Combination of carotenoids and inulin and uses thereof

Abstract

The present invention provides a combination of at least one carotenoid and inulin for use in affecting the gut microbiota in a subject suffering from IBS. Also provided is a combination of at least one carotenoid and inulin for use in affecting the gut microbiota in a subject suffering from overweight or obesity. The combination can be used as a medicament, a dietary supplement, or a food supplement. Preferably, the combination is for use in affecting the gut microbiota in a human IBS subject or overweight subject in comparison to the administration / consumption of inulin alone and the carotenoid alone.

Description

[0001] Combination of carotenoids and inulin and uses thereof

[0002] TECHNICAL FIELD

[0003] The present invention relates to a combination of at least one carotenoid and inulin. The combination positively affects the gut microbiota in subjects with irritable bowel syndrome (IBS) and in overweight subjects and is useful as a medicament, a dietary supplement, or a food supplement.

[0004] BACKGROUND

[0005] Irritable bowel syndrome (IBS) is a functional gastrointestinal disorder characterized by a range of symptoms that usually include abdominal pain, bloating and changes in the consistency of bowel movements. These symptoms can occur over a long period of time, sometimes for years. It is assumed that around 10-15% of people in industrialized countries are affected by IBS. IBS can have a noticeable impact on quality of life, and disorders such as anxiety, severe depression and chronic fatigue syndrome are common in people with IBS.

[0006] The treatment of IBS is aimed at improving the symptoms. This can include dietary changes, medication, probiotics and counselling. Thus, a healthy diet is an important factor for preventing and managing IBS.

[0007] According to the World Health Organization, in 2022, 2.5 billion adults aged 18 years and older were overweight, including over 890 million adults who were living with obesity. The prevalence of overweight and obesity is a growing problem worldwide and poses major challenges to public health. This growing problem is due to a complex interplay of factors, including lack of exercise, unhealthy eating habits, genetic predispositions and environmental influences. Overweight people have an increased risk of various chronic diseases, including cardiovascular disease, type 2 diabetes, certain cancers and musculoskeletal disorders.

[0008] Adhering to weight-loss interventions is a challenge for many people suffering from overweight or obesity. Weight-loss success varies from person to person and depends on a complex interplay between environmental, genetic, neural and endocrine factors. This variability in weight-loss responses between individual persons could partly depend on the baseline gut microbiota composition. Next to increasing physical activity, a healthy diet is an important factor for preventing and managing overweight and obesity.

[0009] Probiotics and prebiotics can contribute to a healthy diet. Prebiotics are conventionally regarded as indigestible food components that can reach the gut and be selectively utilized by the host microbiota, thereby providing health benefits. There are a number of molecules in foods that are not fully digestible and can therefore reach the colon and its microbiota. Inulin is a prebiotic food additive. Inulin is not absorbed in the small intestine as humans lack the degrading enzyme (inulinase). Instead, it is fermentatively broken down by bacteria in the large intestine to form Short-Chain Fatty Acids (SCFAs).

[0010] Butyrate, a short-chain fatty acid (SCFA), plays several important roles in the gut, and is found to be reduced in patients suffering from irritable bowel syndrome (IBS). M. Pozuelo et al., “Reduction of butyrate- and methane-producing microorganisms in patients with Irritable Bowel Syndrome", Sci. Rep. 5, 12693; doi: 10.1038 / srep12693 (2015).

[0011] Based on the established effects of butyrate in the gut, the following connections between butyrate level in the gut and IBS are possible:

[0012] • Energy Source: Butyrate serves as a crucial energy source for the cells lining the colon (colonocytes). These cells rely heavily on butyrate as a primary fuel. In IBS, disturbances in the gut microbiota can affect the production of butyrate, potentially leading to reduced energy supply to the colonocytes.

[0013] • Anti-inflammatory Effects: Butyrate has anti-inflammatory properties, helping to maintain the health of the gut lining and modulate the immune response. In IBS, where low-grade inflammation and immune dysregulation may play a role in symptomatology, butyrate's antiinflammatory effects could be beneficial.

[0014] • Regulation of Gut Motility: Butyrate has been implicated in regulating gut motility, which refers to the movement of food and waste through the digestive tract. In IBS, abnormal gut motility is a common feature, with symptoms ranging from diarrhea-predominant to constipation-predominant. Butyrate may help to regulate this motility, potentially alleviating symptoms.

[0015] • Maintenance of Gut Barrier Function: Butyrate contributes to maintaining the integrity of the gut barrier, which is essential for preventing the entry of harmful substances into the bloodstream. Dysfunction of the gut barrier, often observed in IBS, can lead to increased permeability (leaky gut), allowing toxins, bacteria, and other substances to cross into systemic circulation and potentially exacerbate symptoms.

[0016] • Modulation of Neurotransmitter Production: The gut microbiota, including butyrate- producing bacteria, play a role in the production of neurotransmitters such as serotonin and gamma-aminobutyric acid (GABA), which are involved in the regulation of gut function and mood. Altered neurotransmitter levels are associated with IBS symptoms, and butyrate may influence these levels indirectly through its effects on the gut microbiota.

[0017] In humans, increased colonic production of the SCFA propionate acutely reduces energy intake. Further, SCFAs can act as a natural appetite suppressant. SCFAs are known to have a beneficial effect on appetite regulation via activating GPCR (FFAR2 and 3) and the release of satiety hormones. “The gut microbiota in obesity and weight management: microbes as friends o / ' foe?”Van Hui M, Cani PD., Nat Rev Endocrinol. 2023 May; 19(5):258-271 ; “Increased colonic propionate reduces anticipatory reward responses in the human striatum to high-energy foods” Byrne CS, Chambers ES, Alhabeeb H, Chhina N, Morrison DJ, Preston T, Tedford C, Fitzpatrick J, Irani C, Busza A, Garcia-Perez I, Fountana S, Holmes E, Goldstone AP, Frost GS., Am J Clin Nutr. 2016 Jul; 104(1):5-14; “Effects of targeted delivery of propionate to the human colon on appetite regulation, body weight maintenance and adiposity in overweight adults” Chambers ES, Viardot A, PsichasA, Morrison DJ, Murphy KG, Zac-Varghese SE, MacDougall K, Preston T, Tedford C, Finlayson GS, Blundell JE, Bell JD, Thomas EL, Mt-lsa S, Ashby D, Gibson GR, Kolida S, Dhillo WS, Bloom SR, Morley W, Clegg S, Frost G, Gut. 2015 Nov; 64(11): 1744-54. “Control of appetite and energy intake by SCFA: what are the potential underlying mechanisms?” Chambers ES, Morrison DJ, Frost G., Proc Nutr Soc. 2015 Aug; 74(3):328-36; “The short chain fatty acid propionate stimulates GLP-1 and PYY secretion via free fatty acid receptor 2 in rodents” Psichas A, Sleeth ML, Murphy KG, Brooks L, Bewick GA, Hanyaloglu AC, Ghatei MA, Bloom SR, Frost G, Int J Obes (Lond). 2015 Mar; 39(3):424-9.

[0018] There is a causal association between the gut microbiota and the development of low-grade inflammation and insulin resistance. Some constituents of gram-negative bacteria, such as lipopolysaccharides (LPS), were the key factors triggering the onset of low-grade inflammation and insulin resistance, i.e., metabolic endotoxaemia. Disruption of the gut barrier leads to metabolic endotoxaemia and impaired production of circulating gut hormones.

[0019] Butyrate promotes colonocyte function, decreasing inflammation, maintaining the gut barrier, and promoting a healthy microbiome. “Gut microbiome, endocrine control of gut barrier function and metabolic diseases" Regnier M, Van Hui M, Knauf C, Cani PD, J Endocrinol. 2021 Feb; 248(2):R67-R82; “Butyrate's role in human health and the current progress towards its clinical application to treat gastrointestinal disease” Hodgkinson K, El Abbar F, Dobranowski P, Manoogian J, Butcher J, Figeys D, Mack D, Stintzi A, Clin Nutr. 2023 Feb; 42(2):61-75.

