Novel combination of feed enzymes to improve the degradation of Chlorella vulgaris recalcitrant cell wall

Research Areas: Science & TechnologyABSTRACT - In this study, a rational combination of 200 pre-selected Carbohydrate-Active enzymes (CAZymes) and sulfatases were tested, individually or combined, according to their ability to degrade Chlorella vulgaris cell wall to access its valuable nutritional compounds. The disruption of microalgae cell walls by a four enzyme mixture (Mix) in comparison with the control, enabled to release up to 1.21g/L of reducing sugars (p<0.001), led to an eight-fold increase in oligosaccharides release (p<0.001), and reduced the fuorescence intensity by 47% after staining with Calcofuor White (p<0.001). The Mix treatment was successful in releasing proteins (p<0.001), some MUFA (p<0.05), and the benefcial 18:3n-3 fatty acid (p0.05), total carotenoids were increased in the supernatant (p<0.05) from the Mix treatment, relative to the control. Taken together, these results indicate that this four-enzyme Mix displays an efective capacity to degrade C. vulgaris cell wall. Thus, these enzymes may constitute a good approach to improve the bioavailability of C. vulgaris nutrients for monogastric diets, in particular, and to facilitate the cost-efective use of microalgae by the feed industry, in general.info:eu-repo/semantics/publishedVersio

Sequential multi-stage extraction of biocompounds from Spirulina platensis: Combined effect of ohmic heating and enzymatic treatment

A sequential multi-stage procedure was applied on the extraction of biocompounds from Spirulina platensis. The process consisted in three steps: 1) aqueous extraction, using conventional thermal extraction (CE), ohmic heating (OH, 7V/cm), enzymatic treatment (EAE, 0.8 mgLysozyme/mL), or both OH and EAE combined; 2) ethanolic extraction; 3) CHCl3/MeOH extraction. The results evidenced that the combined OH-EAE extraction allowed selective recovery of phycobiliproteins in the 1st step, with increments of more than 100% in yield in comparison with CE. Pigments and lipids were selectively extracted in the 2nd step. The combination of OH and EAE in the 1st step resulted in higher amounts of extracted compounds in the following phases compared to processes using non-combined technologies. Results demonstrate that the intensification of extraction steps facilitates the use of environmentally friendly technologies in a multi-stage process capable of recovering and isolating different fractions with bio-functional properties, targeting waste reduction and circular economy. Industrial relevance Spirulina plantensis represents a potential biomass feedstock due to its potential as a source of compounds of great economic value (including antioxidants, proteins, lipids and natural pigments, in particular blue colorants). The combined use of ohmic heating and enzymes in the aqueous extraction step fosters the use of environmentally friendly technologies to implement sequential high yield and high purity extraction of the different valuable fractions with bio-functional properties, targeting waste reduction and contributing to the implementation of circular economy strategies. This can be integrated with a design of Industry 4.0 driving the development of new products.This research was funded by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UIDB/04469/2020 unit, by program Marie Skłodowska-Curie grant (MSCA-RISE; FODIAC; 778388) and by project OH2O - POCI-01-0145-FEDER-029145 (funded by FCT, COMPETE2020 – Competitiveness and Internationalization Operational Program and European Fund for Regional Development - FEDER). Pedro Santos is recipient of a PhD fellowship supported by a doctoral advanced training (call NORTE-69-2015-15), funded by the European Social Fund under the scope of Norte2020 - Programa Operacional Regional do Norte (NORTE-08-5369-FSE-000036). Sílvia Miranda acknowledges the financial support provided by FCT through the Doctoral grant SFRH/BD/144188/2019. Spirulina platensis was kindly supplied by EVRA S.r.l. (Potenza, Italy).info:eu-repo/semantics/publishedVersio

Impact of dietary incorporation of Spirulina (Arthrospira platensis) and exogenous enzymes on broiler performance, carcass traits and meat quality

This study assessed the effect of Spirulina (Arthrospira platensis), individually and in combination with exogenous enzymes, on growth performance, carcass traits, and meat quality of broiler chickens. One hundred and twenty Ross 308 male chickens were allocated into 40 battery brooders, with 3 birds per cage, and fed ad libitum a corn-based diet during the first 21 D of the trial. The experimental period lasted from day 21 to 35, during which birds were fed 4 different diets: a corn-soybean basal diet, taken as the control group, a basal diet containing 15% Spirulina (MA), a basal diet containing 15% Spirulina plus 0.005% Rovabio Excel AP (MAR), and a basal diet containing 15% Spirulina plus 0.01% lysozyme (MAL). Body weight gain (P , 0.001) and feed conversion rate (P , 0.001) were improved in control chickens, when compared with those fed with Spirulina. In addition, Spirulina increased the length of duodenum plus jejunum in relation to the other treatment (P , 0.01). Chickens on the MAL diet showed a considerable increase in digesta viscosity (P , 0.05) compared with the control group. Breast and thigh meats from chickens fed with Spirulina, with or without the addition of exogenous enzymes, had higher values of yellowness (b*) (P , 0.001), total carotenoids (P , 0.001), and saturated fatty acids (P , 0.001), whereas n-3 polyunsaturated fatty acid (P , 0.01) and a-tocopherol (P , 0.001) decreased, when compared with the control. In conclusion, the incorporation of 15% Spirulina in broiler diets, individually or combined with exogenous enzymes, reduced birds’ performance through a higher digesta viscosity, which is likely associated with the gelation of microalga indigestible proteins. In addition, cell wall of Spirulina was successfully broken by the addition of lysozyme, but not by Rovabio Excel AP. Therefore, we anticipate that the combination of lysozyme with an exogenous specific peptidase could improve the digestibility of proteins from this microalga and avoid their detrimental gelationinfo:eu-repo/semantics/publishedVersio

