Improved brewing of millet beer – (OYOKPO) and production of single cell proteins from the spent grain

The improved method for preparing Oyokpo a Nigerian fermented beverage from millet, and the preparation of single cell proteins from the spent grain is described. Improvement of the brew was made by controlled malting, mashing and brewing with a pure culture of Saccharomyces cerevisiae. It had a reducing sugar content of 19.73 g/100 ml before fermentation and after fermentation 5.56% alcohol, 0.58 g/100 ml titratable acidity as acetic acid, a final pH of 4.2 and consisted of a yellowish clear liquid, slightly sour. The native brew had a reducing sugar content of 7.37 g/100 ml before fermentation and after fermentation, 2.40% alcohol, 0.43 g/100 ml titratable acidity, a final pH of 3.8 and consisted of a creamy yellowish liquid with a very sour taste. Fermented spent grain gave a higher protein yield compared to unfermented or ground millet. The lipids, proteins and crude fibre were 4.94%, 11.20% and 4.33% respectively for ground millet, 12,79%, 23.77% and 19.46% respectively for unfermented spent grain and 19.61%, 47.28% and 32.09% respectively for fermented spent grain. The high protein and fibre content of the fermented spent grain points to its potential as a feed supplement for ruminants.     

Improvement of the Antioxidant Activity of Fenugreek Protein Isolates by <Emphasis Type="Italic">Lactococcus lactis</Emphasis> Fermentation

This study investigated the antioxidant capacity of fenugreek protein isolate and its improvement by Lc. lactis fermentation through bioactive peptides production and the effect of molecular weight variation on fenugreek fractions antioxidant activity. Fenugreek protein isolate showed a significant increase of antioxidant and radical scavenging activity after 24 h of fermentation, by about 23.7, 42.9 and 40% for respectively antioxidant activity coefficient AAC, DPPH and ABTS radical scavenging activity. FI fermentation led to a hydrolysis of peptide bands with MW?>?35 kDa and a generation of new bands with a MW?<?25 kDa. A significant reduction in molecular-mass distribution of hydrolysates and a great increase of total free amino acids content, especially an increase on isoleucine, leucine, glutamic acid, serine, histidine, glutamine and lysine was noted. The infrared results showed that different reactions may take place after fermentation, and showed an increase of proteins, amides and aromatic compounds. However, fenugreek fraction (F2) with MW 15–50 kDa presented the highest activity instead of fraction (F1) with lower MW. Lc. lactis had the ability to degrade and convert fenugreek proteins into bioactive peptides that contribute positively in the improvement of antioxidant activity of FI and fractions. FI presents a significant antioxidant activity and thus, can be considered as a potential source of high added value natural antioxidants and may be employed as a functional food ingredient with good potential applications in food products.     

A two-step fermentation of distillers’ grains using Trichoderma viride and Rhodopseudomonas palustris for fish feed

It is important to provide added value or to make full use of the co-product of grains from ethanol production. In order to convert distillers’ grains into a high-quality feed, the Trichoderma viride and Rhodopseudomonas palustris fermentation were combined and investigated in this study. The T. viride fermentation was carried out in an aerobic fermentation installation in favoring of the growth of the fungi and the degradation of the cellulose, and then the fermentation of R. palustris was performed to increase the content of protein with an anaerobic installation. After the two step fermentations, the true protein content of dried distiller’ grains increased from 11.4 to 33.6 % (w/w) (the content of crude protein from 14.5 to 39.7 %), the crude fiber content decreased from 21.3 to 7.6 % (w/w), the crude fat content increased from 5.5 to 7.9 % (w/w), the crude ash decreased from 14.6 to 10.2 % (w/w), the total phosphorus content increased from 0.4 to 1.2 % (w/w), and the water content was 11.8 % (w/w). The dried and fermented grains contain the R. palustris viable count of 5.3 × 10¹¹ CFU/g dry matter. The results may support a new application of an active photosynthetic bacteria fish feed in fisheries industry and offer a reference for the further study of lignocellulosic materials as raw materials converting into high-quality feed.     