[0020] Further, there is a known correlation between low grade gut inflammation and IBS and studies have shown that inflammation of the intestinal mucosa and nerves causes the altered Gl dysfunction seen in IBS (^‘Low-grade inflammation plays a pivotal role in gastrointestinal dysfunction in irritable bowel syndrome" H. Akiho, E. Ihara, K. Nakamura, World J Gastrointest Pathophysiol. 2010 Aug 15; 1 (3): 97-105).

[0021] Given its multifaceted role in gut health, butyrate supplementation or interventions aimed at increasing SCFA-producing bacteria in the gut have been proposed as potential therapeutic strategies for IBS and overweight and / or obesity.

[0022] Compounds of the prior art used to influence gut microbiota have several disadvantages such as the requirement of a high dose which limits application formats and high water activity which limits combination with probiotics. In contrast, carotenoids offer several advantages such as low dose, synergistic effect with classical carbohydrate prebiotics and fibres that are normally ingested as food. Moreover, they have low water activity which allows combination with other probiotics.

[0023] Carotenoids have garnered significant attention in the functional food industry owing to their potential health benefits. Originally sourced from plants, where they serve as accessory pigments during photosynthesis, carotenoids are lipophilic antioxidants associated with a reduced risk of various chronic diseases, including cardiovascular disease and macular degeneration. Although some bacteria and fungi also produce carotenoids for protection against UV damage and oxidative stress, humans and other animals rely on dietary intake due to their inability to synthesize these compounds. The primary recognized function of several carotenoids in humans is their provitamin A activity. Beyond this, carotenoids exhibit various biological functions, such as antioxidant properties, inhibition of lipid peroxidation, and tumor-suppressive activity. Lutein and zeaxanthin can help protect the eyes from harmful high-energy light waves like ultraviolet rays in sunlight. Studies suggest that a high level of both in eye tissue is linked with better vision, especially in dim light or where glare is a problem. With over 600 naturally occurring carotenoids identified, further exploration of their multifaceted biological functions presents promising avenues for food supplementation and health promotion. However, carotenoids within the prebiotic space are under-investigated in the prior art. In the prior art, there is only initial evidence on the impact of carotenoids on the gut microbiota.

[0024] WO 2017 / 220708 A1 discloses carotenoids for use in restoring and / or maintaining a health- beneficial gut microbial composition in a subject. This effect is achieved by a specific microbial carotenoid compound resulting in an increased butyrate production by the gut microbiota. In particular, said microbial carotenoid compound modulates the gut microbial composition in favour of butyrate-producing bacteria. Data indicates that the specific bacterial carotenoids can prevent or decrease gastro-intestinal symptoms which are also typically described for IBS.

[0025] US 2019 / 0240168 A1 relates to a method for increasing Akkermansia in intestinal bacterial flora by ingesting the carotenoid astaxanthin. It is reported that Akkermansia muciniphila enterobacteria are associated with obesity and a disease associated therewith. Thus, increase of Akkermansia (muciniphila) in the intestinal bacterial flora allows for, for example, ameliorating a symptom of irritable bowel syndrome (IBS) and ameliorating or preventing obesity.

[0026] In “Prebiotic Effect of Lycopene and Dark Chocolate on Gut Microbiome with Systemic Changes in Liver Metabolism, Skeletal Muscles and Skin in Moderately Obese Persons", Petyaev et al, BioMed Research International Volume 2019, Article ID 4625279, the effect of lycopene and dark chocolate on the gut microbiota in subjects with moderate obesity is investigated. It is reported that continuous administration of a lycopene, especially formulated for effective bioavailability, for 4 weeks resulted in changes in the gut microbiota profile. The authors of the study observed a significant decrease in the abundance of Bacteroidetes and assume that this could be explained either by direct action of this carotenoid, or its indirect activity via stimulation of some species of Bifidobacteria, or a combination of both factors.

[0027] “Carotenoid supplementation and retinoic acid in immunoglobulin A regulation of the gut microbiota dysbiosis", Lyu et al, Experimental Biology and Medicine 2018; 243: 613-620 is a review article focusing on the effect of carotenoid supplementation on the gut microbiota dysbiosis. The authors conclude that carotenoids may contribute to the gut immune homeostasis by directly regulating IgA production, thereby preventing of and / or delaying the development of dysbiosis, but that future investigations are warranted to elucidate the precise mechanisms by which carotenoids are preventive / protective in the gut dysbiosis. The publication also mentions the correlation between SCFAs and disorders, such as IBD (inflammatory bowel disease) and obesity. In particular, it reports that the SCFA butyrate could be used as an energy source for intestinal epithelial cells and microbiomes, which helps protect the intestinal epithelial barrier integrity and make them resistant to potential pathogens. Similar to IBD, a specific individual microbiota signature seems to be related to the development of obesity.

[0028] “Carotenoids and Their Health Benefits as Derived via Their Interactions with Gut Microbiota" , Eroglu et al, Advances in Nutrition 14 (2023) 238-255, is a review article that focuses on the gap of carotenoid research and the interactions of carotenoids with gut microbiota in the colon. Due to limited absorption in the small intestine, the majority of carotenoids are passed on to the colon, where they may act via different mechanisms. The authors conclude that there is mounting evidence that carotenoids can contribute to colonic health, but that more studies on the relationship of carotenoids to the gut microbiota and colon-related health are needed.

[0029] According to the prior art, it is assumed that increasing butyrate-producing bacteria in the gut could be a potential therapeutic strategy for treatment of IBS, but there remains a need for the development of ways to improve the gut microbiota of IBS patients.

[0030] Further, there is a known correlation between increased SCFAs levels in the gut and appetite regulation, but there remains a need for the development of ways to improve the gut microbiota of overweight subjects.

[0031] SUMMARY OF THE INVENTION

[0032] The present invention provides a combination of at least one carotenoid and inulin for use in affecting the gut microbiota in a subject suffering from IBS. Further, the present invention provides a combination of at least one carotenoid and inulin for use in affecting the gut microbiota in a subject suffering from overweight or obesity. Preferably, the combination is for use as a medicament, a dietary supplement, or a food supplement. The present invention further provides a composition comprising inulin and at least one carotenoid. Preferred embodiments are described in the description and in the subclaims.

[0033] More specifically, the present invention relates to the modulation of the gut microbiota of IBS or overweight subjects by carotenoids, i.e., the effects on its composition and metabolic activity. Modulating the gut microbiota composition preferably means that the proportion of SCFA- producing bacteria is increased. In vitro fermentation investigations surprisingly showed that the combination of inulin with carotenoids results in increased production of SCFAs in comparison to inulin alone or carotenoids alone when simulating the colonic fermentation of carotenoids by the gut microbiota from IBS or overweight human adults.

[0034] Preferably, the present invention provides a combination of at least one carotenoid and inulin for use in increasing the amount of SCFAs in the gut of a human IBS or overweight subject in comparison to the administration / consumption of inulin alone and the carotenoid alone.

[0035] BRIEF DESCRIPTION OF THE FIGURES

[0036] Figure 1 shows the results of a kinetic, ex vivo study, simulating the colonic fermentation of carotenoids, incubated with inulin, by the gut microbiota derived from six IBS human adults. The amount of total SCFA production (nM) is shown for inulin alone (IN+) and a combination of inulin with different types and amounts of carotenoids (BC_L+ to ZE_H+). The values measured for each subject (♦, •, •, H, 0, +) are plotted graphically and the average values across for the six individual treatments are indicated at the bottom of the figure. Statistical differences between NSC (No-substrate control; blank) and the individual treatments are visualized via * (0.1 < p adjusted < 0.2), ** (0.05 < p adjusted < 0.1) or *** (p adjusted < 0.05). Significant differences between treatment and IN+ are indicated using $ / $$ / $$$. The rank of the average values per treatment are indicated at the bottom of the figure, with the lowest and highest average being highlighted.

