In mammals, DNA methylation (DNAme) erasure and reinstatement during embryo development and germline establishment are sensitive to the intrauterine environment. Maternal intake of a high-fat diet (HFD), associated with excessive gestational weight gain, has transgenerational effects on offspring health, which may be mediated by changes in DNAme in the germline. Here, we tested the impact of a maternal HFD on embryonic germline DNAme erasure using a rat strain that expresses green fluorescent protein specifically in germ cells. DNAme was analysed by methyl-seq capture in germ cells collected from male and female F1 gonads at gestational day 16. Our data show that although HFD induced global hypomethylation in both sexes, DNAme erasure in female germ cells was more advanced compared to male germ cells. The delay in DNAme erasure in males and the greater impact of HFD suggest that male germ cells are more vulnerable to alterations by exogenous factors.

Polycystic ovary syndrome is associated with increased risks for certain metabolic disorders such as insulin resistance, non-alcoholic fatty liver disease and suppressed ovarian follicular development. This study aimed to examine whether soya isoflavones (ISF) mitigate these polycystic ovary syndrome-associated metabolic disorders in a rat model. Weanling Sprague-Dawley female rats were randomly divided into six groups and were treated with either 0 or 83 µg/d dihydrotestosterone (DHT) to induce polycystic ovary syndrome and fed diets containing 0, 0·5, or 1 g ISF/kg diet for 8 weeks. DHT treatment increased food intake, body weight gain (P < 0·001), percentage of primordial follicles (60 % v. 50·9 %, P < 0·05) and accumulation of lipid droplets in the livers. It also elevated serum total cholesterol, free cholesterol, TAG, NEFA and leptin and hepatic total cholesterol and NEFA. Additionally, DHT treatment reduced the percentage of primary follicles (13·8 % v. 30·2 %, P < 0·05), ovary weight and length (P < 0·001), as well as insulin sensitivity (P < 0·01) compared with the Control. ISF intake at 1 g/kg reduced body weight gain, serum total cholesterol, free cholesterol, NEFA, leptin and hepatic TAG and DHT-induced insulin resistance (P < 0·01). ISF intake at both levels decreased DHT-induced lipid droplet accumulation in the livers and changes in the percentages of primordial and primary follicles. Dietary soya ISF alleviated DHT-induced body weight gain, insulin resistance and hepatic lipid droplet accumulation, as well as suppressed ovarian follicular development. This suggests that the consumption of soya foods or ISF supplements may be beneficial for individuals with polycystic ovary syndrome, mitigating the associated metabolic disorders such as diabetes and non-alcoholic fatty liver disease.

The primary goal of the investigation was to analyse the anti-inflammatory and antioxidant properties of Gamma-linolenic acid (GLA) on rats with indomethacin (IND)-induced gastric ulcers. Thirty rats were divided into five groups: Control, IND (50 mg/kg, p.o.), IND pretreated with GLA 100 mg/kg (p.o. for 14 d), IND pretreated with GLA 150 mg/kg (p.o. for 14 d) and IND pretreated with omeprazole (20 mg/kg, p.o. for 14 d). The stomach tissues were examined to calculate the ulcer index and pH and analyse biochemical markers (prostaglandin E2 (PGE2), cyclooxygenase 1 (COX1), TNF-1, IL-6 and intercellular adhesion molecule-1 (ICAM1)) and oxidative stress parameters (malondialdehyde: (MDA), superoxide dismutase (SOD), glutathione (GSH) and CAT (catalase)) as well as undergo histopathological assessment. GLA 100 and 150 mg/kg showed a protective effect against IND-induced gastric damage. It reduced levels of COX1, TNF-1, IL-6 and ICAM and increased PGE2 levels. GLA also normalised antioxidant function by modulating MDA, SOD, GSH and CAT. GLA intervention protects against IND-induced gastric ulcers by restoring oxidant/antioxidant balance and reducing inflammation.