Solid-state fermentation of industrial solid wastes from the fruits of milk thistle Silybum marianum for feed quality improvement

The industrial solid wastes generated during the production of silymarin from the fruits of milk thistle Silybum marianum was used as the substrate. Preparation and evaluation of the feeds produced by solid-state fermentation (SSF) of the industrial solid wastes was carried out. The protein content of the fermented feed (FF) from a combination of Aspergillus niger and Candida tropicalis was the highest among the examined strains. The optimal process parameters for protein enrichment with SSF using A. niger and C. tropicalis included incubation temperature of 30.8 °C, fermentation time of 87.0 h, and initial moisture content of 59.7 %. Under these conditions, the value additions of FF occurred. The fiber of FF was decreased by 25.07 %, while the digestibility of protein, protein content, and the ratio of total essential amino acids to total amino acids were increased by 79.85, 16.22, and 8.21 %, respectively. The analysis indicated that FF contained 1.44 mg/kg flavonoids and 0.5 mg/kg silybin, which significantly increased by 2.42 and 1.63 times, respectively than those in unfermented substrates. FF recorded reduced molecular weight of proteins from 20.1 to 44.3 kDa to below 14.3 kDa. The results of feeding trial of FF replacement with soybean meal in broilers diets for 8 weeks showed that FF significantly improved carcass characteristics including abdominal fat rate, serum biochemical parameters including aspartate transaminase, blood urea nitrogen and high density lipoprotein cholesterol, and immune responses of broilers. A potential feed quality improvement was achieved through mixed strains SSF of industrial solid wastes of S. marianum fruits.     

Simultaneous saccharification and fermentation of broken rice: an enzymatic extrusion liquefaction pretreatment for Chinese rice wine production

Broken rice, pretreated by enzymatic extrusion liquefaction, was used to produce Chinese rice wine by simultaneous saccharification and fermentation (SSF) process in this study. The study compared the novel process and traditional process for Chinese rice wine fermentation utilizing broken rice and head rice, respectively. With the optimum extrusion parameters (barrel temperature, 98 °C; moisture content, 42 % and amylase concentration, 1 ‰), 18 % (v/v at 20 °C) alcoholic degree, 37.66 % fermentation recovery and 93.63 % fermentation efficiency were achieved, indicating enzymatic extrusion-processed rice wine from broken rice exhibited much higher fermentation rate and efficiency than traditional-processed rice wine from head rice during SSF. The starch molecule distribution data indicated that the alcoholic degree was related to the oligosaccharides’ formation during enzymatic extrusion. Sum of amino acid (AA) in the extrusion-processed wine was 53.7 % higher than that in the traditional one. These results suggest that the enzymatic extrusion pretreatment for broken rice is a feasible and alternative process in the fermentation of Chinese rice wine.     

Comparative study of mycelia growth and sporophore yield of Auricularia polytricha (Mont.) Sacc on selected palm oil wastes as fruiting substrate

The potential for using agricultural and industrial by-products as substrate for the production of the edible mushroom, Auricularia polytricha, was evaluated using several formulations of selected palm oil wastes mixed with sawdust and further supplemented with selected nitrogen sources. The best substrate formulations were sawdust (SD) mixed with oil palm frond (OPF; 90:10) added with 15 % spent grain (SG) and sawdust mixed with empty fruit bunch (EFB; 50:50) added with 10 % spent grain (SG) with mycelia growth rate of 8 mm/day and 7 mm/day respectively. These two substrate formulations were then subjected to different moisture content levels (65 %, 75 % and 85 %). Highest total fresh sporophore yield at 0.43 % was obtained on SD?+?OPF (90:10)?+?15 % SG at 85 % moisture content, followed closely by SD?+?EFB (50:50)?+?10 % SG with 0.40 % total yield, also at 85 % moisture content. Each of the substrate formulations at 85 % moisture content gave the highest biological efficiency (BE) at 288.9 % and 260.7 %, respectively. Both yield and biological efficiency of A. polytricha on these two formulations were almost three times higher when compared to sawdust substrate alone, thus proving the potential of these formulations to improve yield of this mushroom.     

Cellulosic Butanol (ABE) Biofuel Production from Sweet Sorghum Bagasse (SSB): Impact of Hot Water Pretreatment and Solid Loadings on Fermentation Employing <Emphasis Type="Italic">Clostridium beijerinckii</Emphasis> P260