Identification of Synbiotics Conducive to Probiotics Adherence to Intestinal Mucosa Using an In Vitro Caco-2 and HT29-MTX Cell Model

The ability of bacteria to adhere to the intestinal mucosa is a critical property necessary for the long-term colonization of the intestinal tract. This ability can be highly sensitive to the presence of prebiotics. However, limited data are available in this respect for beneficial bacteria such as probiotics or resident gut microbiota. We previously demonstrated that the presence of prebiotics may decrease adherence in several pre- and probiotic combinations. Thus, characterizing the interactions between numerous combinations involving different classes of pre- and probiotics can be crucial in identifying new synbiotics. Accordingly, here, we extend our prior analyses to evaluate the adhesion of five lactobacilli, six bifidobacteria, and one probiotic Escherichia coli strains, as commercial probiotics or promising probiotic candidates, together with the cariogenic Bifidobacterium dentium strain. As an in vitro intestinal mucosa model, Caco-2 and mucin-secreting HT29-MTX cells were co-cultured at 9:1 in the presence or absence of prebiotics. Commercial inulin-type fructooligosaccharide prebiotics Orafti® GR, Orafti® P95, and galactooligosaccharide-based prebiotic formula Vivinal®, including purified human milk oligosaccharides (HMOs) were added into the cultivation media as the sole sugar source (2.5% each). Adherence was tested using microtiter plates and was evaluated as the percentage of fluorescently labeled bacteria present in the wells after three washes. Consistent prebiotics-mediated enhanced adherence was observed only for the commercial probiotic strain E. coli O83. For the remaining strains, the presence of HMO or prebiotics Orafti® P95 or Orafti® GR decreased adherence, reaching statistical significance (p &lt; 0.05) for three of out of eight (HMO) or five of out of 11 strains tested, respectively. Conversely, Vivinal® enhanced adhesion in six out of the 12 strains tested, and notably, it significantly attenuated the adherence of the cariogenic Bifidobacterium dentium Culture Collection of Dairy Microorganisms (CCDM) 318. To our knowledge, this represents the first report on the influence of commercial prebiotics and HMOs on the adhesion of the cariogenic Bifidobacterium sp. Vivinal® seems to be a promising prebiotic to be used in the formulation of synbiotics, supporting the adhesion of a wide range of probiotics, especially the strains B. bifidum BBV and BBM and the probiotic Escherichia coli O83

Curve fitting in Fourier transform near infrared spectroscopy used for the analysis of bacterial cells

International audienceInfrared spectroscopy is a prominent molecular technique for bacterial analysis. Within its context, near infrared spectroscopy in particular brings benefits over other vibrational approaches; these advantages include, for example, lower sensitivity to water, high penetration depth and low cost. However, near infrared spectroscopy is not popular within microbiology, because the spectra of organic samples are difficult to interpret. We propose a comparison of spectral curve-fitting methods, namely, techniques that facilitate the interpretation of most peaks, simplify the spectra and improve the prediction of bacterial species from the relevant near infrared spectra. The performances of three common curve-fitting algorithms and the technique based on the differential evolution were compared via a synthesized experimental spectrum. Utilizing the obtained results, the spectra of three different bacterial species were curve fit by optimized algorithm. The proposed algorithm decomposed the spectra to specific absorption peaks, whose parameters were estimated via the differential evolution approach initialized through Levenberg-Marquardt optimization; subsequently, the spectra were classified with conventional procedures and using the parameters of the revealed peaks. On a limited data set, the correct classification rate computed by partial least squares discriminant analysis was 95%. When we employed the peak parameters for the classification, the rate corresponded to 91.7%. According to the Gaussian formula, the parameters comprise the spectral peak position, amplitude and width. The most important peaks for bacterial discrimination were identified by analysis of variance and interpreted as N-H stretching bonds in proteins, cis bonds and CH2 absorption in fatty acids. We examined some aspects of the behaviour of standard curve-fitting algorithms and proposed differential evolution to optimize the fitting process. Based on the correct use of these algorithms, the near infrared spectra of bacteria can be interpreted and the full potential of near infrared spectroscopy in microbiology exploited