Resultat från epidemiologiska studier har antytt att flavonolrik kakao sänker blodtrycket. Detta sker troligen genom att flavonoider i kakao leder till aktivering av eNOS som sedan katalyserar NO-syntesen i kärlendotel vilket leder till vasodilation och sänkt blodtryck. En nyare studie har funnit att kakao dessutom inhiberar enzymet ACE. Mycket tyder även på att NO-koncentrationen är kopplad till ACE-aktiviteten då dessa faktorer visat sig följa varandra inverst vid varierande doser. Det finns också ett samband mellan ACE-genotyp och serumnivåer av ACE. Någon dosberoende effekt av kakao på blodtrycket har inte påvisats, men ihållande dosering med kakao över en längre period visar på en progressiv sänkning av blodtrycket. Inga biverkningar har rapporterats, men fett och socker i kakao kan leda till diabetes och hjärt-kärlsjukdomar. Syftet med blodtrycksbehandling är prevention av nyss nämnda sjukdomar och adekvat blodtryckssänkning krävs därmed för att en behandling ska kunna övervägas. Än så länge ses endast en signifikant men inte en tillräcklig blodtryckssänkande effekt av kakao. Dessutom saknas studier som visar att kakao förebygger kardiovaskulära händelser, exempelvis hjärtinfarkter. Vidare studier krävs innan kakao kan rekommenderas som behandling.

The intestinal microbiome contributes in immune system function. It contains a large number of microorganisms that interact with each other and thus affect the host. Currently, attention is being directed towards investigating the influence of the intestinal microbiome on hosts, but also on foreign substances. Foreign substances may influence its composition and subsequent microbial metabolism. Crohn's disease patients have been found to have lower bacterial representation of beneficial bacteria. Therefore it is appropriate to examine the intestinal microbiome of these patients and so understand in greater detail the influence of bacteria on the course of the disease progression and on the medication used. The RP-HPLC method were analysed the faecal samples collected (B, C, D), which were incubated at 0, 3 and 6 hours. The incubation took place the addition of myricetin in the McDougall buffer and BHI medium. The analysis was found that myricetin degradation takes place in faecal samples during incubation, regardless of the medium used. In the faecal sample B, degradation of myricetin occurs faster in the BHI medium than in the McDougall's buffer. In faecal samples C and D, degradation is similar in both media. From these results, it is impossible to judge which medium is more suitable for...

Flavonols are a major subclass of flavonoids and are considered the most abundant subclass of flavonoids. Flavonols are classified as having a hydroxyl group on the 3rd carbon of the C ring. The most prevalent modification to flavonols is glucosylation. The flavonol specific 3-O glucosyltransferase (Cp3GT) enzyme from grapefruit (Citrus paradisi) is the topic of this research and specifically adds glucose to flavonols at the 3-OH position. The level of activity varies depending on the flavonol structure. This makes Cp3GT an ideal model system for studying the structure/function relationship of Cp3GT using site-directed mutants. S20G+T21S is a mutant form of Cp3GT. As compared to the wild type Cp3GT, S20G+T21S has significantly higher activity with kaempferol, quercetin, and gossypetin. Another interesting difference of S20G+T21S is its ability to add glucose to the 7-OH position of the flavanone naringenin, thus showing a change in flavonoid class specificity as well as regiospecificity for position of glucose attachment. The S20G+T21S mutant was first made by site-directed mutagenesis and verified by DNA sequencing. The linearized pPICZa plasmid containing S20G+T21S was then transformed into Pichia pastoris via electroporation. Transformation was verified by colony PCR and DNA sequencing and a time course analysis of methanol-induced expression conducted to identify optimal expression. Optimal expression was identified at 24 hours and was verified by SDS-page gel and a western blot. S20G+T21S was purified by IMAC column in preparation for crystallization.

Flavonoid and limonoid glycosides influence taste properties as well as marketability of Citrus fruit and products, particularly grapefruit. In this work, nine grapefruit putative natural product glucosyltransferases (PGTs) were resolved by either using degenerate primers against the semiconserved PSPG box motif, SMART-RACE RT-PCR, and primer walking to full-length coding regions; screening a directionally cloned young grapefruit leaf EST library; designing primers against sequences from other Citrus species; or identifying PGTs from Citrus contigs in the harvEST database. The PGT proteins associated with the identified full-length coding regions were recombinantly expressed in Escherichia coli and/or Pichia pastoris and then tested for activity with a suite of substrates including flavonoid, simple phenolic, coumarin, and/or limonoid compounds. A number of these compounds were eliminated from the predicted and/or potential substrate pool for the identified PGTs. Enzyme activity was detected in some instances with quercetin and catechol glucosyltransferase activities having been identified.

