Pepsin Trypsin Research Articles

The Potential Anti-inflammatory Effect of Spirulina Platensis on an in Vitro Model of Celiac Disease

Background: Celiac disease (CD) is a prevalent autoimmune enteropathy triggered by the ingestion of gluten. The management of CD involves adhering to a gluten-free diet (GFD). Recent studies have been actively exploring potential supplementary or alternative therapies for individuals with CD. The primary objective of the present study was to assess the effectiveness of Spirulina platensis in regulating the intestinal barrier-related gene expression and alleviating inflammation and oxidative stress associated with CD in PT-gliadin-triggered Caco-2 cells. Methods: S. platensis extracts and a pepsin/trypsin (PT) digest of gliadin were prepared and exposed to the human colon carcinoma Caco-2 cell line. Cell viability was assessed. Total RNA was extracted from Caco-2 cells and cDNA synthesis was performed. A quantitative real-time polymerase chain reaction (qRT-PCR) assay was conducted to evaluate the mRNA levels of interleukin (IL)-6, transforming growth factor beta (TGF-β), COX-2, nuclear factor kappa B (NF-κB), ZO-1, and occludin. Results: Treating Caco-2 cells with S. platensis alone (P=0.01 for both) or in combination with PT-gliadin (P=0.004 and P=0.02, respectively) resulted in decreased IL-6 expression and increased occludin mRNA expression. Additionally, S. platensis extract enhanced Zo-1 mRNA levels (P=0.002) and reduced NF-κB mRNA expression (P=0.02). The combination of gliadin and S. platensis led to decreased mRNA expression of COX-2 (P=0.03) and NF-κB (P=0.04). No significant differences were observed in TGF-β mRNA expression between the studied groups (P>0.05). Conclusion: Additional investigation is needed to examine the influence of interactions between S. platensis and gliadin regarding the comprehensive response of CD to gliadin, encompassing the activation of gluten-sensitive immune cells.

Preparation of pepsin trypsin digested gliadin for stimulation experiments.

Celiac disease (CD) is a complex immune disorder of the intestine that developes in genetically susceptible individuals. CD develops as an intolerance to ingested gluten proteins (gliadins, secalins, hordeins and avenins), being gliadin one of the most immunogenic. Here we present a protocol for the preparation of digested gliadin for laboratory use, a fundamental axis for in vitro and in vivo stimulation studies related to celiac disease research. The importance of a scrupulous handling of materials, products and laboratory instruments to achieve a lipopolysaccharide free gliadin is explained and emphasized. Therefore, in the present chapter, a step-by-step set-up of the protocol for pepsin trypsin gliadin digestion is explained.

EVALUATION OF THE ANTIFUNGAL ACTIVITY OF HYDROLYZED CAMEL WHEY PROTEIN AGAINST SOME FUNGI IN SOFT CHEESE

Purpose: The current study’s aim is to investigate the effect of hydrolyzed whey protein concentrate (WPC) derived from camel’s milk on growth of some fungi inoculated in soft cheese during refrigerated storage.
 Methodology: The pepsin-trypsin (P-T) camel´s WPC hydrolysate (20 mg/g) was incorborated in to soft cheese and their effects on the survival of Candida albicans, Asperigillus fumigatus Asperigillus flavus and Asperigillus niger (103-104cfu/g) were examined till cheese deterioration. 
 Findings: The results revealed that P-T hydrolysate had the ability to decrease the viability of C.albicans, A.fumigatus A.flavus and A.niger in soft cheese. C.albicans was the most sensitive strain.
 Unique contribution to theory, practice and policy: Thus, camel´s WPC hydrolysates can be exploited as. This study was conducted to elaborate antifungals from camel´s WPC after enzymatic hydrolysis which could serve as a safe alternative of natural antifungal agents in soft cheese

VALUATION OF THE ANTIFUNGAL ACTIVITY OF HYDROLYZED CAMEL WHEY PROTEIN AGAINST SOME FUNGI IN SOFT CHEESE

Purpose: The current study’s aim is to investigate the effect of hydrolyzed whey protein concentrate (WPC) derived from camel’s milk on growth of some fungi inoculated in soft cheese during refrigerated storage.
 Methodology: The pepsin-trypsin (P-T) camel´s WPC hydrolysate (20 mg/g) was incorborated in to soft cheese and their effects on the survival of Candida albicans, Asperigillus fumigatus Asperigillus flavus and Asperigillus niger (103-104cfu/g) were examined till cheese deterioration. 
 Findings: The results revealed that P-T hydrolysate had the ability to decrease the viability of C.albicans, A.fumigatus A.flavus and A.niger in soft cheese. C.albicans was the most sensitive strain.
 Unique contribution to theory, practice and policy: Thus, camel´s WPC hydrolysates can be exploited as. This study was conducted to elaborate antifungals from camel´s WPC after enzymatic hydrolysis which could serve as a safe alternative of natural antifungal agents in soft cheese