[0037] Figure 2 shows the results of a kinetic, ex vivo study, simulating the colonic fermentation of carotenoids, incubated with inulin, by the gut microbiota derived from six IBS human adults. The amount of acetate production (nM) is shown for inulin alone (IN+) and a combination of inulin with different types and amounts of carotenoids (BC_L+ to ZE_H+). The values measured for each subject (♦, •, •, H, , +) are plotted graphically and the average values across for the six individual treatments are indicated at the bottom of the figure. Statistical differences between NSC and the individual treatments are visualized via * (0.1 < p adjusted < 0.2), ** (0.05 < p adjusted < 0.1) or *** (p adjusted < 0.05). Significant differences between treatment and IN+ are indicated using $ / $$ / $$$. The rank of the average values per treatment are indicated at the bottom of the figure, with the lowest and highest average being highlighted. Figure 3 shows the results of a kinetic, ex vivo study, simulating the colonic fermentation of carotenoids, incubated with inulin, by the gut microbiota derived from six IBS human adults. The amount of propionate production (nM) is shown for inulin alone (IN+) and a combination of inulin with different types and amounts of carotenoids (BC_L+ to ZE_H+). The values measured for each subject (♦, •, •, H, 0, +) are plotted graphically and the average values across for the six individual treatments are indicated at the bottom of the figure. Statistical differences between NSC and the individual treatments are visualized via * (0.1 < p adjusted < 0.2), ** (0.05 < p adjusted < 0.1) or *** (p adjusted < 0.05). Significant differences between treatment and IN+ are indicated using $ / $$ / $$$. The rank of the average values per treatment are indicated at the bottom of the figure, with the lowest and highest average being highlighted.

[0038] Figure 4 shows the results of a kinetic, ex vivo study, simulating the colonic fermentation of carotenoids, incubated with inulin, by the gut microbiota derived from six IBS human adults. The amount of butyrate production (nM) is shown for inulin alone (IN+) and a combination of inulin with different types and amounts of carotenoids (BC_L+ to ZE_H+). The values measured for each subject (♦, •, •, H, , +) are plotted graphically and the average values across for the six individual treatments are indicated at the bottom of the figure. Statistical differences between NSC and the individual treatments are visualized via * (0.1 < p adjusted < 0.2), ** (0.05 < p adjusted < 0.1) or *** (p adjusted < 0.05). Significant differences between treatment and IN+ are indicated using $ / $$ / $$$. The rank of the average values per treatment are indicated at the bottom of the figure, with the lowest and highest average being highlighted.

[0039] Figure 5 shows the results of a kinetic, ex vivo study, simulating the colonic fermentation of carotenoids, incubated with inulin, by the gut microbiota derived from six overweight (OW) human adults. The amount of propionate production (nM) is shown for inulin alone (IN+) and a combination of inulin with different types and amounts of carotenoids (BC_L+ to ZE_H+). The values measured for each subject (▼, •, ♦, ■, ▲ , o) are plotted graphically and the average values across for the six individual treatments are indicated at the bottom of the figure.

[0040] Statistical differences between NSC (No-substrate control; blank) and the individual treatments are visualized via * (0.1 < p adjusted < 0.2), ** (0.05 < p adjusted < 0.1) or *** (p adjusted < 0.05). Significant differences between treatment and IN+ are indicated using $ / $$ / $$$. The rank of the average values per treatment are indicated at the bottom of the figure, with the lowest and highest average being highlighted.

[0041] Figure 6 shows the results of a kinetic, ex vivo study, simulating the colonic fermentation of carotenoids, incubated with inulin, by the gut microbiota derived from six overweight (OW) human adults. The amount of butyrate production (nM) is shown for inulin alone (IN+) and a combination of inulin with different types and amounts of carotenoids (BC_L+ to ZE_H+). The values measured for each subject (▼, •, ♦, ■, ▲ , o) are plotted graphically and the average values across for the six individual treatments are indicated at the bottom of the figure. Statistical differences between NSC and the individual treatments are visualized via * (0.1 < p adjusted < 0.2), ** (0.05 < p adjusted < 0.1) or *** (p adjusted < 0.05). Significant differences between treatment and IN+ are indicated using $ / $$ / $$$. The rank of the average values per treatment are indicated at the bottom of the figure, with the lowest and highest average being highlighted.

[0042] Figure 7 shows the results of a kinetic, ex vivo study, simulating the colonic fermentation of carotenoids, incubated with inulin, by the gut microbiota derived from six overweight (OW) human adults. The amount of acetate production (nM) is shown for inulin alone (IN+) and a combination of inulin with different types and amounts of carotenoids (BC_L+ to ZE_H+). The values measured for each subject (▼, •, ♦, ■, ▲ , o) are plotted graphically and the average values across for the six individual treatments are indicated at the bottom of the figure. Statistical differences between NSC and the individual treatments are visualized via * (0.1 < p adjusted < 0.2), ** (0.05 < p adjusted < 0.1) or *** (p adjusted < 0.05). Significant differences between treatment and IN+ are indicated using $ / $$ / $$$. The rank of the average values per treatment are indicated at the bottom of the figure, with the lowest and highest average being highlighted.

[0043] Figure 8 shows the results of a kinetic, ex vivo study, simulating the colonic fermentation of carotenoids, incubated with inulin, by the gut microbiota derived from six overweight (OW) human adults. The amount of total SCFA production (nM) is shown for inulin alone (IN+) and a combination of inulin with different types and amounts of carotenoids (BC_L+ to ZE_H+). The values measured for each subject (▼, •, ♦, ■, ▲ , o) are plotted graphically and the average values across for the six individual treatments are indicated at the bottom of the figure. Statistical differences between NSC and the individual treatments are visualized via * (0.1 < p adjusted < 0.2), ** (0.05 < p adjusted < 0.1) or *** (p adjusted < 0.05). Significant differences between treatment and IN+ are indicated using $ / $$ / $$$. The rank of the average values per treatment are indicated at the bottom of the figure, with the lowest and highest average being highlighted.

[0044] DETAILED DESCRIPTION OF THE INVENTION

[0045] The present invention relates to a combination of at least one carotenoid and inulin for use in affecting the gut microbiota in a subject suffering from IBS.

[0046] Further, the present invention relates to a combination of at least one carotenoid and inulin for use in affecting the gut microbiota in a subject suffering from overweight or obesity.

[0047] Preferably, the combination is for use as a medicament, a dietary supplement, or a food supplement. The present invention further relates to the non-medical use of the inventive combination as a dietary supplement or a food supplement, preferably for overweight subjects. It is understood that any reference to “combination for use as a dietary supplement” and “combination for use as a food supplement” in this application encompasses the non-medical use thereof, i.e. “use of the combination as a dietary supplement” and “use of the combination as a food supplement”.

[0048] The inventive combination can comprise either a single kind of carotenoid or a mixture of two or more carotenoids. The kind of carotenoid is not particularly limited. Any suitable kind of carotenoid (carotenes and xanthophylls) can be used. Preferably, the carotenoid is selected from the group consisting of: a-carotene (alpha), p-carotene (beta), y-carotene (gamma), 6- carotene (delta), s-carotene (epsilon), ^-carotene (zeta), i , ip-carotene (lycopene), lutein (including its esterified version), zeaxanthin (including its esterified version), astaxanthin, canthaxanthin, and p-zeacarotene; isomers thereof, derivatives thereof, and combinations thereof. Particularly preferred are p-carotene, i , ip-carotene (lycopene), lutein, zeaxanthin, and combinations thereof.

[0049] Preferred is a combination of inulin with a single carotenoid. Particularly preferred are a combination of inulin with p-carotene, a combination of inulin with lycopene, a combination of inulin with lutein, and a combination of inulin with zeaxanthin.

[0050] As is shown in the examples below, a combination of inulin and at least one carotenoid positively influences the gut microbiota of IBS subjects.

[0051] According to the Rome IV Consensus Criteria of the American Gastroenterological Association (B. E. Lacy and N. K. Patel, “Rome Criteria and a Diagnostic Approach to Irritable Bowel Syndrome", J Clin Med. 2017 Nov; 6(11): 99) and other medical societies, irritable bowel syndrome can be diagnosed if the following criteria are met: Recurrent abdominal pain, on average at least once a week within the past three months, associated with two of the following three factors. These criteria should be met for the last three months, while the onset of symptoms should be at least six months ago.

