Fermentation at non-conventional conditions in food- and bio-sciences by the application of advanced processing technologies.

The interest in improving the yield and productivity values of relevant microbial fermentations is an increasingly important issue for the scientific community. Therefore, several strategies have been tested for the stimulation of microbial growth and manipulation of their metabolic behavior. One promising approach involves the performance of fermentative processes during non-conventional conditions, which includes high pressure (HP), electric fields (EF) and ultrasound (US). These advanced technologies are usually applied for microbial inactivation in the context of food processing. However, the approach described in this study focuses on the use of these technologies at sub-lethal levels, since the aim is microbial growth and fermentation under these stress conditions. During these sub-lethal conditions, microbial strains develop specific genetic, physiologic and metabolic stress responses, possibly leading to fermentation products and processes with novel characteristics. In some cases, these modifications can represent considerable improvements, such as increased yields, productivities and fermentation rates, lower accumulation of by-products and/or production of different compounds. Although several studies report the successful application of these technologies during the fermentation processes, information on this subject is still scarce and poorly understood. For that reason, the present review paper intends to assemble and discuss the main findings reported in the literature to date, and aims to stimulate interest and encourage further developments in this field.

Application of High Pressure with Homogenization, Temperature, Carbon Dioxide, and Cold Plasma for the Inactivation of Bacterial Spores: A Review.

Formation of highly resistant spores is a concern for the safety of low-acid foods as they are a perfect vehicle for food spoilage and/or human infection. For spore inactivation, the strategy usually applied in the food industry is the intensification of traditional preservation methods to sterilization levels, which is often accompanied by decreases of nutritional and sensory properties. In order to overcome these unwanted side effects in food products, novel and emerging sterilization technologies are being developed, such as pressure-assisted thermal sterilization, high-pressure carbon dioxide, high-pressure homogenization, and cold plasma. In this review, the application of these emergent technologies is discussed, in order to understand the effects on bacterial spores and their inactivation and thus ensure food safety of low-acid foods. In general, the application of these novel technologies for inactivating spores is showing promising results. However, it is important to note that each technique has specific features that can be more suitable for a particular type of product. Thus, the most appropriate sterilization method for each product (and target microorganisms) should be assessed and carefully selected.

Metabolic stability and metabolite profiling of emerging synthetic cathinones.

Synthetic cathinones constitute the second largest groups of new psychoactive substances (NPS), which are especially popular among adolescents and young adults. Due to their potential toxicity, the recreational use of these NPS constitute a serious worldwide public health problem. However, their fast appearance in the market renders the continuous updating of NPS information highly challenging for forensic authorities. The unavailability of pharmacokinetic data for emerging NPS is critical for forensic and clinical verifications. With the ultimate goal of having a proactive approach towards the NPS issue, high resolution mass spectrometry was used in the current work to assess preliminary pharmacokinetic data for 8 selected cathinones: 4 reported substances (4-CIC, 3-CMC, 4-CMC and 4-MEAP) and 4 previously unreported ones (3-CIC, 4-MDMB, 4-MNEB and 4-MDMP) for which the emergence on the NSP market is expected to be eminent, were also included in this study. Based on the calculation of pharmacokinetic parameters, half-life and intrinsic clearance, 4-CMC and 4-MDMB are low and high clearance compounds, respectively, and all the remaining cathinones included in this study are intermediate clearance compounds. This fact anticipates the key role of metabolites as suitable biomarkers to extend detection windows beyond those provided by the parent cathinones. Reduction of the keto group and hydroxylation on the alkyl chains were the common metabolic pathways identified for all cathinones. However, the relative importance of these metabolic transformations is dependent on the cathinone substituents. The glucuronic acid conjugation to metabolites stemming for keto group reduction constituted the sole Phase II transformation identified. To our knowledge, this study constitutes the first metabolite profiling of the already reported synthetic cathinones 4-CIC, 3-CMC and 4-CMC. Noteworthy is the fact that 3-CMC accounts for almost a quarter of the quantity of powders seized during 2020. The analytical methods developed, and the metabolites characterized, are now available to be included in routine screening methods to attest the consumption of the 8 cathinones studied.

