Ion chromatography coupled with optical emission spectrometry (IC-ICP-OES) methodology for the analysis of inositol phosphates in food and feed.

This study presents the development of ion chromatography coupled with inductively coupled plasma optical emission spectrometry (IC-ICP-OES) for the simultaneous determination and quantification of inositol phosphates (InsPx). Using a CarboPac PA100 column with a nitric acid-water gradient, 28 InsPx isomers (InsP6 to InsP2) were separated within 33 min. The method eliminates baseline drift and post-column derivatization thereby simplifying detection and quantification. It achieves low detection limits of 63 µg/L P across a range of 63-3200 µg/L P. Various extraction and sample preparation methods for food and feed matrices were tested, including acidic and alkaline agents, C18 SPE and spin concentrators. The analysis shows intra-day and intra-laboratory reproducibility with deviations smaller than 1 % for standard solutions and under 4 % for feed samples (80 % recovery rate of phytate). This methodology is applicable to explore enzymatic degradation pathways and the analysis of InsPx in complex food and animal feed matrices.

Safety of a feed additive consisting of 3-phytase produced with Komagataella phaffii CECT 13171 (FSF10000/FLF1000) for poultry species, pigs for fattening and minor porcine species for fattening (Fertinagro Biotech S.L.).

Following a request from the European Commission, EFSA was asked to deliver a scientific opinion on the safety of the feed additive consisting of 3-phytase produced with a genetically modified strain of Komagataella phaffii (CECT 13171). The additive is intended to be used as a zootechnical additive for poultry species, pigs for fattening and minor porcine species for fattening. In a previous opinion, the EFSA Panel on Additives and Products or Substance used in Animal Feed (FEEDAP) Panel could not conclude on the taxonomic identification of the production strain. Moreover, the presence of viable cells of the production strain in the final formulations of the product could not be excluded. Therefore, no conclusions could be drawn on the safety for the target species, consumers, users and environment. In the present submission, the applicant provided supplementary information regarding the characterisation of the production strain and the absence of its viable cells in the final product. Based on the data provided, the identification of CECT 13171 as K. phaffii was confirmed and no viable cells of the production strain were detected in the final formulations of the additive. The FEEDAP Panel concluded that the 3-phytase produced with the genetically modified strain K. phaffii CECT 13171 does not give rise to any safety concern as regard to the production strain. Consequently, the additive, in both its liquid and solid formulations, is safe for the target species, consumers, users and environment.

Response of YPM x Ross 708 male broilers to diets containing varying inclusions of phytase, calcium butyrate, and bacitracin methylene disalicylate from 1 to 42 d of age-part 1: performance, processing yields, and nutrient digestibility.

This 42-d study evaluated the effects of phytase, calcium butyrate (CB), and bacitracin methylene disalicylate 50 (BMD) on broiler performance, processing yields, and nutrient digestibility. Ross YPM x 708 male broilers (2,880 total) were distributed in 72 floor pens and assigned to 1 of 9 treatments (8 replicates/treatment) on d of hatch. This experiment was a 2 × 4 + 1 factorial arrangement, including 2 phytase concentrations (500 or 1,500 FTU/kg), 4 microbiota modulating feed additive groups (MMFA; none, CB (0.5 g/kg of diet), BMD (55 mg/kg of diet), or both CB and BMD), and a negative control without feed additives. Broiler performance (d 14, 28, and 42), apparent ileal nutrient digestibility (d 28 and 42), and processing yields (d 43) were determined. Day 14 BW increased with BMD inclusion compared to CB and no MMFA in the 1,500 FTU/kg group but BW were similar between all MMFA combined with 500 FTU/kg (P ≤ 0.05). Supplementing BMD increased d 28 BW and reduced d 1 to 28 feed conversion ratio compared to CB and no MMFA (main effect, P ≤ 0.05). Day 42 BW varied depending on dietary phytase concentrations. When diets contained 500 FTU/kg, broilers fed both CB and BMD had a higher BW than broilers fed only CB. Whereas when the inclusion of phytase was increased to 1,500 FTU/kg, broilers fed diets with only BMD or both CB and BMD had higher BW than broilers fed diets with no MMFA (P ≤ 0.05). Phytase concentrations at 1,500 FTU/kg increased (P ≤ 0.05) digestibility of fat (main effect, d 42), phosphorus (d 28 and 42), and apparent ileal digestible energy (main effect, d 42) compared to 500 FTU/kg. In this study, dietary BMD improved broiler growth compared to CB and no MMFA. However, these observed differences between CB and BMD were dependent on dietary phytase concentrations.

A study of solely used phytase or in combination with multi-carbohydrase on growth performance along with tibia mineralization, and carcass traits in broilers fed nutrient-deficient diets.