IMPACT OF HYDROLYZED CAMEL´S WHEY PROTEIN CONCENTRATE ON SOFT CHEESE QUALITY

Purpose: The current study’s aim is to investigate the effect of hydrolyzed whey protein concentrate (WPC) derived from camel’s milk on quality and organoleptic properties of soft cheese during refrigerated storage.
 Methodology: Two concentrations (10 and 20 mg/g) of camel´s WPC and its hydrolysates [pepsin (P) and pepsin-trypsin (P-T) hydrolysates] were incorborated in to soft cheese and their effects on total bacterial, phsychrophilic, aerobic spore formers, coliforms and yeast & mold counts were calculated till end of refrigerated storage peroid. Also, flavour, texture and appearance of treated cheese groups were evaluated compared to control one. 
 Findings: The results revealed that the higher concentration of WPC and its hydrolysates, the more significant decrease in the microbial load and increase the shelf life up to 34th days with P-T hydrolysate (20 mg/g) compared with the control with a shelf-life of 18th days only at refrigerated temperature (4oC).This hydrolysate showed also the highest degree of hydrolysis (DH%) of 34.06% ± 1.53 and protein concentration of 30.72%± 3.16. The maximum score for body, and texture and appearance was recorded for the cheese sample containing P-T hydrolysate (20 mg/g), while the maximum flavour score was recorded to pepsin (P) hydrolysate (10 mg/g), compared with unhydrolyzed WPC concentrations and control soft cheese groups. 
 Unique contribution to theory, practice and policy: This study was conducted to elaborate antimicrobials from camel´s WPC after enzymatic hydrolysis which could serve as a potential natural presrvatives in soft cheese without altering the sensory characteristics.

ACE inhibitory effect of the protein hydrolysates prepared from commercially available nori product by pepsin–trypsin digestion

ACE inhibitory effect of the protein hydrolysates prepared from commercially available nori product by pepsin–trypsin digestion

Human gut-derived commensal suppresses generation of T-cell response to gliadin in humanized mice by modulating gut microbiota

The human intestinal tract is colonized by a large number of diverse microorganisms that play various important physiologic functions. In inflammatory gut diseases including celiac disease (CeD), a dysbiotic state of microbiome has been observed. Interestingly, this perturbed microbiome is normalized towards eubiosis in patients showing recovery after treatment. The treatment has been observed to increase the abundance of beneficial microbes in comparison to non-treated patients. In this study, we investigated the effect of Prevotella histicola or Prevotella melaninogenica, isolated from the duodenum of a treated CeD patient, on the induction and maintenance of oral tolerance to gliadin, a CeD associated subgroup of gluten proteins, in NOD.DQ8.ABo transgenic mice. Conventionally raised mice on a gluten free diet were orally gavaged with bacteria before and after injection with pepsin trypsin digested gliadin (PTD-gliadin). P. histicola suppressed the cellular response to gliadin, whereas P. melaninogenica failed to suppress an immune response against gliadin. Interestingly, tolerance to gliadin in NOD.DQ8.ABo mice may be associated with gut microbiota as mice gavaged with P melaninogenica harbored a different microbial diversity as compared to P. histicola treated mice. This study provides experimental evidence that gut microbes like P. histicola from treated patients can suppress the immune response against gliadin epitopes.