Hypercholesterolaemia is a major risk factor for CVD. Fish intake is associated with lower risk of CVD, whereas supplementation with n-3 long-chain PUFA (LC-PUFA) has little effect on the cholesterol concentration. We therefore investigated if cetoleic acid (CA), a long-chain MUFA (LC-MUFA) found especially in pelagic fish species, could lower the circulating total cholesterol (TC) concentration in rodents. A systematic literature search was performed using the databases PubMed, Web of Science and Embase, structured around the population (rodents), intervention (CA-rich fish oils or concentrates), comparator (diets not containing CA) and the primary outcome (circulating TC). Articles were assessed for risk of bias using the SYRCLE’s tool. A meta-analysis was conducted in Review Manager v. 5.4.1 (the Cochrane Collaboration) to determine the effectiveness of consuming diets containing CA-rich fish oils or concentrates on the circulating TC concentration. Twelve articles were included in the systematic review and meta-analysis, with data from 288 rodents. Consumption of CA-rich fish oils and concentrates resulted in a significantly lower circulating TC concentration relative to comparator groups (mean difference −0·65 mmol/l, 95 % CI (−0·93, −0·37), P < 0·00001), with high statistical heterogeneity (I2 = 87 %). The risk of bias is unclear since few of the entries in the SYRCLE’s tool were addressed. To conclude, intake of CA-rich fish oils and concentrates prevents high cholesterol concentration in rodents and should be further investigated as functional dietary ingredients or supplements to reduce the risk for developing CVD in humans.

The influence of the method of evaluating developmentally competent oocytes on their viability after cryopreservation still needs to be better understood. The objective of this study was to determine the cleavage and embryo developmental rates after parthenogenetic activation of cumulus–oocyte complexes (COCs) selected by different concentrations of brilliant cresyl blue (BCB) and cryopreservation. In the first experiment, COCs were separated into groups and incubated for 1 h in medium containing BCB (13 μM, 16 μM, or 20 μM). The control group was not exposed to BCB staining. In the second experiment, COCs were divided into four groups: 13 μM BCB(+), 13 μM BCB(−), fresh control (selected by morphologic observation and immediately in vitro matured) and vitrified control (selected by morphologic evaluation, vitrified, and in vitro matured). In the first experiment, the 13 μM BCB group displayed greater development rates at the morula stage (65.45%, 36/55) when compared with the other groups. In the second experiment, cleavage (47.05%, 72/153) and morula development (33.55%, 51/153) of the control group of fresh COCs were increased compared with the other groups. However, when comparing morula rates between vitrified COC control and BCB(+) groups, the BCB(+) group had better results (19.23%, 5/26 and 64.7%, 11/17, respectively). Our best result in rat COC selection by BCB staining was obtained using a concentration of 13 μM. This selection could be a valuable tool to improve vitrification outcomes, as observed by the BCB(+) group that demonstrated better results compared with the vitrified COC control.

The rat model can be used to assess ileal protein digestibility rapidly and in first intention, but no standardised method exists. Our objective was to compare methods to assess protein digestibility, depending on collection site (ileum/caecum) and use of a non-absorbable marker. A meal containing either casein, gluten or pea protein and chromium oxide as non-absorbable marker was given to male Wistar rats and the entire digestive content was collected 6 h later. Total chromium recovery was incomplete and variable, depending on protein source. We observed no significant difference in digestibility between the methods for any of the protein sources tested. Although none of the methods tested is optimal, our results suggest that caecal digestibility can be used as a proxy of ileal digestibility in rats without using a non-absorbable marker. This simple method makes it possible to evaluate protein digestibility of new alternative protein sources for human consumption.

Paul-Louis Simond’s 1898 experiment demonstrating fleas as the vector of plague is today recognised as one of the breakthrough moments in modern epidemiology, as it established the insect-borne transmission of plague. Providing the first exhaustive examination of primary sources from the Institut Pasteur’s 1897–98 ‘India Mission’, including Simond’s notebooks, experiment carnets and correspondence, and cross-examining this material with colonial medical sources from the first years of the third plague pandemic in British India, the article demonstrates that Simond’s engagement with the question of the propagation of plague was much more complex and ambiguous than the teleological story reproduced in established historical works suggests. On the one hand, the article reveals that the famous 1898 experiment was botched, and that Simond’s misreported its ambiguous findings for the Annales de l’Institut Pasteur. On the other hand, the article shows that, in the course of his ‘India Mission’, Simond framed rats as involved in the propagation of plague irreducibly in their relation to other potential sources of infection and not simply in terms of a parasitological mechanism. The article illuminates Simond’s complex epidemiological reasoning about plague transmission, situating it within its proper colonial and epistemological context, and argues for a new historical gaze on the rat as an ‘epidemiological dividual’, which highlights the relational and contingent nature of epidemiological framings of the animal during the third plague pandemic.