The objective of this study aimed to investigate the effectiveness of phytase alone and in combination with multi-carbohydrase in available phosphorus (AP) and energy-deficient diets on growth performance, nutrient digestibility, tibia traits, and carcass quality of broilers. A total of 288 1-d-old Ross 308 broiler chicks were randomly assigned to one of 6 dietary treatments in a completely randomized design, with each treatment having 6 replicates and 8 birds per cage. The treatments were as follows: 1) positive control with adequate nutrition (PC); 2) 0.20% AP-deficient diet without phytase (NC-1); 3) 0.25% AP-deficient diet without phytase (NC-2); 4) NC-1 diet plus 500 FTU/kg phytase (NCP-1); 5) NC-2 diet plus 750 FTU/kg phytase (NCP-2); and 6) NC-2 with 100 kcal/kg ME deficient diet plus 1,000 FTU/kg phytase and multi-carbohydrase: 2,200 U/kg galactomannanase, 30,000 U/kg xylanase, 22,000 U/kg ß-glucanase, and 700 U/kg α-galactosidase (NCPM). Broilers in the PC group and those fed enzyme-supplemented diets exhibited greater (P < 0.05) growth performance in terms of body weight, average daily gain, and feed conversion ratio, along with enhanced tibia mineralization through the ash, calcium (Ca), phosphorus (P) composition, and tibia breaking strength compared to NC diets throughout the study. Among the treatments, broilers assigned to the NCPM group showed greater (P < 0.05) levels of energy and P digestibility on days 21 and 35. Concerning carcass characteristics, the leg meat yield was greater (P < 0.05) in broilers fed the NCP-2 diet compared to all other treatments on days 21 and 35. Supplementation of phytase at doses of 500 FTU/kg and 750 FTU/kg effectively recovers AP deficiencies of 0.20% and 0.25%, respectively, in broiler diets without compromising the growth performance of broilers. Additionally, combining multi-carbohydrase and phytase led to AP and energy loss recovery, which improved tibia mineralization and nutrient digestibility through their synergistic interaction.

Poultry encounter challenges in synthesizing endogenous enzymes to completely degrade antinutritional factors, including phytic acid (PA) and nonstarch polysaccharides (NSP). These exogenous enzymes are present in plant-based feed ingredients, which encapsulate nutrients and adversely affect their digestion in the gastrointestinal tract (GIT). Incorporating exogenous enzymes, including phytase and multi-carbohydrase, in broiler diets is a promising strategy for enhancing energy, protein, mineral digestibility, and feed efficiency. However, there remains inconsistency in studies presenting both challenges and opportunities for optimizing enzyme utilization. In the present study, different concentrations of phytase in available phosphorus (AP)-deficient diets, along with its combination with multi-carbohydrase (i.e., galactomannanase, xylanase, ß-glucanase, and α-galactosidase) in both AP and energy-deficient diets, were examined for their effects on the growth performance, nutrient digestibility, tibia traits, and carcass quality of broilers fed conventional corn-soybean meal diets. The findings indicate that supplementing nutrient-deficient diets with phytase alone or with multi-carbohydrase did not affect broiler growth, as their body weight, weight gain, feed intake, and feed conversion ratio were similar to those of broilers on a control diet. Those outcomes could be due to the enhancement of nutrient digestibility and bone mineralization, which suggests successful PA and NSP degradation in the GIT of broilers.

Effects of feed particle size, calcium concentration and phytase supplementation on InsP6 degradation in broiler chickens fed pelleted diets.

1. The objective of the trial was to study the single and interactive effects of feed particle size in pelleted feed, dietary calcium (Ca) concentration and microbial phytase supplementation in broiler chickens. The studied traits were myo-inositol (1,2,3,4,5,6) hexakis (dihydrogen phosphate) (InsP6) degradation, pre-caecal digestibility of phosphorus (P), Ca and amino acids (AA) and retention of P, Ca and nitrogen (N).2. Male Ross 308 broiler chickens were housed in metabolism units in groups of 10 and allocated to one of eight diets with seven pen replicates per diet. The 2 × 2 × 2-factorial arrangement included coarse and fine feed particle size (309 or 222 µm), low and high Ca concentration (4.9 and 7.2 g/kg) and without or with phytase supplementation (1,000 FTU/kg).3. Pre-caecal InsP6 disappearance was higher with coarse than fine feed particle size when no phytase was added (54 vs. 48%) but not when phytase was added (74%; p = 0.046). High dietary Ca feeds decreased pre-caecal InsP6 disappearance (67 to 59%) and P digestibility (65 to 55%; p < 0.001). Gizzard pH was lower with coarse than fine feed particle size and higher with high Ca than low Ca (p < 0.001). Pre-caecal digestibility of most AA was approximately 3.5%-points lower with high Ca without phytase compared to the other treatments (p ≤ 0.047). Coarse feed particle size caused higher pre-caecal AA digestibility than fine particle size (~2%-points; p ≤ 0.031). InsP6 disappearance in the crop increased at high Ca concentration when phytase was added (22 vs. 37%; p = 0.011).4. Coarser feed particle size in pellets increased gastrointestinal InsP6 degradation and nutrient digestibility, likely owing to effects on the gizzard functions. Additional Ca supply exerted antinutritive effects that was not compensated for by using coarser feed particles.

Effects of dietary calcium and microbial phytase on the bioavailability of two sources of zinc in broilers.