[0052] • Defecation

[0053] • Change in stool frequency

[0054] • Change in stool consistency

[0055] Preferably, the subject is a human subject in which IBS has been diagnosed according to the Rome IV Consensus Criteria.

[0056] As further shown in the examples below, a combination of inulin and at least one carotenoid positively influences the gut microbiota of overweight subjects. Accordingly, the subject may be a subject suffering from overweight or obesity. Preferably, the subject is suffering from overweight, i.e., the subject has a body mass index (BMI) of 25 < BMI < 30. The subject can be any kind of mammal, including humans, pets (e.g., dogs) and livestock. Preferably, the subject is a human, more preferably a human adult.

[0057] In the context of the present invention, affecting the gut microbiota includes any effects on the composition of the gut microbiota and the metabolic activity of the gut microbiota.

[0058] Preferably, in a subject with IBS, it involves that the development of low grade gut inflammation is prevented, reduced or slowed down, and / or that gut motility is beneficially regulated, and / or that the gut barrier function is restored or maintained and / or that the neurotransmitter production in the gut is beneficially modulated. These effects are particularly beneficial for subjects with IBS. Preferably, the prevention, reduction or slowing down of development of low grade gut inflammation is in comparison to a non-administered subject, in comparison to a subject administered carotenoids alone, or in comparison to a subject administered inulin alone. More preferably, the prevention, reduction or slowing down is both in comparison to a subject administered carotenoids alone and in comparison to a subject administered inulin alone (synergistic effect). Preferably, by combining inulin and at least one carotenoid, the known modulating effect of inulin on the gut microbiota is increased.

[0059] Preferably, in a subject with overweight or obesity, it involves that the development of low-grade gut inflammation and / or the development of insulin resistance is prevented, reduced or slowed down. These effects are particularly beneficial for subjects with overweight or obesity.

[0060] Preferably, it involves effecting appetite regulation in the subject, which is also a particularly beneficial effect for subjects with overweight or obesity. Preferably, the prevention, reduction or slowing down of development of low-grade gut inflammation and / or the development of insulin resistance is in comparison to a non-administered subject, in comparison to a subject administered carotenoids alone, or in comparison to a subject administered inulin alone. More preferably, the prevention, reduction or slowing down is both in comparison to a subject administered carotenoids alone and in comparison to a subject administered inulin alone (synergistic effect). Preferably, by combining inulin and at least one carotenoid, the known modulating effect of inulin on the gut microbiota is increased.

[0061] Preferably, affecting the gut microbiota involves increasing the amount of SCFAs in the gut of the subject. SCFAs (Short-Chain Fatty Acids) are fatty acids with fewer than six carbon atoms in their chain length. Preferably, the amount of SCFAs is increased in comparison to a nonadministered subject, in comparison to a subject administered carotenoids alone, or in comparison to a subject administered inulin alone. More preferably, the amount of SCFAs is increased both in comparison to a subject administered carotenoids alone and in comparison to a subject administered inulin alone (synergistic effect). Preferably, by combining inulin and at least one carotenoid, the known modulating effect of inulin on the gut microbiota is increased. In the examples, it is shown how an increase of SCFAs in comparison to subject administered carotenoids alone and in comparison to a subject administered inulin alone can be determined. Preferably, the SCFAs are selected from the group consisting of acetate, propionate, and butyrate, and combinations thereof. More preferably, the SCFAs are selected from the group consisting of propionate and butyrate.

[0062] In subjects with IBS, most preferably, the SCFAs comprise butyrate.

[0063] Preferably, the combination is for simultaneous administration or consumption. Inulin and the at least one carotenoid can be administered or consumed in a fixed combination or a free combination. However, since both inulin and carotenoids are preferably administered / consumed in oral form, a fixed combination is preferred due to the ease of handling. Preferably, the combination of inulin and the at least one carotenoid is a fixed combination, i.e. preferably, the combination of inulin and the at least one carotenoid is a composition comprising inulin and the at least one carotenoid.

[0064] The combination can be administered / consumed in any suitable form known to the skilled person. Preferably, the combination is administered / consumed in oral form. Preferably, the combination is administered in the form of a tablet, a capsule, a sachet of dried powder or as ingredient of functional food. Examples for functional food include yoghurt or smoothies.

[0065] In addition to inulin and at least one carotenoid, the inventive combination can comprise vitamins, further prebiotics, suitable excipients, and combinations thereof. Examples for vitamins are all lipid and water soluble vitamins. Examples for prebiotics are inulin-type fructans (ITF), GOS (Galactooligosaccharides); XOS (Xylooligosaccharides), pectin, beta glucan, and HMOs (human milk oligosaccharides). Examples for excipients are known to the skilled person and include binders (e.g., hydroxypropyl methylcellulose, microcrystalline cellulose, starches, and gelatine); fillers (e.g., lactose, mannitol, sorbitol, and dicalcium phosphate); disintegrants (e.g., croscarmellose sodium, crospovidone, and sodium starch glycolate); glidants (e.g., colloidal silicon dioxide and talc); lubricants (e.g., magnesium stearate, stearic acid, and sodium lauryl sulphate); colorants; flavouring agents; and coating agents (e.g., cellulose derivatives, shellac, and polymethacrylates).

[0066] More specifically, the present inventors have found that a combination of inulin and different carotenoids has a dose-specific effect on the gut microbiota of IBS subjects and overweight subjects. Thus, the present invention relates to a composition comprising inulin and at least one carotenoid in an effective amount, wherein the carotenoid is selected from the group consisting of: a-carotene, p-carotene, y-carotene, 6-carotene, s-carotene, ^-carotene, i , ip-carotene (lycopene), , lutein (including its esterified version), zeaxanthin (including its esterified version), astaxanthin, canthaxanthin, and p-zeacarotene; isomers thereof, derivatives thereof, and combinations thereof. Preferably, the effective amount results in a synergistic effect as described herein. The composition of the present invention comprises inulin in an amount of 0.1 to 40 g, preferably 0.3 to 40 g, more preferably 0.5 to 40 g, more preferably 1 to 40 g, more preferably 1 to 30 g, more preferably 2 to 25 g, more preferably 2 to 20 g, most preferably 3 to 15 g. For instance, a suitable amount of inulin is 2 to 15 g, 3 to 10 g or 5 g, as in the examples of the present application The composition of the present invention comprises at least one carotenoid in an amount of 1 to 70 mg, preferably 2 to 65 mg, more preferably 2 to 60 mg. The preferred amount of carotenoid typically depends on the kind of carotenoid used. Exemplarily ranges for p-carotene, lutein, lycopene and zeaxanthin are described below. Preferably, the composition is used as a dietary supplement or a food supplement.

[0067] The combination of the present invention preferably comprises inulin in an amount of 0.1 to 40 g and at least one carotenoid in an amount of 1 to 70 mg, more preferably inulin in an amount of 0.3 to 40 g and at least one carotenoid in an amount of 1 to 70 mg, more preferably inulin in an amount of 0.5 to 40 g and at least one carotenoid in an amount of 1 to 70 mg, more preferably inulin in an amount of 1 to 40 g and at least one carotenoid in an amount of 1 to 70 mg, more preferably inulin in an amount of 1 to 30 g and at least one carotenoid in an amount of 2 to 65 mg, more preferably inulin in an amount of 2 to 20 g and at least one carotenoid in an amount of 2 to 60 mg, most preferably inulin in an amount of 3 to 15 g and at least one carotenoid in an amount of 2 to 60 mg.

[0068] When p-carotene is comprised as carotenoid, it is preferably comprised in an amount of 2 to 25 mg, more preferable 2 to 21 mg, more preferable 3 to 15 mg, more preferable 3 to 10 mg, most preferable 3.5 to 7 mg.