The Combined Effect of Pressure and Temperature on Kefir Production-A Case Study of Food Fermentation in Unconventional Conditions.

Food fermentation under pressure has been studied in recent years as a way to produce foods with novel properties. The purpose of this work was to study kefir production under pressure (7-50 MPa) at different temperatures (17-32 °C), as a case study of unconventional food fermentation. The fermentation time to produce kefir was similar at all temperatures (17, 25, and 32 °C) up to 15 MPa, compared to atmospheric pressure. At 50 MPa, the fermentation rate was slower, but the difference was reduced as temperature increased. During fermentation, lactic and acetic acid concentration increased while citric acid decreased. The positive activation volumes (Va) obtained indicate that pressure decreased the fermentation rate, while the temperature rise led to the attenuation of the pressure effect (lower Va). On the other hand, higher activation energies (Ea) were observed with pressure increase, indicating that fermentation became more sensitive to temperature. The condition that resulted in a faster fermentation, higher titratable acidity, and higher concentration of lactic acid was 15 MPa/32 °C. As the authors are aware, this is the second work in the literature to study the combined effect of pressure and temperature on a fermentative process.

Combined effect of pressure and temperature for yogurt production.

Fermentation under non-conventional conditions has gained prominence in the last years, due to the possible process improvements. Fermentation under sub-lethal pressures is one of such cases, and may bring novel characteristics and features to fermentative processes and products. In this work, the effect of both pressure (10-100 MPa) and temperature (25-50 °C) on yogurt production fermentation kinetics was studied, as a case-study. Product formation and substrate consumption were evaluated over fermentation time and the profiles were highly dependent on the fermentation conditions used. For instance, the increase of pressure slowed down yogurt fermentation, but fermentative profiles similar to atmospheric pressure (0.1 MPa) were obtained at 10 MPa at almost all temperatures tested. Regarding temperature, higher fermentative rates were achieved at 43 °C for all pressures tested. Moreover, the inhibitory effect of pressure increased when temperature decreased, with complete inhibition of fermentation occurring at 50 MPa for 25-35 °C, contrasting to 43 °C where inhibition occurred only at 100 MPa. Therefore, an antagonistic effect seems to occur, since yogurt fermentation was slowed down by pressure increasing, on one hand, and by temperature decreasing, on the other hand. Additionally, some kinetic parameters were calculated and fermentation at 43 °C presented the best results for yogurt production, with lower fermentation times and higher lactic acid productivities. Interestingly, fermentation at 10 MPa/43 °C presented the optimal conditions, with improved yield and lactic acid production efficiency, when compared to fermentation at 0.1 MPa (efficiency of 75% at 10 MPa, against 40% at 0.1 MPa). As the authors are aware, this work gives the first insights about the simultaneous effect of pressure and temperature variation on a microbial fermentation process, which can be combined to modulate the metabolic activity of microorganisms during fermentation in order to improve the fermentative yields and productivities of the desired product.

Adaptation of Saccharomyces cerevisiae to high pressure (15, 25 and 35 MPa) to enhance the production of bioethanol.