1. The objective of this study was to test the effect of microbial phytase and calcium (Ca) levels on the Zn bioavailability depending on the Zn source.2. A total of 144, one-day-old broilers received a diet containing 40 mg Zn/kg for 1 week. They were then assigned to one of the eight experimental treatments during 2 weeks. Diets contained 27 mg native Zn and were formulated according to a 2 × 2 × 2 factorial design with two added Zn sources (23 mg Zn/kg), sulphate (ZnSO4) or oxide (ZnO); two dietary Ca levels of either 6 (low) or 10 g/kg (moderate); and two microbial phytase levels, of either 0 or 750 phytase unit/kg.3. Moderate Ca plus phytase improved body weight gain (BWG) and low Ca level without microbial phytase decreased tibia weight (Ca × Phy; p = 0.049). Zinc oxide resulted in a higher tibia development (i.e. tibia weight and length; p < 0.05) and BWG (p = 0.009) and lower FCR (p < 0.001) compared to ZnSO4.4. Phytase improved tibia Zn concentration especially in birds given ZnSO4 compared to ZnO (Zn × Phy; p = 0.049). Moderate Ca diets improved tibial characteristics and Zn deposition compared to low Ca diets (p < 0.05).5. Soluble Zn in the gizzard increased in the presence of phytase (p = 0.011), while higher dietary Ca reduced this in birds receiving ZnO (p = 0.004). In the jejunum, Zn solubility was higher with phytase (p = 0.008).6. Under the conditions of this study, dietary levels of Ca and microbial phytase affect Zn availability in broilers more than the Zn source.

Calcium and phosphorus digestibility in broilers as affected by varying phytate concentrations from corn.

Dietary phytate P (PP) concentration impacts Ca and P digestibility in broilers. Research was conducted to determine the impact of increasing concentration of dietary PP, with and without phytase, on broiler standardized ileal digestibility (SID) of Ca and P. Digestible (Dig) Ca and P were calculated by multiplying SID and the analyzed dietary Ca and P concentrations. The experiment was a factorial arrangement of 2 phytase (0 and 1,000 U/kg) and 4 PP (0.16, 0.23, 0.29, and 0.34%) concentrations. Treatments were fed for 36 h from 20 to 22 d of age (4 b/pen, n ≥ 7 replicate pens/treatment). Different ratios of corn and corn germ were used to achieve the desired PP concentrations. A limestone with 800 µm geometric mean diameter was used as the sole Ca source to achieve 0.7% Ca in the final diets (96% Ca from limestone). An additional diet was fed that was N, Ca- and P-free, for the determination of endogenous losses of each nutrient. Distal ileal digesta were pooled from all birds in a pen. There were no interactions between PP and phytase on SID Ca or Dig Ca from limestone. Irrespective of phytase inclusion, increasing PP from 0.16 to 0.34% decreased SID Ca from 53.8 to 38.1% (P < 0.05). The SID Ca averaged 41.5 and 51.4% in diets containing 0 and 1000 U phytase/kg, respectively, across all PP concentrations (P < 0.05). Interactions were seen between PP and phytase on SID and Dig P (P < 0.05) with SID P of 31.1, 24.0, 20.1, and 16.3% for broilers fed 0.16, 0.23, 0.29, and 0.34% PP diets without phytase, respectively. When phytase was included at 1000 U/kg, SID P was 89.9, 87.5, 73.9 and 60.4% for diets containing 0.16, 0.23, 0.29 and 0.34% PP, respectively (P < 0.05). Overall, phytase improved SID Ca and P independent of PP concentration. However, with increasing PP concentration, both SID Ca and P were negatively affected.

Phytase, 25-hydroxyvitamin D3 and cocci vaccination to broilers fed a calcium and phosphorus-reduced diet under Eimeria spp. challenge: effects on growth performance and intestinal health.

A study evaluated the effects of phytase, 25-hydroxyvitamin D3 (25OHD), and cocci vaccination on broilers fed a diet reduced in calcium (Ca) and available phosphorus (avP) under Eimeria challenge. A total of 840 one-day-old male broilers were assigned to a 2 × 5 factorial arrangement based on cocci vaccination and dietary treatments. Half of the birds were vaccinated against coccidia on d 1, and all birds were orally challenged with Eimeria spp. (sporulated oocysts: 12,500 of E. maxima, 12,500 of E. tenella, and 62,500 of E. acervulina) on d 14. Dietary treatments included: 1) a nutrient adequate diet (PC); 2) a diet reduced by 0.2% in Ca and avP (NC); 3) NC plus 1,500 FTU/kg of phytase (NC+PHY); 4) NC plus 3,000 IU/kg of 25OHD (NC+25OHD); 5) NC with both PHY and 25OHD (NC+PHY+25OHD). SAS was used for data analysis, with significance set at P ≤ 0.05. Pre-infection growth performance was comparable across the treatments. However, vaccinated birds exhibited higher body weight (BW) and body weight gain (BWG) from 0 to 6 d postinoculation (DPI; P < 0.05). The NC diet reduced BWG from 6 to 12 DPI and increased the feed conversion ratio (FCR) during 6 to 12 DPI and the overall period (0-26 d) compared to the PC birds. In contrast, the supplementation with phytase, 25OHD, or both, returned BWG and FCR to levels seen with the PC diet (P < 0.01). Vaccinated birds also had reduced gut permeability at 5 DPI, increased intestinal villus height, and lower expression levels of the tight junction proteins junctional adhesion molecule 2 (JAM2) and occludin (OCLN) at 6 DPI (P < 0.05). Interestingly, the cocci vaccine resulted in lower E. acervulina but higher E. tenella oocyst shedding at 6 DPI (P < 0.01). Interaction effects were observed for duodenal lesion scores and ileal crypt depth at 6 DPI (P < 0.05). In conclusion, coccidial vaccination improved growth performance, decreased intestinal permeability, enhanced intestinal morphology, and modulated tight junction protein gene expression under Eimeria infection. Reducing dietary Ca and avP levels adversely affected growth performance and FI during the recovery phase, but these negative effects could be mitigated by supplementing with phytase or 25OHD.