Female laboratory rats (Rattus norvegicus; Wistar, Alderley Park) were housed as singletons or groups of three in units of two cages. Units were divided by different types of barrier which allowed varying degrees of social contact across the barrier. Singletons were established either with another singleton on the other side of the barrier or with a group of three as neighbours. Single-housing among females had markedly less effect on time budgeting and pathophysiological measures than among males in a similar, earlier study. In particular, singletons showed a less marked increase in self-directed behaviours, particularly tail chasing, and a smaller reduction in undirected movement around the cage. The smaller reduction in mobility may reflect a greater tendency for singly housed females to attempt escape. Females generally showed much higher levels of escape-oriented behaviours than males and up to a threefold increase in such behaviours when housed singly. Differences in time budgeting and in the apparent significance of social separation between the sexes can be interpreted in terms of differences in socio-sexual strategy and potential mating opportunity, with singleton males responding to their cage as a territory, but singleton females seeking to re-establish social contact. Such an interpretation is consistent with the effects of barrier type on behaviour in singleton females, in which time spent in escape-oriented behaviours reflected the extent to which the barrier facilitated, or frustrated, contact with neighbours.

Textbook recommendations for gavaging rats vary between 1-5 ml for an adult rat. Rats weighing either 130 g or 250 g were gavaged with varying dosages of barium sulphate (BaSO4). After dosing, radiographs were taken at 0, 15 and 60 min. Animals showing a section of the small intestine totally filled with BaSO4 were scored as displaying spontaneous release. Other rats of the same sizes were gavaged with similar doses and subsequently tested in an open-field arena for behavioural abnormalities that might indicate stress or pain resulting from the procedure. Body temperature before and after treatment was recorded using microchip transponders. None of the 250 g rats in the 1 ml dosage group showed spontaneous release through the pyloric sphincter. In the 2 ml and 4 ml dosage groups, only one out of five animals showed spontaneous release. In the 6 ml dosage group, half of the animals showed spontaneous release. In the 8 ml and 10 ml dosage groups, five out of six and four out of five, respectively, showed spontaneous release. If doses were higher than 12 ml, no animal was able to keep all of the BaSO4 in its stomach. In the rats weighing 130 g, the 3 ml dosage group showed only one out of four rats with spontaneous release, whereas in the 5 ml and 7 ml dosage groups, all animals showed spontaneous release. After 15 min, all of the rats in both weight groups showed BaSO4 in the duodenum. Ambulation, rearing up onto the hind legs and defecation, as well as body temperature immediately after dosing, correlated very strongly with the dose (ml kg−1); increasing the dose resulted in reduced ambulation, rearing, defecation and body temperature. However, 10 min after performance of the open-field test, neither body temperature, serum corticosterone nor serum glucose showed any correlation with dose. This study indicates that high doses (ie doses up to 10 ml for a 250 g rat) might be safe to use; however, if an adverse impact on the rat is to be avoided, use of much lower doses should be considered—for example, doses that do not enforce opening of the pyloric sphincter in any rat. This would be less than 4 ml kg−1 in a 250 g rat.

Male laboratory rats (Rattus norvegicus; Wistar, Alderley Park) were housed as singletons or groups of three in units of two joined, but divided cages. Units were divided by different types of barrier that allowed different degrees of social contact across the barrier. Singletons were established either with another singleton as a neighbour on the other side of the barrier, or with a group of three as neighbours. Relative to group-housed animals, singly-housed rats showed reduced activity and a greater incidence of self-directed behaviours and behaviours apparently related to escape or seeking social information. Pathophysiological evidence was consistent with Baenninger's (1967) suggestion that tail manipulation in singletons is a surrogate social response, but was also consistent with an overall increase in self-directed activity, reflecting elasticity in time budgeting. Variation in the degree of increase in self-directed activity among singletons and the negative correlation between self-directed activity and organ pathology may have reflected differences in the ability of individuals to avoid an activity limbo. While reduced corticosterone concentration and organ pathology compared with grouped rats implied that separation may remove social stress, responses to contact with neighbours, and correlations between behaviours and organ pathology suggested that rats may actively seek social interaction. Broad differences in stress responses between single and grouped housing conditions may therefore be an inadequate yardstick to the animals’ welfare. However, exposure to neighbours reduced the aggressiveness of singly-housed males when they were eventually introduced into an unfamiliar group, suggesting that a degree of exposure to neighbours (separation, but not isolation) may have some welfare benefits for laboratory-housed rats, depending on procedures.