Saccharomyces cerevisiae is a yeast of great importance in many industries and it has been frequently used to produce food products and beverages. More recently, other uses have also been described for this microorganism, such as the production of bioethanol, as a clean, renewable and sustainable alternative fuel. High pressure processing (HPP) is a technology that has attracted a lot of interest and is increasingly being used in the food industry as a non-thermal method of food processing. However, other applications of high pressure (HP) are being studied with this technology in different areas, for example, for fermentation processes, because microbial cells can resist to pressure sub-lethal levels, due to the development of different adaptation mechanisms. The present work intended to study the adaptation of S. cerevisiae to high pressure, using consecutive cycles of fermentation under pressure (at sub-lethal levels), in an attempt to enhance the production of bioethanol. In this context, three pressure levels (15, 25 and 35 MPa) were tested, with each of them showing different effects on S. cerevisiae fermentation behavior. After each cycle at 15 and 25 MPa, both cell growth and ethanol production showed a tendency to increase, suggesting the adaptation of S. cerevisiae to these pressure levels. In fact, at the end of the 4th cycle, the ethanol production was higher under pressure than at atmospheric pressure (0.1 MPa) (8.75 g.L-1 and 10.69 g.L-1 at 15 and 25 MPa, respectively, compared to 8.02 g.L-1 at atmospheric pressure). However, when the pressure was increased to 35 MPa, cell growth and bioethanol production decreased, with minimal production after the 4 consecutive fermentation cycles. In general, the results of this work suggest that consecutive cycles of fermentation under sub-lethal pressure conditions (15 and 25 MPa) can stimulate adaptation to pressure and improve the bioethanol production capacity by S. cerevisiae; hence, this technology can be used to increase rates, yields and productivities of alcoholic fermentation.

Effect of High Pressure on Paracoccus denitrificans Growth and Polyhydroxyalkanoates Production from Glycerol.

The performance of fermentation under non-conventional conditions, such as high pressure (HP), is a strategy currently tested for different fermentation processes. In the present work, the purpose was to apply HP (10-50 MPa) to fermentation by Paracoccus denitrificans, a microorganism able to produce polyhydroxyalkanoates (PHA) from glycerol. In general, cell growth and glycerol consumption were both reduced by HP application, more extensively at higher pressure levels, such as 35 or 50 MPa. PHA production and composition was highly dependent on the pressure applied. HP was found to decrease polymer titers, but increase the PHA content in cell dry mass (%), indicating higher ability to accumulate these polymers in the cells. In addition, some levels of HP affected PHA monomeric composition, with the polymer produced at 10 and 35 MPa showing considerable differences relative to the ones obtained at atmospheric pressure. Therefore, it is possible to foresee that the changes in polymer composition may also affect its physical and mechanical properties. Overall, the results of this study demonstrated that HP technology (at specific levels) can be applied to P. denitrificans fermentations without compromising the ability to produce PHA, with potentially interesting effects on polymer composition.

Ethnopharmacology, phytochemistry and biological activity of Erodium species: A review.

Erodium spp. is a genus that can be found in all continents that has been traditionally used in folk medicine to treat many diseases such as hemorrhage, dermatological disorders, indigestion, and inflammatory diseases. Moreover, Erodium leaves have been used for the preparation of salads, omelets, sandwiches, sauces and soups, among other food products. The objective of this review was to show the recent and relevant studies about extraction of bioactive compounds, the phytochemical characterization, the potential biological activities and toxicological evidence reported in both in vitro and in vivo studies from Erodium spp. In addition, the use of Erodium spp. as natural compounds against the development of diseases were also showed. This review highlights the traditional use of Erodium species in several countries as a therapeutic agent to treat several diseases (such as constipation, dermatological disorders, diabetes, indigestion, urinary inflammations, and as carminative agent), the factors influencing the extraction of bioactive compounds (mainly species and solvent composition on phenolic compounds) and phytochemical profile (presence of essential oils and alkaloids), the scientific evidence about its anti-inflammatory, antimicrobial (against both spoilage and pathogenic microorganisms), antiviral and other health-related activities (anti-protozoal and anti-viral activity) as well as the toxicological evidence. Erodium spp. is a relevant source of compounds with antioxidant, antimicrobial, and biological activity, which support its potential exploration in pharmacological and food area. Major efforts are necessary to advance the knowledge about Erodium genus regarding the relation between traditional use and scientific evidence, optimization of extraction conditions, the influence on biological mechanisms at animal and clinical levels, and bioaccessibility and bioavailability of bioactive compounds.