A novel butanol fermentation process was developed in which sweet sorghum bagasse (SSB) was pretreated using liquid hot water (LHW) pretreatment technique followed by enzymatic hydrolysis and butanol (acetone butanol ethanol (ABE)) fermentation. A pretreatment temperature of 200 °C resulted in the generation of a hydrolyzate that inhibited butanol fermentation. When SSB pretreatment temperature was decreased to 190 °C (0-min holding time), the hydrolyzate was successfully fermented without inhibition and an ABE productivity of 0.51 g L^?1 h^?1 was achieved which is comparable to the 0.49 g L^?1 h^?1 observed in the control fermentation where glucose was used as a feedstock. These results are based on the use of 86 g L^?1 SSB solid loadings in the pretreatment reactors. We were also able to increase SSB solid loadings from 120 to 200 g L^?1 in the pretreatment step (190 °C) followed by hydrolysis and butanol fermentation. As pretreatment solid loadings increased, ABE yield remained in the range of 0.38–0.46. In these studies, a maximum ABE concentration of 16.88 g L^?1 was achieved. Using the LHW pretreatment technique, 88.40–96.00 % of polymeric sugars (cellulose + hemicellulose) were released in the SSB hydrolyzate. The LHW pretreatment technique does not require chemical additions and is environmentally friendly, and the hydrolyzate can be used successfully for butanol fermentation.     

Cultivation of <Emphasis Type="Italic">Chlorella protothecoides</Emphasis> in anaerobically treated brewery wastewater for cost-effective biodiesel production

The use of wastewater has been investigated to overcome the economic challenge involved with a production of microalgae-based biodiesel. In this study, to achieve economical biodiesel production along with effective wastewater treatment at the same time, anaerobically treated brewery wastewater (ABWW) was utilized as a low-cost nutrient source, in the cultivation of Chlorella protothecoides. About 96 and 90 % of total nitrogen and phosphorus in ABWW were removed, respectively, while C. protothecoides was accumulating 1.88 g L^?1 of biomass. The C. protothecoides grown in ABWW showed increases in cell size and cell aggregation, resulting in a near 80 % enhanced harvesting efficiency within 20 min, as compared with only 4 % in BG-11. In addition, the total fatty acid content of the C. protothecoides grown in ABWW increased by 1.84-fold (35.94 ± 1.54 % of its dry cell weight), relative to that of BG-11.     

CAROTENOIDS PRODUCTION IN DIFFERENT CULTURE CONDITIONS BY Sporidiobolus pararoseus

Carotenoids produced by Sporidiobolus pararoseus were studied. It was found that biomass was connected with carbon source, temperature, and pH, but carotenoids proportion was seriously influenced by dissolved oxygen and nitrogen source. Different carotenoids could be obtained by using selected optimum conditions. In the end we established the strategies to produce β-carotene or torulene. Fed-batch fermentation in fermentor was used to prove the authenticity of our conclusions. The cell biomass, β-carotene content, and β-carotene proportion could reach 56.32 g/L, 18.92 mg/L and 60.43%, respectively, by using corn steep liquor at 0–5% of dissolved oxygen saturation. β-Carotene content was 271% higher than before this addition. The cell biomass, torulene content, and torulene proportion could reach 62.47 g/L, 31.74 mg/L, and 70.41%, respectively, by using yeast extract at 30–35% of dissolved oxygen saturation. Torulene content was 152% higher than before this addition. The strategy for enhancing specific carotenoid production by selected fermentation conditions may provide an alternative approach to enhance carotenoid production with other strains.     

Ex situ product recovery for enhanced butanol production by <Emphasis Type="Italic">Clostridium beijerinckii</Emphasis>

In situ butanol recovery fermentation has been intensively studied as an effective alternative to conventional butanol production, which is limited due to the cellular toxicity of butanol. However, the low biocompatibility of adsorbents often leads to failure of in situ recovery fermentations. In this study, Clostridium beijerinckii NCIMB 8052 was cultured in flasks without shaking and in situ recovery fermentation was performed by using an adsorbent L493. The amounts of acetone, butanol, and ethanol (ABE) increased by 34.4 % in the presence of the adsorbent. In contrast, cell growth and production of organic acids and ABE were retarded in the 7-L batch fermentations with in situ butanol recovery. Cell damage occurred in the fermentor upon agitation in the presence of the adsorbent, unlike in static flask cultures with in situ recovery. Ex situ recovery fermentation using circulation of fermentation broth after mid-exponential phase of cell growth was developed to avoid adsorbent-cell incompatibility. No apparent cell damage was observed and 25.7 g/L of ABE was produced from 86.2 g/L glucose in the fed-batch mode using 7 L fermentors. Thus, ex situ recovery fermentation with C. beijerinckii is effective for enhancing butanol fermentation.     