Dose of phytase from either Aspergillus niger or Escherichia coli on performance of nursery piglets.

Supplementing swine diets with phytase increases phosphorus release by approximately 50% from cereal phytates. The increase in phosphorus availability allows for a reduction in dietary phosphorus supplementation from mineral sources and decreases the environmental impact of pork production through a decrease in phosphorus excretion. Superdosing phytase has been reported to boost swine productivity, improve the digestibility of other nutrients, and mitigate the antinutritional effects of phytates. However, there are significant cost differences among phytase products. Bacterial phytases are considered more modern, often with a higher cost of inclusion. A study was conducted with 288 piglets that were 21 d of age and weighed 6.43 ±â€…0.956 kg. Pigs were divided into four groups. Each group of pigs was fed a different experimental diet varying in phytase source and level: fungal phytase (Aspergillus niger) at 500 FTU/kg of diet, fungal phytase at 2,000 FTU/kg, bacterial phytase (Escherichia coli) at 500 FTU/kg, and bacterial phytase at 2,000 FTU/kg. No differences were found for phytase sources or doses on productivity at 14 and 21 d postweaning. However, piglets supplemented with 2,000 FTUs/kg of phytase in the diet during the first 21 d of nursery exhibited a 5.8% better feed conversion (P = 0.02). An interaction between phytase source and dose was observed for average live weight and daily weight gain over the 42-d nursery period (P < 0.05). Supplementing the diet with 2,000 FTU/kg of fungal phytase improved daily weight gain and live weight throughout the experimental period compared to piglets supplemented with 500 FTU/kg of the same phytase source. Additionally, it resulted in better final weights compared to piglets supplemented with 500 FTU/kg of bacterial phytase. Phytase inclusion at 2,000 FTU/kg improved feed conversion by 2.07% over the 42-d nursery period. The most economically favorable feed conversion ratios were observed when supplementing the diet with fungal phytase at 2,000 FTUs/kg.

Effects of Dietary Supplementation of Quillaja Saponin or Phytase on the Growth Performance, Nutrient Digestibility, Faecal Gas Emissions, and Carcass Grade in Growing-Finishing Pigs.

This experiment aimed to evaluate the effects of low doses of Quillaja saponin (QS) or phytase (PHY) on growth performance, nutrient digestibility, faecal gas emissions, and carcass grade in growing-finishing pigs. A total of 72 pigs (Landrace × Yorkshire × Duroc), each weighing 25.82 ± 1.68 kg, were selected and randomly assigned to three treatment groups. Each group had six replicates, with four pigs per pen, and the allocation was based on the four initial body weight and sex of the pigs. They were randomly divided into the following three diet groups: the basal diet as a control (CON) group, the basal diet + 0.02% PHY; and the basal diet + 0.01% QS. The experiment period lasted for 110 days. The results of adding 0.01% QS to the basal diet of pigs show that it can significantly increase the body weight (BW) of growing-finishing pigs on the 110th day (p < 0.05). QS can significantly increase the average daily weight gain (ADG) on Days 80-110 of the experiment (p < 0.05). QS can significantly increase the total average daily weight gain (TADG) of growing-finishing pigs during the entire experimental period (p < 0.05) and has a tendency to improve the average daily feed intake and feed conversion rate during the entire experimental period. However, QS has no significant effect on pig nutrient digestibility and carcass grade. In addition, we also found that QS has a tendency to reduce carbon dioxide emissions. However, adding 0.02% PHY to the basal diet of growing-finishing pigs can only increase the TADG during the entire experimental period. Throughout the experiment, adding PHY to the diet had no significant impact on the nutrient digestibility, faecal gas emissions, and carcass grade of growing-finishing pigs. In summary, adding QS to feed can significantly improve the growth performance of growing-finishing pigs, and has a tendency to improve faecal gas emissions. PHY can only improve the growth performance of growing-finishing pigs.

Microalgae and phytase dietary supplementation improved growth and gut microbiota in juvenile European seabass (Dicentrarchus labrax).