Enhancing the complexity of the environments of captive animals is often referred to as environmental enrichment, and aims to have positive effects on the animals’ well-being. Such enrichments may have consequences both for so-called ‘normal’ behaviour and for the pathophysiology of the animals in question. The effects of a lack of environmental complexity, including social isolation, on home cage behaviour and on pathophysiology in rats is considered in this review. Several preference tests on rats — choice tests and operant tests — indicate a preference for bedding, nesting material and social contact. Contradictory research results concerning the need for gnawing objects per se are more difficult to interpret, and it is argued that excessive gnawing may be indicative of primary frustration and hence reduced welfare. One disadvantage of providing environmental enrichment to laboratory animals is a possible increase in subject variability, resulting in the need to use a greater number of test animals. However, this increased variability seems to be inconsistent and is not very well documented. It is argued that in cases where the behavioural benefits of environmental enrichment justify the use of more animals, better welfare should be more highly valued than a reduction in the number of animals used.

High concentrations of carbon dioxide (CO2), used for killing laboratory rodents, are known to be more strongly aversive to rats than sweet food items are attractive. This study investigated whether the maintenance of a high oxygen (O2) concentration, using a gas mixture of 70% CO2 and 30% O2, would reduce aversion to CO2 during a gradual-fill procedure. Eight male Wistar rats, aged 10 months, were housed individually in an apparatus consisting of two cages, one higher than the other and joined by a tube. In a series of trials, subjects entered the lower cage for a reward of 20 sweet food items. The gas was turned on at the moment the rat started eating the reward items and flowed into the lower cage at a fixed rate. There were four treatments: 1) 100% CO2 at 14.5% cage volume min–1; 2) gas mixture at 14.5% min–1; 3) gas mixture at 21.0% min–1, which delivered CO2 at approximately 14.5% min–1 and 4) air, with each subject tested with each treatment four times. Measures of willingness to stay and eat in the lower cage (latency to stop eating, latency to leave and the number of reward items eaten) were much lower in all three gas treatments than in air, indicating that the CO2 and the CO2 + O2 mixture were both more strongly aversive than sweet food items were attractive. Comparing the gas mixture with 100% CO2, the latency to leave and the number of reward items eaten were slightly higher in the CO2 + O2 mixture at 21% min–1 than in CO2 at 14.5% min–1, indicating that the addition of O2 slightly reduced the aversiveness of CO2 in the gradual-fill procedure. This reduction is not enough to warrant recommending the use of CO2 + O2 mixtures for killing rats.

To study the influence of maternal stress on neonatal locomotor development, rat pups of mothers housed singly and in groups were treated orally with corticosterone from 2 to 15 days of age. Control animals received almond oil vehicle only. The rat pups were subjected to swim-tests from 8 to 20 days of age to evaluate locomotor development. Swim-test performance demonstrated a retardation of locomotor development in pups treated with corticosterone (P <0.05). Retardation was most marked in the pups from group-housed mothers and between 13 and 15 days of age. Comparing pups not treated with hormones, the pups born to group-housed mothers showed significantly (P <0.05) better performance on swim-testing. The weight gain of pups from group-housed mothers was significantly (P < 0.05) higher than that of pups from individually caged mothers. Corticosteroid treatment had no effect on weight gain.

Metabolic cages are used for housing rats and mice for up to five days for collection of urine and/or faeces. The small, barren area of the metabolic cage compromises animal welfare as the animals lack a solid floor, shelter, nest material and social contact. We constructed and tested a practically-applicable enrichment device designed to meet behavioural needs for environmental complexity. The influence of this device on the cage preferences and stress levels of the animals was evaluated. A box-shaped enrichment device was designed and implemented in existing metabolic cages. Male Tac:SD rats were housed for five days in an enriched metabolic cage (EMC; n = 12) or a standard metabolic cage (SMC; n = 12), and data were collected on bodyweight, food and water intake, urination and defaecation, as well as urinary corticosterone and creatinine. Moreover, open-field behaviour and cage preferences were assessed. Rats in both groups gained significantly less weight when housed in metabolic cages. Furthermore, SMC rats failed to increase their weight gain after being housed in the metabolic cage. Defaecation was significantly higher in the SMC than in the EMC and so was urinary creatinine. No group differences were found in open-field behaviour. However, in comparing activity before and after housing in the metabolic cage, only SMC animals exhibited significantly lower total activity. In a preference test, a preference for the tunnel connecting the cages in the preference test and a side preference for the left side were found. This side preference was eliminated when the EMC was placed on the right side, whereas the right side was significantly avoided when the EMC was placed on the left side. Based on these results, we conclude that, to some extent, the enrichment device improved the welfare of rats housed in EMC, compared to those in SMC.