Effect of Pepsin–Trypsin In Vitro Gastro-Intestinal Digestion on the Antioxidant Capacities of Ultra-Filtrated Rice Bran Protein Hydrolysates (Molecular Weight > 10 kDa; 3–10 kDa, and < 3 kDa)

The antioxidant capacities of rice bran protein hydrolysates (RBPH) obtained by in vitro gastro-intestinal (GI) digestion by pepsin–trypsin (P–T) system were investigated. The hydrolysates were separated by ultrafiltration into three fractions (FI: molecular weight (MW) > 10 kDa, FII: MW 3–10 kDa, and FIII: MW < 3 kDa). Furthermore, the protein content and free amino acids of hydrolysates were studied. The results showed that fraction FII with MW 3–10 kDa had the highest antioxidant activity: DPPH of 28.34 ± 0.16%, ABTS of 30.66 ± 0.29%, and metal chelating activity of 61.04 ± 0.38%, and was further purified by RP-HPLC. Four sub-peptide fractions (F1, F2, F3, and F4) were collected and their antioxidant activities were assessed. The results displayed that F2 sub-peptide fraction owned a higher metal chelating activity (72.09 ± 0.32%) than other sub-peptide fractions. The amino acids composition of the purified peptide (F2) was evaluated, which could play an essential role in its antioxidant activity. The results indicated that rice bran proteins are rich in antioxidant peptides which can be employed as a potential source of natural bioactive compounds. The peptides produced can resist physiological digestion in the GI tract.

Identification and Active Evaluation of Antioxidant Peptides from Protein Hydrolysates of Skipjack Tuna (Katsuwonus pelamis) Head.

For the full use of fish by-products to produce antioxidant peptides, skipjack tuna (Katsuwonus pelamis) heads generated during can processing were defatted and hydrolyzed using the in vitro gastrointestinal (GI) digestion (pepsin–trypsin system) method and six antioxidant peptides (P1 to P6) were purified from the head hydrolysate (KPH) using ultrafiltration and serial chromatography methods. Six isolated peptides (P1 to P6) were identified as Val-Glu-Glu (VEE, P1), Trp-Met-Phe-Asp-Trp (WMFDW, P2), Asp-Ala-Gly-Pro-Tyr-Gly-Pro-Ile (DAGPYGPI, P3), Trp-Met-Gly-Pro-Tyr (WMGPY, P4), Glu-Arg-Gly-Pro-Leu-Gly-Pro-His (ERGPLGPH, P5), and Glu-Met- Gly-Pro-Ala (EMGPA, P6), respectively, using a protein sequencer and electrospray ionization-mass spectrometer. Among skipjack tuna head hydrolysates, fractions, and six isolated peptides (P1 to P6), WMFDW (P2), WMGPY (P4), and EMGPA (P6) showed the highest radical scavenging activities on 2,2-diphenyl-1-picrylhydrazyl (DPPH) (EC50 values of 0.31, 0.33, and 0.46 mg/mL for WMFDW, WMGPY, and EMGPA, respectively), hydroxyl (EC50 values of 0.30, 0.43, and 0.52 mg/mL for WMFDW, WMGPY, and EMGPA, respectively), and superoxide anion (EC50 values of 0.56, 0.38, and 0.71 mg/mL for WMFDW, WMGPY, and EMGPA, respectively). Moreover, WMFDW, WMGPY, and EMGPA showed strong capability in reducing power and lipd peroxidation inhibition in the linoleic acid system. In addition, WMFDW, WMGPY, and EMGPA can retain strong antioxidant activity at temperatures lower than 60 °C and pH values ranged from 5 to 9. The results showed that six isolated peptides (P1 to P6) from skipjack tuna heads, especially WMFDW, WMGPY, and EMGPA, might be applied in health care products acting as powerful antioxidant agents.

Variable Immunogenic Potential of Wheat: Prospective for Selection of Innocuous Varieties for Celiac Disease Patients via in vitro Approach.

Celiac Disease (CD) is a multifactorial, autoimmune enteropathy activated by cereal proteins in genetically predisposed individuals carrying HLA DQ2/8 genes. A heterogenous gene combination of the cereal prolamins is documented in different wheat genotypes, which is suggestive of their variable immunogenic potential. In the current study, four wheat varieties (C591, C273, 9D, and K78) identified via in silico analysis were analyzed for immunogenicity by measuring T-cell proliferation rate and levels of inflammatory cytokines (Interferon-γ and Tumor Necrosis Factor-α). Peripheral Blood Mononuclear Cells and biopsy derived T-cell lines isolated from four CD patients in complete remission and two controls were stimulated and cultured in the presence of tissue transglutaminase activated pepsin-trypsin (PT) digest of total gliadin extract from test varieties. The immunogenicity was compared with PBW 621, one of the widely cultivated wheat varieties. Phytohaemagglutinin-p was taken as positive control, along with unstimulated cells as negative control. Rate of cell proliferation (0.318, 0.482; 0.369, 0.337), concentration of IFN- γ (107.4, 99.2; 117.9, 99.7 pg/ml), and TNF- α (453.8, 514.2; 463.8, 514.2 pg/ml) was minimum in cultures supplemented with wheat antigen from C273, when compared with other test varieties and unstimulated cells. Significant difference in toxicity levels among different wheat genotypes to stimulate celiac mucosal T-cells and PBMC's was observed; where C273 manifested least immunogenic response amongst the test varieties analyzed.

