral Another side aspect of fossil fuel use concerns greenhouse gas (GHG) emissions, particu- with regard to jurisdictional claims in larly carbon dioxide (CO2), during the combustion reaction [3]. In particular, in the years published maps and institutional affil- between 1998 and 2018, CO2 emissions increased by 48%, making it necessary to implement iations. carbon capture and storage (CCS) technology to limit serious and detrimental effects on climate change [2]. In this worrying scenario, fossil fuels still provide more than 80% of primary energy needs, although the interest in renewable sources, together with their consumption, are steadily increasing [4]. Among renewable options, hydrogen shows the important advantage of CO -free com- Copyright: © 2021 by the authors. 2 Licensee MDPI, Basel, Switzerland. bustion, with water as a by-product. Thermodynamic properties, compared to traditional This article is an open access article fuels, confirm the interest in its investments in research, although several aspects related distributed under the terms and to production and storage are still to be mastered. Today, several industrial applications conditions of the Creative Commons depend on hydrogen, but the most is obtained by techniques, such as steam reforming or Attribution (CC BY) license (https:// electrolysis, not entirely free from the involvement of fossil fuels [5]. Some living organisms, creativecommons.org/licenses/by/ such as microalgae and bacteria, are the basis of processes capable of producing hydrogen 4.0/). in a completely eco-sustainable way. Microalgal hydrogen production is made possible Appl. Sci. 2021, 11, 1616. https://doi.org/10.3390/app11041616 https://www.mdpi.com/journal/applsci Appl. Sci. 2021, 11, x FOR PEER REVIEW 2 of 14 Appl. Sci. 2021, 11, 1616 2 of 14 capable of producing hydrogen in a completely eco-sustainable way. Microalgal hydrogen production is made possible by biological processes directly or indirectly, dependingby biological on processessunlight, directlyor by fermentation or indirectly, processes depending and on thermochemical sunlight, or by fermentationtechnologies forprocesses biomass and conversion thermochemical (Figure technologies1). for biomass conversion (Figure1). FigureFigure 1. 1. HydrogenHydrogen production production pr processesocesses in in microalgae microalgae.. In recent years, many green systems have focused on algal biomass to obtain energy In recent years, many green systems have focused on algal biomass to obtain energy from living matter. Microalgae store and exploit light energy thanks to the photosynthetic from living matter. Microalgae store and exploit light energy thanks to the photosynthetic process. To support cell growth and metabolic activities, they use resources widely avail- process. To support cell growth and metabolic activities, they use resources widely able in nature, such as water and CO2. These organisms can concentrate considerable available in nature, such as water and CO2. These organisms can concentrate considerable CO2 amounts and obtain nutrients necessary for growth, even from substrates or waters CO2 amounts and obtain nutrients necessary for growth, even from substrates or waters deriving from industrial waste. Algal cultivation systems, which are simple and relatively derivinginexpensive, from ensureindustrial advantageous waste. Algal growth cultivation rates. systems, Furthermore, which are they simple can be and set relatively up on in- inexpensive,fertile territories ensure without advantageous competing growth with agricultural rates. Furthermore, areas that can they be can exploited be set for up food on infertileresources. territories In particular, without the competing general interest with ag inricultural microalgae areas has that increased can be significantly exploited for in foodrecent resources. decades dueIn particular, to the variety the general and versatility interest ofin themicroalgae metabolites has increased present in significantly various and innumerous recent decades species due [6]. to the variety and versatility of the metabolites present in various and numerousThe present species review [6]. article is a collection of the most recent evidence on hydrogen productionThe present in green review microalgae article is and a collection the efforts of tothe understand most recent and evidence improve on thehydrogen above- productionmentioned processesin green microalgae in the most and widely the usedefforts species. to understand and improve the above- mentioned processes in the most widely used species. 2. Hydrogen Metabolism in Green Microalgae: Biophotolysis 2. HydrogenHydrogen Metabolism metabolism in wasGreen firstly Microalgae: observed Biophotolysis in eukaryotic microalgae in 1939 by Ger- manHydrogen physiologist metabolism Hans Gaffron was [firstly7]. Due observed to the oxygenin eukaryotic incompatibility, microalgae it occursin 1939 only by Germantransiently physiologist in photosynthetic Hans Gaffron organisms. [7]. Due to the oxygen incompatibility, it occurs only transiently in photosynthetic organisms. 2.1. Photosynthetic Electrons and Hydrogenase 2.1. PhotosyntheticThe pivotal processElectrons of and microalgal Hydrogenase metabolism consists of oxygenic photosynthesis andThe complex pivotal redox process reactions of microalgal that take placemetabolis at them levelconsists of theof oxygenic thylakoid photosynthesis membranes in andchloroplasts complex throughredox reactions two successive that take phases. place at During the level the of first the light-depending thylakoid membranes reactions, in chloroplastsATP and reduced through NADH, two successive and NADPH, phases. are During generated the tofirst be light-depending involved in the nextreactions, dark ATPreactions and reduced where the NADH, atmospheric and NADPH, CO2 is fixedare generated by a RuBiSco to be (ribulose-1,5- involved in the bisphosphate next dark carboxylase/oxygenase) enzyme to ultimately generate energy rich-carbohydrate stores. reactions where the atmospheric CO2 is fixed by a RuBiSco (ribulose-1,5- bisphosphate carboxylase/oxygenase)Specifically, during the lightenzyme phase, to ultimately an electron generate transport energy chain rich-carbohydrate is generated along stores. with Specifically,photosystems during II (PSII) the via light the phase, plastoquinone an electron (PQ) transport pool, cytochrome chain is generated b6f complex along (Cyt with b6f), and photosystems I (PSI) due to the light energy received as photosystems are associated photosystems II (PSII) via the plastoquinone (PQ) pool, cytochrome b6f complex (Cyt b6f), with light-harvesting complexes I and II (LHC I and LHCII), consisting of numerous and photosystems I (PSI) due to the light energy received as photosystems are associated photoreceptive