Fishmeal and fish oil have been the main sources of protein and fatty acid for aquaculture fish. However, their increasing price and low sustainability have led the aquafeed industry to seek sustainable alternative feedstuffs to meet the nutritional requirements of fish and improve their health and performance. Plant proteins have been successfully used to replace fishery derivatives in aquafeeds, but the presence of anti-nutritional substances is a potential drawback of this approach. Thus, it has been reported that phytate breakdown can be caused by feed supplementation with exogenous phytase. The inclusion of microalgae has been proposed to improve gut functionality in fish fed diets with a high vegetable protein content. The aim of this study was to evaluate the effect on the growth and gut microbiota of European seabass (Dicentrarchus labrax) juveniles of a diet containing a blend of microalgae (Arthrospira platensis and Nannochloropsis gaditana) and different concentrations of phytase. An 83-day feeding trial was conducted, comprising four experimental diets with 2.5% microalgae and 500, 1,000, 2,000, or 10,000 phytase units (FTU)/kg feed and a microalgae- and phytase-free control diet. At the end of the trial, a significantly increased body weight was observed in fish fed the diet with the highest phytase concentration (10,000 FTU/kg) versus controls, although the gut bacterial composition did not differ from controls in alpha or beta diversity with either majority (Weighted UniFrac) or minority bacterial strains (Unweighted UniFrac). In comparison to the control group, the groups fed diets with 1,000 or 2,000 FTU/kg diets had a lower alpha diversity (Shannon's diversity index), while those fed diets with 500 FTU/kg or 1,000 FTU/kg showed distinct clusters in beta diversity (involving minority ASVs). According to these findings, the diet containing the 2.5% microalgae blend with 10,000 FTU/kg may be useful to increase the aquafeed quality and sustain the growth performance of juvenile European seabass.

Production, purification and characterization of phytase from Pichia kudriavevii FSMP-Y17and its application in layers feed.

INTRODUCTION: Phytase, recognized for its ability to enhance the nutritional value of phytate-rich foods, has has gained significant prominence. The production of this enzyme has been significantly boosted while preserving economic efficiency by utilizing natural substrates and optimizing essential factors. This study focuses on optimizing phytase production through solid-state fermentation and evaluating its effectiveness in enhancing nutrient utilization in chicken diets. OBJECTIVE: The objective is to optimize phytase production via solid-state fermentation, characterize purified phytase properties, and assess its impact on nutrient utilization in chicken diets. Through these objectives, we aim to deepen understanding of phytase's role in poultry nutrition and contribute to more efficient feed formulations for improved agricultural outcomes. METHODOLOGY: We utilized solid-state fermentation with Pichia kudriavzevii FSMP-Y17 yeast on orange peel substrate, optimizing variables like temperature, pH, incubation time, and supplementing with glucose and ammonium sulfate. Following fermentation, we purified the phytase enzyme using standard techniques, characterizing its properties, including molecular weight, optimal temperature and pH, substrate affinity, and kinetic parameters. RESULTS: The optimized conditions yielded a remarkable phytase yield of 7.0 U/gds. Following purification, the enzyme exhibited a molecular weight of 64 kDa and displayed optimal activity at 55 °C and pH 5.5, with kinetic parameters (Km = 3.39 × 10-3 M and a Vmax of 7.092 mM/min) indicating efficient substrate affinity. CONCLUSION: The addition of purified phytase to chicken diets resulted in significant improvements in nutrient utilization and overall performance, including increased feed intake, improved feed conversion ratio, enhanced bird growth, better phosphorus retention, and improved egg production and quality. By addressing challenges associated with phytate-rich diets, such as reduced nutrient availability and environmental pollution, phytase utilization promotes animal welfare and sustainability in poultry production.

Quinoa snack elaborated with Lactiplantibacillus plantarum CRL 1964 sourdough increases the mineral bioavailability in mice.

BACKGROUND: Consumption of pseudocereal-based foods decreased in phytate concentration can provide better nutrition concerning mineral bioavailability. This study aimed to evaluate the mineral bioavailability of quinoa sourdough-based snacks in a murine model. The mice were divided into five groups. One group was fed with basal snacks; three control groups received quinoa-based snacks made from non-fermented dough, dough without inoculum, and chemically acidified dough; and the test group (GF) received quinoa snacks elaborated from sourdough fermented by a phytase-positive strain, Lactiplantibacillus plantarum CRL 1964. Food intake, body weight, and mineral concentration in blood and organs (liver, kidney, and femur) were determined. RESULTS: Food consumption increased during the feeding period and had the highest (16.2-24.5%) consumption in the GF group. Body weight also increased during the 6-weeks of trial. The GF group showed higher (6.0-10.2%) body weight compared with the other groups from the fifth week. The concentrations of iron, zinc, calcium, magnesium, and phosphorus in blood, iron and phosphorus in the liver, manganese and magnesium in the kidney, and calcium and phosphorus in the femur increased significantly (1.1-2.7-fold) in the GF group compared to the control groups. CONCLUSION: The diet that includes quinoa snacks elaborated with sourdough fermented by phytase-positive strain L. plantarum CRL 1964 increased the concentrations of minerals in the blood, liver, kidney, and femur of mice, counteracting the antinutritional effects of phytate. This study demonstrates that the diminution in phytate content and the consequent biofortification in minerals are a suitable tool for producing novel foods. © 2024 Society of Chemical Industry.

A comparative study on the effects of fungal and bacterial phytase with or without citric acid on growth performance, serum mineral profile, bone quality, and nutrient retention in broilers.