Properties of bovine gelatin as affected by a cross-linking induced by horseradish peroxidase, glucose oxidase and glucose

Gelatin is an important protein-based hydrocolloid, providing excellent gelling properties in processed foods. Horseradish peroxidase (100 U/g protein), glucose oxidase (1 U/g protein), and glucose (0.025 mmol/g protein) were applied to cross-link bovine gelatin at 37 °C for 3 h, and the gelling properties of cross-linked gelatin were evaluated. Obviously, the cross-linking process did not change the composition of amino acids. The cross-linked gelatin had 39.2% increase in surface hydrophobicity, 54.7% increase in emulsion stability index, but 28.2% decrease in emulsifying activity index, in comparison with bovine gelatin. Laser scanning confocal microscopy observation indicated that cross-linked gelatin rather than bovine gelatin conferred better droplet stability on the generated emulsion after long-storage period of 48 h or 7 days. Moreover, cross-linked gelatin had decreased in vitro digestibility during pepsin hydrolysis (38.5%) and pepsin–trypsin hydrolysis (14.5%) than bovine gelatin. Finally, the gelling and melting temperatures of cross-linked gelatin were 2 °C higher than those of bovine gelatin. Overall, the cross-linking process of bovine gelatin led to easier gelation and improved heat resistance of the subsequently gels.

Biofunctional properties of bioactive peptide fractions from protein isolates of moringa seed (Moringa oleifera).

Moringa oleifera (Moringaceae) is a specie of significant importance because of its multiple nutraceutical properties, that has led to increase in its consumption. The seeds contain a high percentage of protein (37.48%). However, little is known about the bioactive properties of these proteins and peptides, especially those generated by enzymatic hydrolysis. The objective of this study was to evaluate the biofunctional properties of total hydrolysates (TH) and peptide fractions from protein isolates of moringa seeds. Isoelectric protein isolates were prepared and TH were obtained by digestion with trypsin, chymotrypsin and pepsin-trypsin for 2.5 and 5h. TH were fractioned by ultrafiltration (UF) with a 10kDa membrane to generate the peptide fractions. In all treatments, the antioxidant capacity was significantly higher in peptide fractions>10kDa with 5h of hydrolysis. The results showed that the fraction>10kDa of pepsin-trypsin digested for 5h presented a better Angiotensin Converting Enzyme inhibition (ACE-I) activity with an IC50 of 0.224μg/μl. Also, antidiabetic activity was enhanced in pepsin-trypsin treatment with 5h of hydrolysis showing an IC50 of 0.123μg/μl. Finally, this study showed that hydrolysates of moringa seed proteins had excellent in vitro nutraceutical potential.

Preparation and characterisation of glyceollin‐enriched soya bean protein using solid‐state fermentation

SummaryGlyceollins, stress‐induced phytoalexins from parent soya bean isoflavones, were elicited with Aspergillus oryzae. This solid‐state fermentation facilitated the conversion of isoflavone glycosides into aglycones and glyceollins, which could mainly enrich in soya bean protein isolate (SPI) during protein preparation due to protein–polyphenol interactions. Glyceollin‐enriched SPI exhibited less flavour volatiles, higher solubility, lower whiteness and higher antioxidant activity than unfermented SPI. Fermented SPI was more easily to be digested during in vitro pepsin–trypsin digestion. This may be attributed to partial degradation of protein, especially α′ and α subunits of β‐conglycinin and acidic subunit of glycinin. The antioxidant activity of digestive products derived from fermented SPI was obviously enhanced with increasing digestion time due to simultaneous release of antioxidant peptides and glyceollins involved in the interior of protein molecule. These results suggest an effective technique for producing a nutrient‐enhanced SPI as novel functional ingredients applied in food industry.