[0069] When lutein is comprised as carotenoid, it is preferably comprised in an amount of 5 to 70 mg, more preferable 10 to 70 mg, more preferable 10 to 60 mg, more preferable 20 to 60 mg, most preferable 40 to 60 mg.

[0070] When lycopene is comprised as carotenoid, it is preferably comprised in an amount of 5 to 50 mg, more preferable 10 to 50 mg, more preferable 15 to 50 mg, more preferable 15 to 45 mg, most preferable 25 to 45 mg.

[0071] When zeaxanthin is comprised as carotenoid, it is preferably comprised in an amount of 1 to 10 mg, more preferable 2 to 9 mg, more preferable 2 to 8 mg, more preferable 2 to 7 mg, most preferable 2 to 6 mg.

[0072] When different kinds of carotenoids are present in the combination, the amounts of each kind of carotenoid can be adjusted accordingly, as will be evident to the skilled person. For instance, a combination comprising inulin, p-carotene and lutein, preferably comprises 0.1 to 40 g inulin, 1 to 12.5 mg p-carotene (half of the amount indicated above for p-carotene alone) and 2.5 to 35 mg lutein (half of the amount indicated above for lutein alone). When the combination is used as a medicament, a dietary supplement, or a food supplement, the above amounts correspond to the administered daily dose. Accordingly, when the combination is not administered as a single dose but as separate doses in the course of the day, the above amounts can be divided into several smaller doses, such that the total daily dose corresponds to the above amounts. Preferably, the inventive combination is for a use comprising administering to the subject inulin in an effective amount and at least one carotenoid in an effective amount. Preferably, the effective amount results in a synergistic effect as described herein. The combination of the present invention is preferably for a use comprising administering the combination to the subject in a combination of inulin in an amount of 0.1 to 40 g / day, more preferably 0.3 to 40 g / day, more preferably 0.5 to 40 g / day, more preferably 1 to 40 g / day, more preferably 1 to 30 g / day, more preferably 2 to 25 g / day, more preferably 2 to 20 g / day, most preferably 3 to 15 g / day; and the at least one carotenoid in an amount of 1 to 70 mg / day, more preferably 2 to 65 mg / day, more preferably 2 to 60 mg / day. For instance, a suitable dose of inulin is 2 to 15 g / day or 3 to 10 g / day or 5 g / day, as in the examples of the present application. A daily dose of 0.1 to 40 g inulin typically corresponds to a dose of around 1 to 600 mg / kg body weight per day.

[0073] There are no specific limitations regarding the duration of the administration / consumption of the inventive combination. However, to enhance the beneficial effects thereof, it is advantageous to administer / consume it in the above-described daily doses for a period of at least 2 weeks, preferably at least 3 weeks, more preferably at least 4 weeks, most preferably at least 6 weeks. Further, there is no upper limit for the duration of administration / consumption and the inventive combination can be administered / consumed for several months or several years.

[0074] Preferably, the combination of the present invention comprises inulin and carotenoids in the following amounts:

[0075] The following preferred embodiments apply to IBS subjects:

[0076] • Preferably, the inventive use involves increasing the amount of total SCFA in the gut of the subject, and the combination comprises inulin in an amount of 3 - 15 g / day, and p-carotene in an amount of 3.5 - 21 mg / day or lycopene in an amount of 25 - 45 mg / day, more preferable inulin in an amount of 3 - 15 g / day and lycopene in an amount of 25 - 45 mg / day, more preferable 35 - 45 mg / day.

[0077] • Preferably, the inventive use involves increasing the amount of acetate in the gut of the subject, and the combination comprises inulin in an amount of 3 - 15 g / day, and p-carotene in an amount of 3 - 5 mg / day or lycopene in an amount of 25 - 45 mg / day; more preferable inulin in an amount of 3 - 15 g / day and lycopene in an amount of 25 - 45 mg / day, more preferable 35 - 45 mg / day.

[0078] • Preferably, the inventive use involves increasing the amount of propionate in the gut of the subject, and the combination comprises inulin in an amount of 3 - 15 g / day, and p-carotene in an amount of 3.5 - 21 mg / day or lycopene in an amount of 25 - 45 mg / day; more preferable inulin in an amount of 3 - 15 g / day and lycopene in an amount of 25 - 45 mg / day, more preferable 35 - 45 mg / day.

[0079] • Preferably, the inventive use involves increasing the amount of butyrate in the gut of the subject, and the combination comprises inulin in an amount of 3 - 15 g / day, and p-carotene in an amount of 3.5 - 21 mg / day or lutein in an amount of 10 - 60 mg / day or lycopene in an amount of 10 - 20 mg / day or zeaxanthin in an amount of 2 - 6 mg / day or combinations thereof.

[0080] • Preferably, the inventive use involves increasing the amount of total SCFA, acetate and propionate in the gut of the subject, and the combination comprises inulin in an amount of 3

[0081] - 15 g / day and lycopene in an amount of 25 - 45 mg / day, more preferable 35 - 45 mg / day.

[0082] The following preferred embodiments apply to overweight subjects:

[0083] • Preferably, the inventive use involves increasing the amount of propionate in the gut of the subject, and the combination comprises inulin in an amount of 3 - 15 g / day, and p-carotene in an amount of 3.5 - 21 mg / day or lycopene in an amount of 25 - 45 mg / day; more preferable inulin in an amount of 3 - 15 g / day g / day and lycopene in an amount of 25 - 45 mg / day, more preferable 35 - 45 mg / day.

[0084] • Preferably, the inventive use involves increasing the amount of butyrate in the gut of the subject, and the combination comprises inulin in an amount of 3 - 15 g / day, and p-carotene in an amount of 3.5 - 21 mg / day or lutein in an amount of 10 - 60 mg / day or lycopene in an amount of 10 - 20 mg / day or zeaxanthin in an amount of 2 - 6 mg / day or combinations thereof; more preferable inulin in an amount of 3 - 15 g / day and lutein in an amount of 10 - 60 mg / day, more preferable 40 - 60 mg / day.

[0085] • Preferably, the inventive use involves increasing the amount of acetate in the gut of the subject, and the combination comprises inulin in an amount of 3 - 15 g / day, and p-carotene in an amount of 2 - 5 mg / day or lycopene in an amount of 25 - 45 mg / day; more preferable inulin in an amount of 3 - 15 g / day and lycopene in an amount of 25 - 45 mg / day, more preferable 35 - 45 mg / day.

[0086] • Preferably, the inventive use involves increasing the amount of total SCFA in the gut of the subject, and the combination comprises inulin in an amount of 3 - 15 g / day, and p-carotene in an amount of 3 - 10 mg / day or lycopene in an amount of 25 - 45 mg / day; more preferable inulin in an amount of 3 - 15 g / day and lycopene in an amount of 25 - 45 mg / day, more preferable 35 - 45 mg / day.

[0087] • Preferably, the inventive use involves increasing the amount of propionate, acetate and total SCFA in the gut of the subject, and the combination comprises inulin in an amount of 3

[0088] - 15 g / day and lycopene in an amount of 25 - 45 mg / day, more preferable 35 - 45 mg / day. Definitions

[0089] The term “affecting the gut microbiota" refers to any influence or change on the microorganisms residing in the gastrointestinal tract of the subject. It includes any effects on the composition of the gut microbiota and the metabolic activity of the gut microbiota such as an increase in the production of SCFA.

[0090] “ANOVA" stands for Analysis of Variance. It is a statistical method used to analyze the differences among group means in a sample. ANOVA assesses whether the means of two or more groups are statistically significantly different from each other.

[0091] The term “administration" is used in connection with a medicament. It describes the process of giving or applying the inventive combination, typically following specific guidelines or instructions.

[0092] The term “consumption" is used in connection with a dietary supplement or food supplement. It describes the process of ingesting the inventive combination.

[0093] In the present application, unless not specified otherwise or obvious from the context, features described in relation to the term “administration” encompass the corresponding features in relation to the term “consumption” and vice versa. For instance, if it is described that the inventive combination is administered in a specific amount, this encompasses the statement that the inventive combination is consumed in the specific amount.