Objective: Current research aimed to compare the effects of fungal and bacterial phytase with or without citric acid (CA) on growth performance, serum mineral profile, bone quality, and nutrient retention in birds given non-phytate phosphorus (nPP)-deficient diets. Materials and Methods: A total of 216 Indian River broiler chicks were disturbed into six groups, namely, i) positive control (PC), ii) negative control (NC) contained 0.2% lower nPP than that in the PC diet, iii) NC + fungal phytase (Aspergillus niger), iv) NC + fungal phytase with 2% CA, v) NC + bacterial phytase (Escherichia coli), and vi) NC + bacterial phytase with 2% CA. Results: Compared to the PC group, the NC group showed poor performance, serum phosphorus (P) content, P retention, and bone quality. However, with the inclusion of phytase, all these phenomena were improved. The addition of bacterial phytase showed better values compared with fungal phytase. The main effects of phytase were significant for the feed conversion ratio (FCR), metabolizable energy conversion ratio (MECR), and P retention. The addition of CA, either with fungal or bacterial phytases, did not show considerable beneficial effects on overall performances. However, the main effects of CA were significant on the FCR, MECR, and crude protein conversion ratio. Conclusion: Incorporating bacterial and fungal phytase into low-nPP diets enhanced the broiler's performance. The effects of bacterial phytase were more apparent than those of fungal phytase. However, the efficacy of phytase based on the source might relate to dose, and other factors need further investigation.

Effect of dietary calcium source, exogenous phytase, and formic acid on inositol phosphate degradation, mineral and amino acid digestibility, and microbiota in growing pigs.

The choice of the calcium (Ca) source in pig diets and the addition of formic acid may affect the gastrointestinal inositol phosphate (InsP) degradation and thereby, phosphorus (P) digestibility in pigs. This study assessed the effects of different Ca sources (Ca carbonate, Ca formate), exogenous phytase, and chemical acidification on InsP degradation, nutrient digestion and retention, blood metabolites, and microbiota composition in growing pigs. In a randomized design, 8 ileal-cannulated barrows (24 kg initial BW) were fed 5 diets containing Ca formate or Ca carbonate as the only mineral Ca addition, with or without 1,500 FTU/kg of an exogenous hybrid 6-phytase. A fifth diet was composed of Ca carbonate with phytase but with 8 g formic acid/kg diet. No mineral P was added to the diets. Prececal InsP6 disappearance and P digestibility were lower (P ≤ 0.032) in pigs fed diets containing Ca formate. In the presence of exogenous phytase, InsP5 and InsP4 concentrations in the ileal digesta were lower (P ≤ 0.019) with Ca carbonate than Ca formate. The addition of formic acid to Ca carbonate with phytase diet resulted in greater (P = 0.027) prececal InsP6 disappearance (87% vs. 80%), lower (P = 0.001) InsP5 concentration, and greater (P ≤ 0.031) InsP2 and myo-inositol concentrations in the ileal digesta. Prececal P digestibility was greater (P = 0.004) with the addition of formic acid compared to Ca carbonate with phytase alone. Prececal amino acid (AA) digestibility of some AA was greater with Ca formate compared to Ca carbonate but only in diets with phytase (P ≤ 0.048). The addition of formic acid to the diet with Ca carbonate and phytase increased (P ≤ 0.006) the prececal AA digestibility of most indispensable AA. Exogenous phytase affected more microbial genera in the feces when Ca formate was used compared to Ca carbonate. In the ileal digesta, the Ca carbonate diet supplemented with formic acid and phytase led to a similar microbial community as the Ca formate diets. In conclusion, Ca formate reduced prececal InsP6 degradation and P digestibility, but might be of advantage in regard to prececal AA digestibility in pigs compared to Ca carbonate when exogenous phytase is added. The addition of formic acid to Ca carbonate with phytase, however, resulted in greater InsP6 disappearance, P and AA digestibility values, and changed ileal microbiota composition compared to Ca carbonate with phytase alone.

The study aimed to investigate the effects of dietary calcium sources, exogenous phytase, and formic acid on inositol phosphate (InsP) degradation and nutrient digestibility in ileal-cannulated growing pigs. It also evaluated the concentrations of phosphorus, calcium, and myo-inositol in the blood, the composition of the microbiota in the ileal digesta and feces, and the concentrations of volatile fatty acids in the feces. Replacing calcium carbonate with calcium formate in the feed reduced prececal InsP6 disappearance and phosphorus digestibility. However, adding formic acid to a diet containing calcium carbonate and phytase enhanced prececal InsP6 disappearance and phosphorus digestibility, and increased InsP2 and myo-inositol concentrations in the ileal digesta. The dietary treatments resulted in more pronounced alterations of the microbiota in the feces than the ileal digesta. In ileal digesta, the shifts in relative abundance were primarily evident among low-abundant genera, while in feces, changes were observed in a larger number among genera with higher levels of abundance. The findings of this study suggest that calcium formate is not a suitable alternative to calcium carbonate for phosphorus digestibility in growing pigs. The release of phosphorus from InsP by exogenous phytase can be increased by adding formic acid.