This study assessed the welfare of rats (Rattus norvegicus) poisoned with a lethal dose of the methaemoglobin (MetHb) inducing compound para-aminovalerophenone (PAVP). Twenty rats were orally gavaged with either PAVP (treated) or the vehicle only (control). Spontaneous and evoked behaviours were recorded and blood samples collected post mortem for analysis of MetHb%. Female and male rats received a mean (± SEM) dose of 263 (± 3) and 199 (± 6) mg PAVP kg−1, respectively. Mean (± SEM) time to death was 67 (± 16) and 354 (± 71) min for female and male rats, respectively. Control animals did not show any signs of intoxication. The time to death from methaemoglobinaemia in rats was significantly shorter than that reported for anticoagulants and there were no obvious signs of distress or pain.

The way in which animals are fed is an important aspect of their welfare. Not only does food provide the energy and nutrients vital for survival, but feeding is also associated with a number of other factors contributing to the well-being of animals. The feeding method can determine the animals’ abilities to fulfil basic behavioural needs, such as foraging. The aim of this paper is to review and discuss the dilemma of choosing between ad libitum feeding (AL) and dietary restriction (DR). AL can produce obese individuals with severe health problems, though it does appear to be compatible with welfare-friendly management systems. On the other hand, DR is often associated with improved physical health and longevity but can leave animals suffering from hunger, frustration or aggression. The species discussed are the laboratory rat, pigs and poultry all of which are omnivores sharing many characteristics in their eating habits. The welfare implications of different feeding methods depend upon the definition of welfare used. Based on a definition of welfare in terms of functioning, DR could be considered the best way to feed animals, because it results in improved physical health and longevity. If welfare is defined in terms of natural living, it is also a requirement for the animal to be able to engage in natural foraging behaviours. From the feelings-based approach, DR can be viewed as preferable only in circumstances when animals are anticipated to live so long that they would otherwise suffer from the negative long-term consequences of AL. It is argued that incentives are needed to make farmers spend resources to ensure that farm animals are allowed to have their foraging-related needs fulfilled. Feeding of laboratory animals creates special dilemmas when it is important either to under- or over-nourish the animals for experimental purposes, in such instances there is a need for Refinement.

Exposure to high concentrations of CO2 is a common means of stunning and killing laboratory rodents. However, there is concern regarding the potential for animals to have aversive experiences, such as pain or breathlessness, prior to loss of awareness. This preliminary study evaluated the electroencephalographic (EEG) responses of rats (Rattus norvegicus) to CO2 inhalation, using a method based on a minimal anaesthesia model previously used to assess nociception in mammals. Fifteen adult female Sprague-Dawley rats were anaesthetised with halothane in oxygen and maintained at a minimal plane of anaesthesia. EEG was continuously recorded throughout a 10-min baseline period followed by sequential exposure to 5, 15, 30 and 50% CO2. The EEG summary variables median frequency (F50), 95% spectral edge frequency (F95) and total power (PTOT) were derived from the raw EEG. The F50 of the EEG, a sensitive indicator of nociception, increased significantly above baseline during exposure to 15% CO2, suggesting this concentration was noxious to rats. This is consistent with behavioural aversion in rats at around the same CO2 concentration. Stimulation of the rat mucosal nociceptors only occurs at CO2 concentrations of 37% or greater; therefore, it is hypothesised that the observed response was a result of what would have been CO2-induced aversive respiratory sensation in conscious animals, rather than pain. This study provides some evidence that an anaesthesia model may be utilised to study the nocuous effects of low-moderate CO2 exposure in rodents.