[0094] “Carotenoids" are a class of chemical compounds that are produced by plants and algae, as well as several bacteria, archaea, and fungi. They are organic pigments consisting of two subgroups, i.e. carotenes and xanthophylls.

[0095] The term carotenoid is certainly understood to include a-carotene, p-carotene, y-carotene, 5- carotene, s-carotene, ^-carotene, i , ip-carotene (lycopene), lutein, including its esterified form, zeaxanthin, including its esterified form, astaxanthin, canthaxanthin, and p-zeacarotene; isomers thereof, derivatives thereof, and combinations thereof.

[0096] Preferred examples of carotenoids include p-carotene, lutein, lycopene and zeaxanthin, isomers thereof, derivatives thereof, and combinations thereof.

[0097] Preferably, isomers of the aforementioned carotenoids include E-isomers, such as all-E isomers, and Z-isomers. For instance, preferred isomers of lutein are zeaxanthin and meso- zeaxanthin.

[0098] Preferably, derivatives of the aforementioned carotenoids include esters and chemical derivatives such as butyrate, propionate. For instance, a preferred derivative of lutein is its esterified form, because lutein typically occurs in esterified form in nature, e.g., in tagetes, and is then saponified to free lutein. Combinations of carotenoids include combinations in any kind of ratios, preferably those ratios that result from processing. For instance, as result of their joined extraction processes from plants, a typical combination of carotenoids is a combination of lutein and zeaxanthin, wherein a weight ratio of lutein / zeaxanthin is in the range of 0.1 / 10 to 10 / 0.1.

[0099] “ / 3-Carotene", also called p, p-Carotene, is the most common carotenoid in plants and the best- known provitamin A carotenoid. In the body, it can be converted to retinol (vitamin A). p- Carotene is industrially made either by chemical synthesis (also called nature identical p- Carotene), or by extraction or fermentation from biological sources and genetically-engineered microbes (also called natural p-Carotene). Natural p-Carotene usually comprises minor amounts of side products as a result of the extraction and fermentation procedure, e.g. a- Carotene and y-Carotene. In the context of the present application, p-Carotene encompasses natural and nature identical p-Carotene.

[0100] “Lutein", also called (3R,6R,3'R)-p,s-Carotene-3,3'-diol, is a xanthophyll which is found in high quantities in green leafy vegetables and tagetes species.

[0101] “Lycopene", also called i , i -Carotene, is a bright red carotenoid hydrocarbon found in tomatoes and other red fruits and vegetables.

[0102] “Zeaxanthin", also called (3R,3'R)-p,p-Carotene-3,3'-diol, is synthesized in plants and some micro-organisms.

[0103] The term “combination" means that inulin and the at least one carotenoid are in a form allowing their administration / consumption at the same time or at a short interval. In the case of a “fixed combination”, the term “combination of’ can be used interchangeably with the term “composition comprising”.

[0104] A “dietary supplement" is as a product taken by mouth that contains a dietary ingredient, e.g. vitamins, minerals, herbs, amino acids, intended to supplement the diet. The term is defined by the DSHEA (Dietary Supplement Health and Education Act) to mean a product (other than tobacco) intended to supplement the diet that bears or contains one or more dietary ingredients, including a vitamin, a mineral, an herb or other botanical, an amino acid, a dietary substance for use by human to supplement the diet by increasing the total dietary intake, or a concentrate, metabolite, constituent, extract, or combination of any of the aforementioned ingredients. Thus, consumption as a dietary supplement can refer to a non-medical use of the inventive combination.

[0105] The term “effecting appetite regulation" refers to the process of influencing or controlling the body's natural mechanisms that govern hunger, satiety, and food intake. Preferably, it means that the subject’s appetite is controlled, resulting in a decreased energy intake. An “effective amount’ of the inventive combination is the combination of an amount of inulin and an amount of at least one carotenoid that render the desired outcome in the subject the combination is administered to. Preferably, the effective amount results in a synergistic effect as described herein.

[0106] A “fixed combination" is a dosage form comprising both inulin and the at least one carotenoid, e.g. in the form of a tablet or capsule, or as a functional food, e.g. a yoghurt comprising both inulin and the at least one carotenoid.

[0107] A “free combination" is a dosage form allowing to administer / consume inulin and the at least one carotenoid one at a time. For instance, inulin and the at least one carotenoid can be contained in two separate tablets or capsules.

[0108] A “food supplement" is any product intended to supplement the diet with additional nutrients, e.g. prebiotics or vitamins. The term “food supplement” can encompass the term “dietary supplement” but includes also substances not falling under the specific definition of a dietary supplement according to the DSHEA. Preferably, food supplements are contained in functional food. Thus, consumption as a food supplement can refer to a non-medical use of the inventive combination.

[0109] “Functional food" is a kind of food that provides health benefits beyond basic nutrition due to its specific bioactive compounds or ingredients. These foods are often designed or modified to provide additional physiological benefits beyond their basic nutritional value. Examples of functional food include yoghurt, cereals, drinks or smoothies.

[0110] “Inulin" is a heterogeneous collection of fructose polymers. It consists of chain-terminating glucosyl moieties and a repetitive fructosyl moiety, which are linked by [3(2,1) bonds. The degree of polymerization (DP) of standard inulin ranges from 2 to 60. Because of the [3(2,1) linkages, inulin is not digested by enzymes in the human alimentary system.

[0111] “Inulin-type fructans (ITF)" are a group of natural carbohydrates that include fructooligosaccharides (FOS), oligofructose, and inulin.

[0112] “Irritable bowel syndrome" (IBS) as used herein refers to a prevalent disorder primarily affecting the large intestine or colon. Characterized by symptoms such as cramping, abdominal pain, bloating, gas, diarrhea, and constipation, IBS is distinguished from other gastrointestinal conditions like Crohn’s disease or ulcerative colitis by its chronic nature and absence of structural changes in bowel tissue or elevated risk of colorectal cancer. It is commonly diagnosed according to the Rome IV Consensus Criteria.

[0113] The term “low grade gut inflammation" refers to gut inflammation without tissue injury, which is not usually detectable by colonoscopy. It can be detected through biopsies from intestine and blood. This term is used to describe low-grade inflammation occurring in IBS in comparison to the inflammatory disorder IBD or an overt inflammation, as seen for instance in Crohn’s disease.

[0114] The term “restoring or maintaining the gut barrier function" refers to the process of preserving the integrity and effectiveness of the gut barrier, which helps prevent harmful substances from passing into the bloodstream; or to restore it if it is compromised in an IBS patient. Dysfunction of the gut barrier is often observed in IBS and can lead to increased permeability (leaky gut), allowing toxins, bacteria, and other substances to cross into systemic circulation and potentially exacerbate symptoms. Thus, by restoring or maintaining the barrier function, potentially symptoms of IBS can be alleviated. medicament' is a substance or preparation used for the prevention, diagnosis, alleviation, treatment, or cure of disease or to affect the structure or function of the body. A medicament is used to treat an existing disease (in particular IBS or obesity) or to prevent a disease (in particular IBS or obesity). Thus, administration as a medicament refers to a medical use of the inventive combination.

[0115] The term “beneficially modulating the neurotransmitter production in the gut' refers to the process of regulating the synthesis and activity of chemical messengers (neurotransmitters) in the gastrointestinal tract, such that IBS symptoms can be alleviated. The gut microbiota plays a role in the production of neurotransmitters such as serotonin and gamma-aminobutyric acid (GABA), which are involved in the regulation of gut function and mood. Altered neurotransmitter levels are associated with IBS symptoms. Thus, by beneficially modulating neurotransmitter production, potentially symptoms of IBS can be alleviated.

[0116] “Obesity" is a chronic complex disease defined by excessive fat deposits that can impair health (e.g. it can lead to increased risk of type 2 diabetes and heart disease, it can affect bone health and reproduction, and it increases the risk of certain cancers). It is defined by the WHO as a BMI greater than or equal to 30 (for adults). The BMI (body mass index) is calculated as follows: weight (kg) / height2(m2). The skilled person knows how to convert the BMI ranges indicated for adults to the corresponding BMI ranges for children taking the definitions provided by the WHO into account (https: / / www.who.int / news-room / fact-sheets / detail / obesity-and-overweight).