Production of 2,3-Butanediol from Sucrose by a <Emphasis Type="Italic">Klebsiella</Emphasis> Species

Chemical 2,3-butanediol is an important platform compound possessing diverse industrial applications. So far, it is mainly produced by using petrochemical feedstock which is associated with high cost and adverse environmental impacts. Hence, finding alternative routes (e.g., via fermentation using renewable carbon sources) to produce 2,3-butanediol are urgently needed. In this study, we report a wild-type Klebsiella sp. strain XRM21, which is capable of producing 2,3-butanediol from a wide variety of carbon sources including glucose, sucrose, xylose, and glycerol. Among them, fermentation of sucrose leads to the highest production of 2,3-butanediol. To maximize the production of 2,3-butanediol, fermentation conditions were first optimized for strain XMR21 by using response surface methodology (RSM) in batch reactors. Subsequently, a fed-batch fermentation strategy was designed based on the optimized parameters, where 91.2 g/L of 2,3-butanediol could be produced from substrate sucrose dosing in 100 g/L for three times. Moreover, random mutagenesis of stain XMR21 resulted in a highly productive mutant strain, capable of producing 119.4 and 22.5 g/L of 2,3-butanediol and ethanol under optimized fed-batch fermentation process within 65 h with a total productivity of 2.18 g/L/h, which is comparable to the reported highest 2,3-butanediol concentration produced by previous strains. This study provides a potential strategy to produce industrially important 2,3-butanediol from low-cost sucrose.     

Comparison of gamma-aminobutyric acid production in Thai rice grains

Gamma-aminobutyric acid (GABA) has many pharmacological functions including being a major inhibitory neurotransmitter. Two comparative methods for GABA production in rice grains as main food source in Thailand were investigated in this study. Fermentation and germination method were separately carried out using seven selected local grain cultivars in northern Thailand. Red yeast rice, obtained from the fermentation method, gave the higher GABA concentration than the germinated rice produced from the germination method in most rice cultivars. The highest GABA concentration was 28.37 mg/g at 3 weeks fermentation time of glutinous rice, O. sativa L. cv. Sanpatong 1 cultivars, while germinated rice from glutinous rice; O. sativa L. cv. Korkor6 (RD6) cultivars contained the highest GABA concentration of 3.86 mg/g. These results provide information for the basis of an appropriate method for GABA production. The fermentation produced higher GABA concentration but required longer production period and red yeast rice was obtained as product. On the other hand, the germination method yielded rice grains with lower GABA but in more suitable form for consumption. Both methods are considered to be economical and efficient methods to increase GABA in rice grains, providing alternative products with higher nutritional values.     

Brewer's spent grain: characterization and standardization procedure for the enzymatic hydrolysis by Bacillus cereus strain

An important way to reuse agroindustrial by-products and to produce added-value products consists of the production of protein hydrolysates. In the current study, we used Brewer's spent grain (BSG), mainly because of its availability and cost, as a substrate for the enzymatic hydrolysis by Bacillus cereus. First, the physicochemical and microbiological characterization of BSG batches from three varieties was carried out. Furthermore, the optimal fermentation upstream processes for enzymatic hydrolysis by B. cereus were defined. Finally, the ability of B. cereus to hydrolyze different fractions of BSG was analyzed and possible synergistic effects of this bacterium along with other proteolytic bacteria were also investigated. Results showed that the naturally associated microflora was predominantly thermophilic aerobic bacteria and the drying process was the better alternative for BSG preservation. Water, lipids, and ash content differed significantly among the three varieties; however, no statistically significant differences were found in the protein content among them. After BSG characterization studies, the following protocol was set to obtain the fermentation substrate (FS): drying at 60°C for 24-48 H; sieving, grinding, and polyphenol extraction with an alcohol-water solution; and finally autoclaving. A synergistic effect was observed when B. cereus was inoculated with Pseudomonas strains in FS.     

Effect of polyphenol content on the hydrolysis and fermentation of grain sorghum starch

The high polyphenol content of birdproff grain sorghum has been associated with impaired nutritional quality of the grain and with reduced brewing value of birdproof grain sorghum malt due to enzyme inhibition. In this investigation, high polyphenol grain sorghum was evaluated as a feedstock for fermentation ethanol production using NaOH pretreatment to inactivate the polyphenolic compounds prior to hydrolysis with commercial amylases. The polyphenolic inhibition of starch hydrolysis was minimal at a grain sorghum slurry concentration of 20% dry solids, but became pronounced at slurry concentrations of 28% and higher. At these high slurry concentrations the liquefaction and subsequent saccharification and fermentation were markedly improved by alkaline pretreatment. The highest ethanol concentration (12·3%, vol/vol), coupled with the best starch conversion efficiency to ethanol (83·5%), was obtained with a 28% grain sorghum slurry using a partial simultaneous saccharification and fermentation procedure. The residual fermented solids had a crude protein content of 45·4%. Tannic acid decreased yeast cell viability in synthetic media, but had no effect on the hydrolysis or fermentation of grain sorghum starch.     