Stress cues can affect the welfare of animals in close proximity and are possibly useful non-invasive indicators of the emitters’ welfare. To facilitate their study in murids, we tested whether rats’ stress odours could be collected and stored using an enfleurage-type technique. ‘Donor’ rats were individually exposed to a compound stressor (carried circa 75 m inside a novel container, then euthanised with rising carbon dioxide) while on blotting paper dotted with melted vegetable lard. These sheets were sealed, left at room temperature for 2-5 h, and then ‘bioassayed’ by a blind observer for their effects on conspecifics. Compared with control sheets (exposed to unstressed rats, to CO2 alone, or untreated), stress-exposed sheets significantly affected the unconditioned behaviour of 16 pairs of detector rats trained to enter an arena from their home cage to obtain sucrose. When used to line this arena, the stress-exposed sheets significantly increased: i) rats’ latencies to eat, to place front feet into, and to completely step into the arena and ii) shuttling movements between arena and home cage. These pilot data thus suggest that odours produced by stressed rats can be simply and successfully collected and stored for several hours, though certain potential confounds (eg urine volume) may conceivably be alternative explanations for the observed effects. Future work should control for urine volume, and assess whether fat is needed for optimal odour absorption by paper and for how long sheets can be stored at various temperatures. Much fundamental work is also still needed on the nature, functions, and sources of stress odours.

In a series of experiments the welfare of para-aminovalerophenone (PAVP) sub-lethally poisoned rats (Rattus norvegicus) was assessed. The experiments: (i) examined the acute methaemoglobin (MetHb) profile over time; (ii) refined the LD50 estimate for PAVP in adult female rats; (iii) developed and validated three neurological tests; and (iv) assessed rats for neurological deficit following prolonged methaemoglobinaemia. The results from the first three experiments were used to refine the sub-lethal study. In the sub-lethal experiment 20 rats were gavaged with a single dose of 40 mg kg−1 PAVP (based on an LD50 estimate of 43.3 mg kg−1). Control rats (n = 10) were treated with the carrier only. Eleven (surviving) PAVP-treated rats and controls were assessed over a two-week period. Rats were tested for forelimb grip strength, stability on an inclined plane and the ability to remove tape wrapped around a forepaw in order to determine deficits in motor functions and sensorimotor integration. Signs of recovery began 3-6 h post-dosing, with all animals showing no outward signs of poisoning within 48 h, and over the 14-day post-treatment monitoring period they gained weight and increased their food consumption. There was no significant overall difference in performance between PAVP-treated and control rats in any of the three neurological tests. In the inclined plane test, performance of sub-lethally PAVP-poisoned rats appeared to be temporarily impaired with treated animals slipping at a lower angle than controls on day two. During the tape removal test, four PAVP-treated rats failed to remove the tape within the 3-min time limit on one occasion each (4/77 occasions) up to seven days post-dosing. The severity and duration of signs following acute sub-lethal PAVP poisoning appeared to be lower than those reported for existing rodenticides. It is likely that the results presented in this study extend to other MetHb-inducers.

A high circulating cholesterol concentration is considered an important risk factor for the development of CVD. Since lean fish intake and fish protein supplementation have been associated with lower cholesterol concentration in some but not all clinical studies, the main aim of this study was to investigate the effect of diets containing proteins from fish muscles and fish by-products on the serum/plasma total cholesterol (TC) concentration in rodents. A systematic literature search was performed using the databases PubMed, Web of Science and Embase, structured around the population (rodents), intervention (type of fish and fraction, protein dose and duration), comparator (casein) and the primary outcome (circulating TC). Articles were assessed for risk of bias using the SYRCLE’s tool. A meta-analysis was conducted in Review Manager v. 5·4·1 (the Cochrane Collaboration) to determine the effectiveness of proteins from fish on the circulating TC concentration. Thirty-nine articles were included in the systematic review and meta-analysis, with data from 935 rodents. The risk of bias is unclear since few of the entries in the SYRCLE’s tool were addressed. Consumption of proteins from fish resulted in a significantly lower circulating TC concentration when compared with control groups (mean difference −0·24 mmol/l, 95 % CI − 0·34, −0·15, P < 0·00001), with high statistical heterogeneity (I2 = 71 %). To conclude, proteins from fish muscles and by-products show promise as a functional dietary ingredient or supplement by preventing high cholesterol concentration in rodents, thus reducing one of the most important risk factors for developing CVD.