Identification of buffalo casein-derived bioactive peptides with osteoblast proliferation activity

Osteoblast cells are the building blocks of bone architecture and most suitable cells for screening osteoanabolic agents. As bone formation begins with osteoblast proliferation, it is the essential stage for screening osteoanbolics. LC–MS/MS analysis of the 11th RP–HPLC fraction of pepsin–trypsin (PT) hydrolysates of buffalo casein showed 15 peptides. Out of these 15 peptides, four peptides of less than 1 kDa were custom-synthesized and evaluated for osteoblast proliferating ability. Proliferation effect of the peptides was studied by formazan crystal formation (MTT method) and relative expression of proliferation marker genes (clay and cdk2). PT hydrolysates and all the four peptides significantly induced osteoblast proliferation indicated by both MTT and relative gene expression studies. This study showed the osteoblast-promoting property of peptides derived from milk proteins, thereby further substantiating the earlier epidemiological data regarding bone health-promoting effect of milk.

Influence of Preparation Conditions on the Properties of Keratin-Based Polymer Hydrogel

In this study, a novel keratin-based polymer hydrogel (K-PmaH) was successfully prepared by graft-copolymerization of methacrylic acid onto keratin, cross-linked with N,N’-methylene-bis-acrylamide, then blended with agar. The influence of reaction temperature, dosage of reductant, initiator, and monomers on graft-copolymerization and that of heating methods on blending process was investigated. The results demonstrated that the dosage of DTT has prominent effects on the graft-copolymerization, and microwave radiation has significant effects on hydrogel properties. The swelling properties and in vitro degradation characteristic of K-PmaH also have been investigated, respectively. Determination results indicate that the K-PmaH has moderate swelling capacity in water and swelling–shrinkage behavior in saline solution. Moreover, K-PmaH reveals an excellent degradable characteristic in pepsin solution and trypsin solution. Consequently, the hydrogel K-PmaH is expected to be applied to the wounds and burns covering materials or bandages.

Immunological characterization of the gluten fractions and their hydrolysates from wheat, rye and barley.

Gluten proteins in wheat, rye and barley cause celiac disease, an autoimmune disorder of the small intestine, which affects approximately 1% of the world population. Gluten is comprised of prolamin and glutelin. Since avoidance of dietary gluten is the only option for celiac patients, a sensitive gluten detection and quantitation method is warranted. Most regulatory agencies have set a threshold of 20 ppm gluten in foods labeled gluten-free, based on the currently available ELISA methods. However, these methods may exhibit differences in gluten quantitation from different gluten-containing grains. In this study, prolamin and glutelin fractions were isolated from wheat, rye, barley, oats and corn. Intact and pepsin-trypsin (PT)-digested prolamin and glutelin fractions were used to assess their immunoreactivity and gluten recovery by three sandwich and two competitive ELISA kits. The Western blots revealed varied affinity of ELISA antibodies to gluten-containing grain proteins and no reactivity to oat and corn proteins. ELISA results showed considerable variation in gluten recoveries from both intact and PT-digested gluten fractions among different kits. Prolamin fractions showed higher gluten recovery compared to their respective glutelin fractions. Among prolamins, barley exhibited higher recovery compared to wheat and rye with most of the ELISA kits used. Hydrolysis resulted in reduced gluten recovery of most gluten fractions. These results suggest that the suitability of ELISA for accurate gluten quantitation is dependent upon various factors, such as grain source, antibody specificity, gluten proteins and the level of their hydrolysis in foods.

A comparative in vitro study of the digestibility of heat- and high pressure-induced gels prepared from industrial milk whey proteins

We undertook this study to compare the digestibility of heat- and high pressure-induced gels produced from whey protein isolate (WPI). To simulate in vivo gastrointestinal digestion of WPI gels, a pepsin–trypsin digestion system was used. The in vitro protein digestibility of WPI gels induced by high pressure (400 MPa and 30 min; P-gel) and those induced by heat (80°C and 30 min; H-gel) was compared using a protein concentration of 0.14 g mL−1. The in vitro protein digestibility of P-gels was significantly greater than that of H-gels (p<0.05). The size-exclusion chromatography profiles of the hydrolysates showed that the P-gel generated more and smaller peptides than natural WPI and H-gels. Furthermore, Sodium dodecyl sulfate–polyacrylamide gel electrophoresis analysis showed some soluble disulfide-mediated aggregation in the P-gel, while there was more insoluble aggregation in the H-gel than the P-gel. The P-gel was more sensitive to proteinase than the H-gel, which was related to the content of S–S bonds, and this in turn could be attributed to the differences in the gelation mechanism between the H-gel and P-gel.