[0117] “Overweight', abbreviated “OW’, is a condition of excessive fat deposits and defined by the WHO as a BMI greater than or equal to 25 (for adults). In the context of the present application, adult human overweight subjects have 25 < BMI < 30.

[0118] “Prebiotics" are compounds contained in or added to food that induce the growth or activity of beneficial microorganisms such as bacteria and fungi in the gut of an animal species or a human. In the gastrointestinal tract, prebiotics can alter the composition of organisms in the gut microbiome. The term “beneficially regulating the gut motility" refers the process of managing and optimizing the movement of the digestive system, i.e., to the movement of food and waste through the digestive tract. By beneficially regulating this motility, potentially symptoms of IBS can be alleviated.

[0119] “SCFAs” (Short-Chain Fatty Acids) are fatty acids with fewer than six carbon atoms in their chain length. They are produced by certain gut bacteria as a result of carbohydrate fermentation of dietary fibres in the colon. Acetate, propionate and butyrate are the primary SCFA products and are produced in an approximate molar ratio of 60:20:20.

[0120] “Simultaneous administration or consumption" in the context of the present application means that inulin and the at least one carotenoid are administered or consumed at the same time or at a short interval. A short interval means any time up to 12 hours.

[0121] EXAMPLES

[0122] The present invention is further illustrated in the following examples, which are provided by way of illustration and not by way of limitation.

[0123] Example: Comparing the ex vivo simulated colonic fermentation of carotenoids, incubated with or without inulin, in human adults with IBS and overweight human adults

[0124] Study set-up and tested compounds.

[0125] A kinetic, ex vivo study was implemented, simulating the colonic fermentation of test products by the gut microbiota derived from human adults with IBS (n = 6) and healthy overweight (OW) human adults (n = 6), respectively. Individual bioreactors were processed in parallel in a bioreactor management device (Cryptobiotix, Ghent, Belgium). Each bioreactor contained 5 ml of nutritional medium-faecal inoculum blend dosed with various test products, sealed individually, before being rendered anaerobic. After preparation, bioreactors were incubated under continuous agitation (140 rpm) at 37°C for 24 h. Upon gas pressure measurement in the headspace, liquid samples were collected for subsequent analysis.

[0126] Test products were carotenoids, incubated with or without inulin (given at the concentration 5 g / l, simulating intake of 5 g per day). List of carotenoids and their doses is given in Table A below. Carotenoids are contained in an amount of 1-60 mg.

[0127] The doses of carotenoids investigated are based on the daily upper limits at which each of the carotenoids are administered / consumed when taken alone (middle doses in Table A). The in vitro system used is designed to allow for 1 :1 extrapolation of effect of doses from in vitro setting to in vivo. Table A. Types and doses of carotenoids.

[0128] The concentration (mM) of SCFAs (acetate, propionate, butyrate) was determined via a gas chromatography-flame ionization detection (GC-FID) approach. GC-FID is an analytical technique that is used to separate and analyze mixtures consisting of volatile components. GC- FID is particularly useful in detecting and quantifying organic compounds, such as fatty acids.

[0129] SCFAs were measured for inulin alone, for each of the carotenoids alone (3 doses), and for combination of each of the carotenoids (3 doses) with inulin.

[0130] Statistics

[0131] For the statistical evaluation of the treatment effects on SCFA production across 6 different donors, a repeated measures ANOVA analysis was performed (based on paired t-testing, thus accounting for fact that values are compared between samples of a given donor). The statistical significance of potential treatment effects was determined via Benjamin-Hochberg post hoc testing. The latter involves that a correction for multiple comparisons was implemented where p- values were adjusted by multiplying them with the total number of comparisons divided by the rank of each original p-value (across all p-values). In this specific case, 30 comparisons were considered (30 carotenoid-related treatments vs IN+). In practice, this means that while the largest obtained p-value remained uncorrected (i.e., multiplied 1), the lowest p-value was multiplied with 30, thus strongly decreasing the chance of type 1 errors (i.e., false positives). In addition, the resulting series of adjusted p-values was rendered non-decreasing (i.e., when an adjusted p-value was higher than any of the subsequent adjusted p-values, its value was equaled to this lowest value). This generates a false discovery rate threshold allowing to estimate and control the chance of type 1 errors (i.e., false positives). Statistical differences were visualized via: • $ (0.1 < p adjusted < 0.2), $$ (0.05 < p adjusted < 0.1) or $$$ (p adjusted < 0.05) for carotenoid-related treatments vs IN+.

[0132] Determination of synergism

[0133] A synergistic effect is displayed by outlining the following measurements:

[0134] • Carotenoid alone (Types and doses of carotenoids, see Table A)

[0135] • Inulin alone (IN)

[0136] • Inulin + carotenoid

[0137] These measurements are used then to calculate the Synergistic effect by adding “carotenoid alone” with “inulin alone” values and comparing it to the measured value of “inulin + carotenoid”. The difference is the value in the Synergistic effect column shown in Tables 1 to 4. The values are given as amount (in mM) of produced SCFA.

[0138] The results in Figures 1 to 4 show measured concentration for each of the SCFAs for inulin alone and combination of carotenoids (3 doses) in the presence of inulin.

[0139] In Tables 1 to 4 data points are provided which display synergistic effect for inulin and carotenoids. In the Tables, the data where there is a synergistic effect present between a carotenoid and inulin, have been extracted. Thus, only combinations with synergistic effect are shown in Tables 1 to 4. A summary of the data is shown in Table 5.

[0140] Example 1-1 : Total SCFA Production

[0141] The results for measuring the concentration (mM) of total SCFA are shown in Table 1 and Figure 1.

[0142] In samples from IBS donors, a synergistic (higher than the sum of the increases of carotenoids only and inulin only) increase of total SCFA production is observed when inulin is combined with different carotenoids. In IBS subjects, inulin combined with lycopene at 45 mg (LY_H) resulted in the highest increase in total SCFA among all treatments. This increase was significantly different to inulin alone. Also p-carotene at 7 mg (BC_M) and nat. p-carotene at 3.5, 7 and 21 mg (NBC_L, NBC_M and NBC_H) led to a statistically significant increase in total SCFA when compared to inulin only (Figure 1). These increases in total SCFA in response to inulin plus carotenoids were synergistic, i.e. higher than the sum of the increases of carotenoids only and inulin only (Table 1). Table 1. Samples in which a synergistic effect for total SCFA production is observed.

[0143] Example 1-2: Acetate Production

[0144] The results for measuring the concentration (mM) of acetate are shown in Table 2 and Figure 2.

[0145] In IBS subjects, inulin combined with lycopene at 45 mg (LY_H) resulted in the highest increase in acetate among all treatments. This increase was significantly different to inulin alone. Also nat. p-carotene at 3.5 mg (NBC_L) led to a statistically significant increase in acetate when compared to inulin only (Figure 2). These increases in acetate in response to inulin plus carotenoids were synergistic, i.e. higher than the sum of the increases of carotenoids only and inulin only (Table 2).

[0146] Table 2: Samples in which a synergistic effect for acetate production is observed.

[0147] Example 1-3: Propionate Production

[0148] The results for measuring the concentration (mM) of propionate are shown in Table 3 and Figure 3.

[0149] In IBS subjects, inulin combined with lycopene at 45 mg (LY_H) resulted in the highest increase in propionate among all treatments. This increase was significantly different to inulin alone. Also P-carotene at 7 mg (BC_M), and nat. p-carotene at 3.5, 7 and 21 mg (NBC_L, NBC_M, NBC_H) led to a statistically significant increase in propionate when compared to inulin only (Figure 3). These increases in propionate in response to inulin plus carotenoids were synergistic, i.e. higher than the sum of the increases of carotenoids only and inulin only (Table 3). Table 3 Samples in which a synergistic effect for propionate production is observed.