Enhanced production of tetramethylpyrazine in <Emphasis Type="Italic">Bacillus licheniformis</Emphasis> BL1 by <Emphasis Type="Italic">bdhA</Emphasis> disruption and 2,3-butanediol supplementation

The 2,3-butanediol (2,3-BD) dehydrogenase gene (bdhA) of Bacillus licheniformis BL1 was disrupted to construct the tetramethylpyrazine (TMP)-producing BLA strain. During microaerobic fermentation, the bdhA-disrupted BLA strain produced 46.98 g TMP/l, and this yield was 23.99 % higher than that produced by the parent BL1 strain. In addition, the yield of acetoin, which is a TMP precursor, also increased by 28.98 % in BLA. The TMP production by BL1 was enhanced by supplementing the fermentation medium with 2,3-BD. The yield of TMP improved from 37.89 to 44.77 g/l as the concentration of 2,3-BD increased from 0 to 2 g/l. The maximum TMP and acetoin yields increased by 18.16 and 17.87 %, respectively with the increase in 2,3-BD concentration from 0 to 2 g/l. However, no increase was observed when the concentration of 2,3-BD in the matrix was ≥3 g/l. This study provides a valuable strategy to enhance TMP and acetoin productivity of mutagenic strains by gene manipulation and optimizing fermentation conditions.     

Genetic diversity of bread wheat genotypes in Iran for some nutritional value and baking quality traits

Genetic variation among 78 irrigated bread wheat genotypes was studied for their nutritional value and baking quality traits as well as some agronomic traits. The experiment was conducted in a randomized complete block design with three replicates under normal and terminal drought stress conditions in Kermanshah, Iran during 2012–2013 cropping season. The results of combined ANOVA indicated highly significant genotypic differences for all traits. All studied traits except grain yield, hectoliter weight and grain fiber content were significantly affected by genotype × environment interaction. Drought stress reduced grain yield, thousand kernel weight, gluten index, grain starch content and hectoliter weight and slightly promoted grain protein and fiber contents, falling number, total gluten and ratio of wet gluten to grain protein content. Grain yield by 31.66% and falling number by 9.20% attained the highest decrease and increase due to drought stress. There were negative and significant correlations among grain yield with grain protein and fiber contents under both conditions. Results of cluster analysis showed that newer genotypes had more grain yield and gluten index than older ones, but instead, they had the lower grain protein and fiber contents. It is thought that wheat breeders have bred cultivars with high grain yield, low protein content, and improved bread-making attributes during last seven decades. While older genotypes indicated significantly higher protein contents, and some of them had higher gluten index. We concluded from this study that it is imperative for breeders to pay more attention to improve qualitative traits coordinated to grain yield.     

Fermentation of quinoa and wheat slurries by Lactobacillus plantarum CRL 778: proteolytic activity

Quinoa fermentation by lactic acid bacteria (LAB) is an interesting alternative to produce new bakery products with high nutritional value; furthermore, they are suitable for celiac patients because this pseudo-cereal contains no gluten. Growth and lactic acid production during slurry fermentations by Lactobacillus plantarum CRL 778 were greater in quinoa (9.8 log?cfu/mL, 23.1 g/L) than in wheat (8.9 log?cfu/mL, 13.9 g/L). Lactic fermentation indirectly stimulated flour protein hydrolysis by endogenous proteases of both slurries. However, quinoa protein hydrolysis was faster, reaching 40–100 % at 8 h of incubation, while wheat protein hydrolysis was only 0–20 %. In addition, higher amounts of peptides (24) and free amino acids (5 g/L) were determined in quinoa compared to wheat. Consequently, greater concentrations (approx. 2.6-fold) of the antifungal compounds (phenyllactic and hydroxyphenyllactic acids) were synthesized from Phe and Tyr in quinoa by L. plantarum CRL 778, an antifungal strain. These promising results suggest that this LAB strain could be used in the formulation of quinoa sourdough to obtain baked goods with improved nutritional quality and shelf life, suitable for celiac patients.     