Effects of graded levels of dietary microbial 6-phytase on performance, intestinal histomorphology, caecal microbial population and short-chain fatty acid composition of Lohmann white-classics.

1. This study was conducted to determine the effects of graded levels of phytase on the performance, egg quality and gut health of white laying hens.2. Treatments consisted of a negative control (NC) diet containing 0.14% available phosphorus (avP), positive control (PC) diet containing 0.35% avP provided via dicalcium phosphate (DCP) and DCP replaced in the PC by with three graded levels of phytase derived from Komagataella phaffii at 500 (PC-500), 750 (PC-750) and 1000 (PC-1000) FTU/kg which provided 0.176%, 0.188% and 0.200% of avP, respectively.3. Egg production, feed intake, feed conversion ratio and jejunal morphometry were negatively affected in NC-fed birds (p < 0.05). Considering the whole period, birds fed a diet supplemented with graded levels of phytase shared the same egg production and feed intake levels with PC birds (p < 0.05). Feed conversion ratio was significantly lowered by 4.9%, 1.6% and 7.6% in hens fed on diets PC-500, PC-750 and PC-1000, respectively compared to those fed the PC (p < 0.05).4. Neither of the dietary treatments affected cracked eggs, dirty eggs, eggshell breaking strength and eggshell thickness. Dietary supplementation of phytase significantly increased villus surface area by 15%, 36% and 40% in PC-500, PC-750 and PC-1000 birds, respectively compared to PC (p < 0.05).5. A significant increase in lactobacillus count was observed in line with increasing the level of phytase (p < 0.05). Dietary treatments had no effect on the caecal coliform or aerobic populations. Furthermore, phytase supplementation significantly increased the concentrations of total caecal short-chain fatty acid (SCFA; p < 0.01).6. In conclusion, along with improving performance parameters, the inclusion of phytase in laying hen diets can ameliorate intestinal morphology and stimulate caecal microflora and increase SCFA concentrations.

Exploring the biotechnological potential of novel soil-derived Klebsiella sp. and Chryseobacterium sp. strains using phytate as sole carbon source.

Phosphorus (P) is essential for biological systems, playing a pivotal role in energy metabolism and forming crucial structural components of DNA and RNA. Yet its bioavailable forms are scarce. Phytate, a major form of stored phosphorus in cereals and soils, is poorly bioavailable due to its complex structure. Phytases, enzymes that hydrolyze phytate to release useable phosphorus, are vital in overcoming this limitation and have significant biotechnological applications. This study employed novel method to isolate and characterize bacterial strains capable of metabolizing phytate as the sole carbon and phosphorus source from the Andes mountains soils. Ten strains from the genera Klebsiella and Chryseobacterium were isolated, with Chryseobacterium sp. CP-77 and Klebsiella pneumoniae CP-84 showing specific activities of 3.5 ± 0.4 nkat/mg and 40.8 ± 5 nkat/mg, respectively. Genomic sequencing revealed significant genetic diversity, suggesting CP-77 may represent a novel Chryseobacterium species. A fosmid library screening identified several phytase genes, including a 3-phytase in CP-77 and a glucose 1-phosphatase and 3-phytase in CP-84. Phylogenetic analysis confirmed the novelty of these enzymes. These findings highlight the potential of phytase-producing bacteria in sustainable agriculture by enhancing phosphorus bioavailability, reducing reliance on synthetic fertilizers, and contributing to environmental management. This study expands our biotechnological toolkit for microbial phosphorus management and underscores the importance of exploring poorly characterized environments for novel microbial functions. The integration of direct cultivation with metagenomic screening offers robust approaches for discovering microbial biocatalysts, promoting sustainable agricultural practices, and advancing environmental conservation.

Impact of phytase supplementation on nutrient variability, growth, and digestibility in weaned pigs fed a corn-SBM complex diet.

This study evaluated the effects of different nutrient matrices, with or without phytase supplementation, on growth performance, nutrient digestibility, ileal amino acid (AA) digestibility, and blood inositol in pigs fed a complex diet based on corn-soybean meal. Four hundred newly weaned cross-bred (Landrace × Yorkshire × Duroc) 21-day-old piglets of initial body weight 6.35 ± 1.91 kg were allotted to one of the five dietary treatments: Control (CNT), a corn-soybean-based standard diet; negative control 1 (NC1), a standard diet with reduced available phosphorus (Av.P) (-0.125%), metabolizable energy (ME) (-40 kcal), and crude protein (CP) (-0.3%); NC1 with 500 phytase units per kilogramme (FTU/kg) (N1P5); negative control 2 (NC2), a standard diet with greater reduction of Av.P (-0.150%), ME (-55 kcal), and CP (-0.45%,); and NC2 with 1000 FTU/kg (N2P10). Piglets were housed in a random arrangement based on sex and body weight and data were analyzed as a randomized complete block design using analysis of variance. Results showed that the body weight and average daily gain of the NC2 treatment were lower (p < 0.05) compared to NC2. Gain to feed ratio was greater (p < 0.05) in the CNT and N1P5 treatments compared to the NC1, NC2, and N2P10 treatments. The CP digestibility was higher (p < 0.05) in N1P5 and N2P10 treatments compared to other treatments. Moreover, the digestibility of phosphorus and calcium was higher (p < 0.05) in N1P5 and N2P10 treatments than in CNT, NC1, and NC2 treatments. The digestibility of non-dispensable AA; histidine, isoleucine, leucine, phenylalanine, and valine were increased (p < 0.05) in N1P5 and N2P10 than in CNT, NC1, and NC2 treatments. Nevertheless, the digestibility of dispensable AA, glutamic acid, was higher (p < 0.05) in N1P5 and N2P10 treatments than in CNT, NC1, and NC2 treatments. Blood myo-inositol concentration was higher (p < 0.05) in N1P5 and N2P10 treatments compared to CNT, NC1, and NC2 treatments in phase 2. These results demonstrated enhanced outcomes under conditions of moderate deficiency, whereas more pronounced deficiencies necessitated increased phytase dosages to observe significant improvements. The efficacy of phytase was evident in its ability to elevate average daily gain, gain to feed ratio, phosphorus and calcium, CP, AA, and blood myo-inositol.