Functional Properties of Protein Isolates from Caragana korshinskii Kom. Extracted by Three Different Methods

Seeking cheap, sustainable protein sources greatly facilitates in alleviating the dependence upon expensive animal-based protein in many developing countries. Caragana korshinskii Kom. offers a good alternative feedstock because of its high-content of protein, low fertilizer and pesticide requirements, excellent stress (high salty and less water) tolerance, wide adaptability, etc. The functional properties of C. korshinskii Kom. protein isolates by three different extraction methods were investigated. The extraction processes greatly influenced the physiological characteristics of protein isolates. C. korshinskii Kom. protein isolate by traditional alkaline extraction (Al-CPI) exhibited good performance on emulsifying activity index, oil and water absorption capacity, and foaming property compared to A-CPI ( C. korshinskii Kom. protein isolate by the acetone precipitation method) and TCA-CPI ( C. korshinskii Kom. protein isolate by trichloroacetic acid-acetone precipitation). The water and oil adsorption capacities of Al-CPI were observed at 4.99 and 3.45 g/g, respectively, even much higher than those of soy protein isolate (SPI) (3.94 and 2.95 g/g, respectively). The highest foaming capacity was observed by Al-CPI at 185.0%, followed by A-CPI (177.5%), TCA-CPI (142.5%), and SPI (141.9%), respectively. It has to be noted that A-CPI showed good solubility at acidic pH and excellent in vitro digestibility. After sequential pepsin-trypsin digestion, the percentage of N release of A-CPI reached up to 83.7%, which was 1.63 times that of Al-CPI (51.2%), 1.19 times that of TCA-CPI (70.1%), and slightly higher than that of the commercial SPI (82.5%). These results indicate that C. korshinskii Kom. holds great potential for application in the animal feed and food additive industry.

Functional properties of limited hydrolysed cross‐linked casein–gelatin composites

SummaryA casein–gelatin composite was prepared by cross‐linking of caseinate and bovine gelatin with a microbial transglutaminase, and the impact of limited proteolysis by trypsin on some functional properties of the composite was investigated in the present work. Two hydrolysed composites were prepared with degree of hydrolysis (DH) of 1 and 2% and analysed by SDS‐PAGE to reflect polypeptide profiles. Some functional properties of the two hydrolysed composites were evaluated, among which solubility, digestibility in vitro, surface hydrophobicity and emulsifying properties showed dependence on the DH. Limited proteolysis of the composite improved its solubility in pH 3–10, especially when the DH was 2%. Compared to the composite, the hydrolysed composites showed an increased emulsifying activity index (about 357–408%), emulsion stability index (about 23–28%) and digestibility in vitro (about 65–80% for pepsin, whereas 2–3% for pepsin–trypsin hydrolysis), together with a decreased surface hydrophobicity (about 55–62%) and oil absorption capacity (about 20–23%). The applied proteolysis also led to the aqueous dispersions of the two hydrolysed composites much lower apparent viscosity, storage and loss modulus.

The sourdough fermentation may enhance the recovery from intestinal inflammation of coeliac patients at the early stage of the gluten-free diet

This study aimed at investigating the effect of corn, rice and amaranth gluten-free (GF) sourdoughs on the release of nitric oxide (NO) and synthesis of pro-inflammatory cytokines by duodenal mucosa biopsies of eight coeliac disease (CD) patients. Selected lactic acid bacteria were used as starters for the manufacture of corn, rice or amaranth sourdoughs. Chemically acidified doughs, without bacterial starters, and doughs started with baker's yeast alone were also manufactured from the same GF matrices. Pepsin-trypsin (PT) digests were produced from all sourdoughs and doughs, and used to assay the rate of recovery of biopsy specimens from eight CD patients at diagnosis. The release of NO and the synthesis of pro-inflammatory cytokines interferon-γ (IFN-γ) were assayed. During fermentation, lactic acid bacteria acidified and grew well (ca. log 9.0 CFU/g) on all GF matrices, showing intense proteolysis. Duodenal biopsy specimens still released NO and IFN-γ when subjected to treatments with basal medium (control), PT-digest from chemically acidified doughs and PT-digest from doughs fermented with baker's yeast alone. On the contrary, the treatment of all the biopsy specimens with PT-digests from all GF matrices subjected to sourdough fermentation significantly decreased the release of NO and the synthesis of IFN-γ. During manufacture of GF baked goods, the use of sourdough fermentation could be considered as an adjuvant to enhance the recovery from intestinal inflammation of coeliac patients at the early stage of the gluten-free diet.