Simultaneous single-cell protein production and COD removal with characterization of residual protein and intermediate metabolites during whey fermentation by K. marxianus

Cheese whey fermentation with Kluyveromyces marxianus was carried out at 40 °C and pH 3.5 to examine simultaneous single-cell protein production and chemical oxygen demand (COD) removal, determine the fate of soluble whey protein and characterize intermediate metabolites. After 36 h of batch fermentation, the biomass concentration increased from 2.0 to 6.0 g/L with 55 % COD reduction (including protein), whereas soluble whey protein concentration decreased from 5.6 to 4.1 g/L. It was confirmed through electrophoresis (SDS-PAGE) that the fermented whey protein was different from native whey protein. HPLC and GC–MS analysis revealed a change in composition of organic compounds post-fermentation. High inoculum concentration in batch fermentation resulted in an increase in biomass concentration from 10.3 to 15.9 g/L with 80 % COD reduction (including protein) within 36 h with residual protein concentration of 4.5 g/L. In third batch fermentation, the biomass concentration increased from 7.3 to 12.4 g/L with 71 % of COD removal and residual protein concentration of 4.3 g/L after 22 h. After 22 h, the batch process was shifted to a continuous process with cell recycle, and the steady state was achieved after another 60 h with biomass yield of 0.19 g biomass/g lactose and productivity of 0.26 g/L h. COD removal efficiency was 78–79 % with residual protein concentration of 3.8–4.2 g/L. The aerobic continuous fermentation process with cell recycle could be applied to single-cell protein production with substantial COD removal at low pH and high temperature from cheese whey.     

Enhancement and modeling of microparticle-added <Emphasis Type="Italic">Rhizopus oryzae</Emphasis> lactic acid production

Lactic acid has a wide industrial application area and can be produced by fungal strains. However, excessive bulk growth form of fungi during the fermentations is a major problem, which limits the fermentation performance. Microparticles are excellent tools to prevent bulk fungal growth and provide homogenized fermentation broth to increase uniformity and the prediction performance of the models. Therefore, in this study, addition of aluminum oxide and talcum microparticles into fermentations was evaluated to enhance the production of lactic acid by Rhizopus oryzae. The results showed that the bulk fungal growth was prevented and the lactic acid concentration increased from 6.02 to 13.88 and 24.01 g/L, when 15 g/L of aluminum oxide or 10 g/L of talcum was used, respectively, in the shake-flask fermentations. Additionally, substrate concentration, pH, and agitation were optimized in the bioreactors using response surface methodology, and optimum values were determined as 126 g/L of glucose, 6.22 pH, and 387 rpm, respectively. Under these conditions, lactic acid production further increased to 75.1 ± 1.5 g/L with 10 g/L of talcum addition. Also, lactic acid production and glucose consumption in the batch fermentation were successfully modeled with modified Gompertz model and modified logistic model. RMSE and MAE values for lactic acid production were calculated as 2.279 and 1.498 for the modified Gompertz model; 3.6 and 4.056 for the modified logistic model. Additionally, modified logistic model predicted glucose consumption with ?2.088 MAE and 2.868 RMSE, whereas these values were calculated as 2.035 and 3.946 for the modified Gompertz model.     

On-Farm Evaluation of an Automatic Enrichment Device with Maize Silage for Laying Hens

Challenges in alternative housing for laying hens are barren functional areas such as winter gardens and the occurrence of behavioral disorders. Environmental enrichment is a measure to deal with these problems. Therefore, an enrichment device offering maize silage automatically was tested in two winter gardens on-farm. The use of the winter gardens and the times individual hens stayed there and occupied themselves with the maize silage were determined in a temporary preference test. The proportion of residing hens was significantly larger in the enriched winter garden. The mean time individual hens stayed in the enriched winter garden ranged from 02:16 ± 02:22 (mm:ss) to 03:17 ± 02:27, whereas the time ranged from 00:18 ± 00:32 to 00:59 ± 01:19 in the other winter garden (p < .05). Once the enrichment device ran in both winter gardens, no differences were found between the observed parameters. On average, the hens occupied themselves with the enrichment material for 03:50 ± 02:12 to 05:01 ± 03:06. Thus, based on its use and acceptance by the laying hens, the automatic device provided adequate environmental enrichment.