Effects of enzyme supplementation on growth performance, digestibility of phosphorus, femur parameters and fecal microbiota in growing pigs fed different types of diets.

A 42-days study was conducted to evaluate the effects of different dietary types (corn-or wheat-soybean meal-based diet) and phytase (Phy) or a multi-carbohydrase and phytase complex (MCPC) supplementation on growth performance, digestibility of phosphorus (P), intestinal transporter gene expression, plasma indexes, bone parameters, and fecal microbiota in growing pigs. Seventy-two barrows (average initial body weight of 24.70 ± 0.09 kg) with a 2 × 3 factorial arrangement of treatments and main effects of diet type (corn-or wheat-soybean meal-based-diets) and enzyme supplementation (without, with Phy or with MCPC). Each group was designed with 6 replicate pens. The MCPC increased (p < 0.05) average daily gain (ADG) and final body weight (BW). A significant interaction (p = 0.01) was observed between diet type and enzyme supplementation on apparent total tract digestibility (ATTD) of P. The ATTD of P was higher (p < 0.05) in wheat soybean meal-based diets compared to corn-soybean meal-based diets. Compared with the corn-soybean meal-based diet, the relative expression of SLC34A2 and VDR genes in the ileum and SLC34A3 in jejunum of growing pigs fed the wheat-soybean meal based diet was lower (p < 0.05). The MCPC significantly reduced (p < 0.05) the relative expression of TRPV5 and CALB1 genes in the ileum and increased the expression of CALB1 in the duodenum compared to control diet. The phytase increased (p < 0.05) the relative expression of SLC34A1 gene in the duodenum in comparison to control diet and MCPC-supplemented diet. The Ca and P contents in plasma from pigs fed corn-soybean meal-based diet were higher (p < 0.05) than those from pigs fed wheat-soybean meal-based diet, and the parathyroid hormone (PTH) and calcitonin (CT) concentrations were lower (p < 0.05) than those fed wheat-soybean meal-based diet. The content of Ca and P in the femur and the bone strength of pigs in the corn-soybean meal group were significantly higher (p < 0.05) than those in the wheat-soybean meal groups. The phytase increased (p < 0.05) the Ca and P content and bone strength of the femur. Additionally, diet type and both enzymes significantly improved fecal microbial diversity and composition. Taken together, diet type and exogenous enzymes supplementation could differently influence the growth performance, utilization of phosphorus, intestinal transporter gene expression, bone mineralization and microbial diversity and composition in growing pigs.

Engineering of the Phytase YiAPPA to Improve Thermostability and Activity and Its Application Potential in Dephytinization of Food Ingredients.

The aim of this study was to modify phytase YiAPPA via protein surficial residue mutation to obtain phytase mutants with improved thermostability and activity, enhancing its application potential in the food industry. First, homology modeling of YiAPPA was performed. By adopting the strategy of protein surficial residue mutation, the lysine (Lys) and glycine (Gly) residues on the protein surface were selected for site-directed mutagenesis to construct single-site mutants. Thermostability screening was performed to obtain mutants (K189R and K216R) with significantly elevated thermostability. The combined mutant K189R/K216R was constructed via beneficial mutation site stacking and characterized. Compared with those of YiAPPA, the half-life of K189R/K216R at 80°C was extended from 14.81 min to 23.35 min, half-inactivation temperature (T50 30) was increased from 55.12°C to 62.44°C, and Tm value was increased from 48.36°C to 53.18°C. Meanwhile, the specific activity of K189R/K216R at 37°C and pH 4.5 increased from 3960.81 to 4469.13 U/mg. Molecular structure modeling analysis and molecular dynamics simulation showed that new hydrogen bonds were introduced into K189R/K216R, improving the stability of certain structural units of the phytase and its thermostability. The enhanced activity was primarily attributed to reduced enzyme-substrate binding energy and shorter nucleophilic attack distance between the catalytic residue His28 and the phytate substrate. Additionally, the K189R/K216R mutant increased the hydrolysis efficiency of phytate in food ingredients by 1.73-2.36 times. This study established an effective method for the molecular modification of phytase thermostability and activity, providing the food industry with an efficient phytase for hydrolyzing phytate in food ingredients.