[0150] Example 1-4: Butyrate Production

[0151] The results for measuring the concentration (mM) of butyrate are shown in Table 4 and Figure 4. In IBS subjects, inulin combined with zeaxanthin at 2 mg (ZE_M) resulted in the highest increase in butyrate among all treatments. This increase was significantly different to inulin alone. Also all other carotenoids led to a statistically significant increase in butyrate when compared to inulin only except for lycopene at 7.5 (LY_L) and 45 mg (LY_H) and zeaxanthin at 1 mg (ZE_L) (Figure 4). The increase in butyrate in response to the combination of inulin and carotenoids were synergistic, i.e. higher than the sum of the increase of carotenoids only and inulin only (Table 4).

[0152] Table 4: Samples in which a synergistic effect for butyrate production is observed.

[0153] Summary of Examples 1-1 to 1-4

[0154] Table 5 summarized the data on synergistic effect shown in Tables 1 to 4. As shown in Table 5, the combination of inulin with each carotenoid tested shows a synergistic effect for the production of at least one of the SCFAs investigated. In summary, the experimental data show a dose-specific synergistic effect of a combination of inulin and different carotenoids for the production of SCFAs in an ex vivo model for simulated colonic fermentation of carotenoids, incubated with or without inulin, in IBS human adults.

[0155] From the prior art, it is known that there is a relationship between the gut microbiota and IBS symptoms. SCFAs, in particular butyrate, play several important roles in the gut, and are found to be reduced in patients suffering from irritable bowel syndrome (IBS). Therefore, the synergistic effect on SCFA production observed in the examples of the present application indicates that the inventive combination of inulin and at least one carotenoid is suitable for use as a medicament, a dietary supplement, or a food supplement in a subject suffering from IBS.

[0156] Table 5’. Overview of samples for which a synergistic effect present between a carotenoid and inulin is observed.

[0157] +: Synergism observed, see Tables 1 to 4

[0158] ++: Sample with highest synergism observed, see Tables 1 to 4

[0159] No synergism observed, see Tables 1 to 4

[0160] Example 2-1 : Propionate Production

[0161] The results for measuring the concentration (mM) of propionate are shown in Table 6 and Figure 5.

[0162] In samples from overweight donors, a synergistic (higher than the sum of the increases of carotenoids only and inulin only) increase of propionate production is observed when inulin is combined with different carotenoids. The effect is strongest with lycopene at 45 mg (LY_H). Table 6’. Samples in which a synergistic effect for propionate production is observed.

[0163] Example 2-2: Butyrate Production

[0164] The results for measuring the concentration (mM) of butyrate are shown in Table 7 and Figure 6.

[0165] In samples from overweight donors, a synergistic (higher than the sum of the increases of carotenoids only and inulin only) increase of butyrate production is observed when inulin is combined with different carotenoids. The effect is strongest with lutein at 60 mg (LU_H).

[0166] Table 7: Samples in which a synergistic effect for butyrate production is observed. Example 2-3: Acetate Production

[0167] The results for measuring the concentration (mM) of acetate are shown in Table 8 and

[0168] Figure 7.

[0169] In samples from overweight donors, a synergistic (higher than the sum of the increases of carotenoids only and inulin only) increase of acetate production is observed when inulin is combined with different carotenoids. The effect is strongest with lycopene at 45 mg (LY_H). Table 8. Samples in which a synergistic effect for acetate production is observed.

[0170] Example 2-4: Total SCFA Production

[0171] The results for measuring the concentration (mM) of total SCFA are shown in Table 9 and Figure 8.

[0172] In samples from overweight donors, a synergistic (higher than the sum of the increases of carotenoids only and inulin only) increase of total SCFA production is observed when inulin is combined with different carotenoids. The effect is strongest with lycopene at 45 mg (LY_H).

[0173] Table 9’. Samples in which a synergistic effect for total SCFA production is observed.

[0174] Summary of Examples 2-1 to 2-4

[0175] Table 5 summarized the data on synergistic effect shown in Tables 6 to 9. As shown in Table 10, the combination of inulin with each carotenoid tested shows a synergistic effect for the production of at least one of the SCFAs investigated. In summary, the experimental data show a dose-specific synergistic effect of a combination of inulin and different carotenoids for the production of SCFAs in an ex vivo model for simulated colonic fermentation of carotenoids, incubated with or without inulin, in overweight human adults.

[0176] From the prior art, it is known that there is a causal association between the gut microbiota and the development of low-grade inflammation and insulin resistance associated with obesity and lipid-rich diets. Further, SCFAs are known to have a beneficial effect on appetite regulation via activating GPCR (FFAR2 and 3) and the release of satiety hormones. Therefore, the synergistic effect on SCFA production observed in the examples of the present application indicates that the inventive combination of inulin and at least one carotenoid is suitable for use as a medicament, a dietary supplement, or a food supplement in a subject suffering from overweight or obesity. Table 10 Overview of samples for which a synergistic effect present between a carotenoid and inulin is observed.

[0177] +: Synergism observed, see Tables 6 to 9 ++: Sample with highest synergism observed, see Tables 6 to 9

[0178] No synergism observed, see Tables 6 to 9

Claims

CLAIMS1 . A combination of at least one carotenoid and inulin for use in affecting the gut microbiota in a subject suffering from irritable bowel syndrome (IBS), wherein the carotenoid is selected from the group consisting of: lycopene, lutein, and zeaxanthin; and isomers, derivatives, and combinations thereof.

2. A combination of at least one carotenoid and inulin for use in affecting the gut microbiota in a subject suffering from overweight or obesity, wherein the carotenoid is selected from the group consisting of: lycopene, lutein, and zeaxanthin; and isomers, derivatives, and combinations thereof.

3. The combination for use according to claim 1 or 2, wherein the carotenoid is lycopene or an isomer or derivative thereof.

4. The combination for use according to claim 1 or 3, wherein affecting the gut microbiota involves:(i) preventing, reducing or slowing down the development of low-grade gut inflammation, and / or(ii) beneficially regulating gut motility, and / or(iii) restoring or maintaining the gut barrier function, and / or(iv) beneficially modulating the neurotransmitter production in the gut.

5. The combination for use according to claim 2 or 3, wherein affecting the gut microbiota involves:(i) preventing, reducing or slowing down the development of low-grade gut inflammation, and / or(ii) preventing, reducing or slowing down the development of insulin resistance.

6. The combination for use according to any of claims 2 or 5, wherein affecting the gut microbiota involves effecting appetite regulation in the subject.

7. The combination for use according to any of claims 1 , 2, 4 or 5, wherein affecting the gut microbiota involves increasing the amount of SCFAs in the gut of the subject.

8. The combination for use according to claim 7, wherein the SCFAs are selected from the group consisting of acetate, propionate, and butyrate, and combinations thereof.

9. The combination for use according to claim 1 or 4, wherein the SCFAs comprise butyrate.

10. The combination for use according to any of claims 4 or 5, wherein the prevention, reduction or slowing down of development of low-grade gut inflammation, and / or the increase in the amount of SCFAs is in comparison to a non-administered subject; incomparison to a subject administered carotenoids alone; in comparison to a subject administered inulin alone; or in comparison to a subject administered carotenoids alone and in comparison to a subject administered inulin alone.11 . The combination for use according to any of claims 1 to 10, wherein the combination is for simultaneous administration or consumption.

12. The combination for use according to any of claims 1 to 11 , wherein said use comprises administering to the subject a combination of inulin in an amount of 0.1 to 40 g / day and the at least one carotenoid in an amount of 1 to 70 mg / day.

13. The combination for use according to any of claims 1 to 11 , wherein the combination is used as a medicament, a dietary supplement, or a food supplement.

14. A composition comprising inulin and at least one carotenoid, wherein the carotenoid is selected from the group consisting of: lycopene, lutein, and zeaxanthin; and isomers, derivatives, and combinations thereof, wherein inulin is comprised in an amount of 0.1 to 40 g, and the at least one carotenoid is comprised in an amount of 1 to 70 mg.

15. Use of the composition according to claim 14 as a dietary supplement or a food supplement.

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