Breast cancer is the most commonly diagnosed cancer in women with an estimated 287,850 cases in 2022. Approximately 684,000 deaths each year are associated with breast cancer across the world. Risk factors of breast cancer include increased estrogen exposure, family history of breast cancer, and environmental factors. Treatment of breast cancer is highly dependent on the presence of HER2, estrogen, and progesterone receptors. Breast cancers that present with increased receptors for estrogen, progesterone, and HER2 are typically the least aggressive and the easiest to treat. The percentage of cases in the United States associated with hormone receptor positive and HER-2 negative or positive are approximately 82%. Absence of receptors for estrogen, progesterone, and HER2 is known as triple negative breast cancer. In the United States, only about 10% of cases are associated with this form. However, it is considered the most aggressive and difficult to treat. Two emerging flavonoids known as CT1 and CT3 have shown cytotoxic activity against cell lines that represent some of the most common breast cancers: MCF7, MDA-MB-231, and SKBr3. CT1 and CT3 were extracted from the leaves of Chromolaena tacotana using a Soxhlet extractor and the compounds then underwent isolation and purification. The cells were then treated with CT1 or CT3 at concentrations of 5, 10, 20, 40 and 80 µM. MTT assays were then used to determine cell viability. MDA-MB-231, the most aggressive type of breast cancer cells, responded to both CT1 and CT3. The most profound cytotoxic effects of CT1 were seen with MCF7 and MDA-MB-231, while CT3 exhibited a greater toxicity against SKBr3 cells. Preliminary results indicate that CT1 and CT3 target the MEK-ERK signaling pathway. Further studies need to be completed to determine mechanistically how these compounds lead to receptor-independent toxicity.

Flavonoids are plant secondary metabolites that are integral to our lives. Grapefruits are well-known for production of unique glucosylated products and the enzymes responsible are UDP-glucose:glucosyltransferases (GTs). The objective of this research was to obtain full-length clones of putative grapefruit GTs, express them, and characterize them. Previously, gene specific primers (from conserved PSPG box) and clone specific primers (from partial 5' clones) were designed, and a compiled sequence attained using SMART RACE RT-PCR. A full-length clone was obtained using primers designed from the extreme ends of the compiled sequence. The full-length clone was inserted into expression vector (pET32a) and transformed into expression host BL21(DE3)RIL. Expressed protein was tested for GT activity using different flavonoid aglycones and UDP-14C-glucose as glucose donor. Results indicated that the expressed protein was probably not a flavonoid GT. A directionally cloned grapefruit leaf cDNA library is undergoing EST mining to identify additional GT candidates.

Flavonoids are polyphenolics compounds that constitute a major group of plant specialized metabolites, biosynthesized via the phenylpropanoid/polymalonate pathways. The resulting specialized metabolites can be due to decoration of flavonoid compounds with sugars, usually glucose, by the action of regiospecific UDP-glycosyltransferase (UGT) enzymes. In some cases, glycosylation can involve enzymatic attachment of other sugar moieties, such as glucuronic acid, galactose, rhamnose or arabinose. These modifications facilitate or impact the bioactivity, stability, solubility, bioavailability and taste of the resulting flavonoid metabolites. The present work shows the limitations of utilizing mammalian UDP-glucuronosyltransferases (UGATs) for flavonoid glucuronidation, and then proceeds to investigate plant UG(A)T candidates from the model legume Medicago truncatula for glucuronidating brain-targeted flavonoid metabolites that have shown potential in neurological protection. We identified and characterized several UG(A)T candidates from M. truncatula which efficiently glycosylate various flavonoids compounds with different/multiple regiospecificities. Biochemical characterization identified one enzyme, UGT84F9, that efficiently glucuronidates a range of flavonoid compounds in vitro. In addition, examination of the ugt84f9 gene knock-out mutation in M. truncatula indicates that UGT84F9 is the major UG(A)T enzyme that is necessary and sufficient for attaching glucuronic acid to flavonoid aglycones, particularly flavones, in this species. Finally, the identified UG(A)T candidates were analyzed via homology modeling and site-directed mutagenesis towards increasing the repertoire of UG(A)Ts applicable for synthesis of flavonoid glucuronides with potential human health benefits in neurological protection.

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Yes / Extracts of Erythrina addisoniae are frequently used in the traditional medicine of Western Africa, but insufficient information about active compounds is available. From the stem bark of E. addisoniae, three (1, 2, 4) and three known (3, 5, 6) flavanones were isolated: addisoniaflavanones I and II, containing either a 2″,3″-epoxyprenyl moiety (1) or a 2″,3″-dihydroxyprenyl moiety (2) were shown to be highly toxic (MTT assay: EC50 values of 5.25 ± 0.7 and 8.5 ± 1.3 μM, respectively) to H4IIE hepatoma cells. The cytotoxic potential of the other isolated flavanones was weaker (range of EC50 values between 15 and >100 μM). Toxic effects of addisoniaflavanone I and II were detectable after 3 h (MTT assay). Both compounds induced an apoptotic cell death (caspase-3/7 activation, nuclear fragmentation) in the hepatoma cells and, at high concentrations, also necrosis (membrane disruption: ethidium bromide staining). Formation of DNA strand breaks was not detectable after incubation with these compounds (comet assay). In conclusion, the prenylated flavanones addisoniaflavanones I and II may be of interest for pharmacological purposes due to their high cytotoxic and pro-apoptotic potential against hepatoma cells.