Lactobacillus reuteri growth and fermentation under high pressure towards the production of 1,3-propanediol.

Lactobacillus reuteri is a lactic acid bacterium able to produce several relevant bio-based compounds, including 1,3-propanediol (1,3-PDO), a compound used in food industry for a wide range of purposes. The performance of fermentations under high pressure (HP) is a novel strategy for stimulation of microbial growth and possible improvement of fermentation processes. Therefore, the present work intended to evaluate the effects of HP (10-35 MPa) on L. reuteri growth and glycerol/glucose co-fermentation, particularly on 1,3-PDO production. Two different types of samples were used: with or without acetate added in the culture medium. The production of 1,3-PDO was stimulated at 10 MPa, resulting in enhanced final titers, yields and productivities, compared to 0.1 MPa. The highest 1,3-PDO titer (4.21 g L-1) was obtained in the presence of acetate at 10 MPa, representing yield and productivity improvements of ≈ 11 and 12%, respectively, relatively to the same samples at 0.1 MPa. In the absence of acetate, 1,3-PDO titer and productivity were similar to 0.1 MPa, but the yield increased ≈ 26%. High pressure also affected the formation of by-products (lactate, acetate and ethanol) and, as a consequence, higher molar ratios 1,3-PDO:by-products were achieved at 10 MPa, regardless of the presence/absence of acetate. This indicates a metabolic shift, with modification of product selectivity towards production of 1,3-PDO. Overall, this work suggests that HP can be a useful tool to improve of 1,3-PDO production from glycerol by L. reuteri, even if proper process optimization and scale-up are still needed to allow its industrial application. It also opens the possibility of using this technology to stimulate other glycerol fermentations processes that are relevant for food science and biotechnology.

Hyperbaric storage of melon juice at and above room temperature and comparison with storage at atmospheric pressure and refrigeration.

Hyperbaric storage (8h) of melon juice (a highly perishable food) at 25, 30 and 37°C, under pressure at 25-150 MPa was compared with atmospheric pressure storage (0.1 MPa) at the same temperatures and under refrigeration (4°C). Comparatively to the refrigerated condition, hyperbaric storage at 50/75 MPa resulted in similar or lower microbial counts (total aerobic mesophiles, enterobacteriaceae, and yeasts/moulds) while at 100/150 MPa, the counts were lower for all the tested temperatures, indicating in the latter case, in addition to microbial growth inhibition, a microbial inactivation effect. At 25 MPa no microbial inhibition was observed. Physicochemical parameters of all samples stored under pressure (pH, titratable acidity, total soluble solids, browning degree and cloudiness) did not show a clear variation trend with pressure, being the results globally similar to refrigeration storage. These results show the potential of hyperbaric storage, at and above room temperature and with potential energy savings, comparatively to refrigeration.

Microorganisms under high pressure--adaptation, growth and biotechnological potential.

Hydrostatic pressure is a well-known physical parameter which is now considered an important variable of life, since organisms have the ability to adapt to pressure changes, by the development of resistance against this variable. In the past decades a huge interest in high hydrostatic pressure (HHP) technology is increasingly emerging among food and biosciences researchers. Microbial specific stress responses to HHP are currently being investigated, through the evaluation of pressure effects on biomolecules, cell structure, metabolic behavior, growth and viability. The knowledge development in this field allows a better comprehension of pressure resistance mechanisms acquired at sub-lethal pressures. In addition, new applications of HHP could arise from these studies, particularly in what concerns to biotechnology. For instance, the modulation of microbial metabolic pathways, as a response to different pressure conditions, may lead to the production of novel compounds with potential biotechnological and industrial applications. Considering pressure as an extreme life condition, this review intends to present the main findings so far reported in the scientific literature, focusing on microorganisms with the ability to withstand and to grow in high pressure conditions, whether they have innated or acquired resistance, and show the potential of the application of HHP technology for microbial biotechnology.