Plasma urolithin metabolites correlate with improvements in endothelial function after red raspberry consumption: A double-blind randomized controlled trial

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Abstract

Raspberries are a rich source of ellagitannins and anthocyanins. The aim of this work was to investigate whether raspberry consumption can improve vascular function and to understand which phenolic metabolites may be responsible for the effects. A 3 arm double-blind randomized controlled crossover human intervention trial was conducted in 10 healthy males. Flow-mediated dilation (FMD) was measured at baseline, 2 h, and 24 h post-consumption of 200 g and 400 g of red raspberries containing 201 or 403 mg of total (poly)phenols, or a matched control drink. Raspberry (poly)phenol metabolites were analyzed in plasma and urine by UPLC-QTOF mass spectrometry using authentic standards. Significant improvements in FMD were observed at 2 h (1.6% (95%CI 1.2, 1.9) and 1.2% (95% CI 0.8, 1.5)) and 24 h (1.0% (95% CI 0.6, 1.2) and 0.7% (95%CI 0.2, 0.9)) post-consumption of the 200 and 400 g raspberry drinks as compared to control, respectively. Plasma ellagic acid, urolithin A-3-glucuronide and urolithin A-sulfate correlated with the improvements in FMD at 2 and 24 h post consumption, respectively. Consumption of dietary achievable amounts of red raspberries acutely improves endothelial function up to 24 h and ellagitannins may be responsible for the observed effect.

Introduction

Red raspberries are one of the most common berry fruits consumed in the US with a yearly average availability of 560 g per capita in 2015 and still increasing in popularity today (https://www.ers.usda.gov). Despite being a very popular fruit, to our knowledge no clinical study has been published investigating the vascular effects of red raspberries in human subjects, although a number of preclinical studies have indicated potential health benefits [[1], [2], [3], [4]].

Red raspberries are good sources of putatively bioactive (poly)phenolic compounds, including ellagitannins and anthocyanins [5,6], which have been associated with lower cardiovascular disease (CVD) risk according to recent meta-analyses [7,8]. Evidence from randomized controlled trials with ellagitannin-rich sources (i.e. strawberries, pomegranate, and walnuts) have shown improvements in surrogate biomarkers of cardiovascular risk such as blood lipids, glycemic index, and blood pressure in at-risk individuals [[9], [10], [11], [12], [13], [14]]. The bioavailability of (poly)phenols in plasma is mainly characterized by an early absorption in the small intestine followed by a late colonic uptake. As early as 1 h after red raspberry consumption, ellagic acid was reported in plasma in low nanomolar concentrations [15]. Most of the ellagitannins, however, travel further down the intestine and reach the colon after a few hours where they are transformed into urolithin catabolites by colonic microbiota [16]. In contrast to other phenolic compounds, urolithins remain in the circulation for up to 80 h after consumption [17,18]. Anthocyanins, also present in red raspberries, were shown to be absorbed and metabolized – both in the early and later phases - with main phenolic breakdown products and metabolites identified in urine and plasma as cinnamic acids, hippuric acids, phenylacetic acids, phenylpropionic acids, and benzoic acids [19,20].

In the current study, we aimed to investigate the potential vascular benefits of red raspberries in healthy humans and identify which of the berry (poly)phenol metabolites may be responsible for the effects.

Section snippets

Study subjects

Ten healthy male volunteers aged 18–35 years were recruited from the University of Düsseldorf and surrounding area. Health was ascertained by a routine clinical physical exam and specific cardiovascular history performed by a cardiovascular specialist. Manifest cardiovascular disease including coronary artery disease, cerebrovascular disease, and peripheral artery disease, diabetes mellitus, acute inflammation, terminal renal failure, malignancies, and heart rhythm other than sinus were

Baseline characteristics of the study population and tolerance of intervention

The baseline characteristics of the group of healthy young non-obese males were all within normal limits (Table 1). All study subjects completed the study, drinks were well tolerated by all subjects, and no adverse events were reported.

(Poly)phenol content of the red raspberry drinks

A total of 27 (poly)phenolic compounds were quantified in the raspberry drinks used in the present study, including 2 ellagitanins, 2 anthocyanins, 5 flavonols, 2 flavan-3-ols, 8 cinnamic acids, 6 benzoic acids, and 2 benzaldehydes (Table 2). In 200 g and 400 g

Discussion

In summary, the current study demonstrate for the first time that the consumption of red raspberries can increase endothelial function for 24 h and that this effect is associated with patterns of circulating ellagitanin metabolites in healthy humans.

In the search for the mechanisms of action of dietary (poly)phenols and causality assumptions related to this, it is essential to consider the pharmacokinetics of these compounds. Most studies investigating the acute effects of (poly)phenols on

Acknowledgments

The authors' responsibilities were as follows: ARM and CH designed the research; GI, RPF, TW and RGV conducted the research; GI, RPF, TW, and RGV analyzed the data and performed the statistical analysis; ARM, GI, CH and FTB wrote the manuscript; ARM, GI and CH had primary responsibility for the final content; and all authors read and approved the final manuscript.

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      All studies had varying intervention duration, ranging from the short duration of < 1 month (Huang et al., 2021; Istas et al., 2018; Mellen et al., 2010; Moazzen & Alizadeh, 2017; Pokimica et al., 2019; Stote et al., 2017), 1–3 months (Amani et al., 2014; Basu et al., 2011; Feresin et al., 2017; Hsia et al., 2020; Johnson et al., 2015; Kojadinovic et al., 2017; Loo et al., 2016; Nilsson et al., 2017; Paquette et al., 2017; Quirós-Fernández et al., 2019; Richter et al., 2021; Stull et al., 2015), and > 3 months (Curtis et al., 2019; Istas et al., 2019; Park et al., 2016; Puupponen‐Pimiä et al., 2013). The daily dosage of dietary polyphenols and ETs supplementation was quite varied among studies, ranging from < 500 mg/daily (Basu et al., 2011; Curtis et al., 2019; Feresin et al., 2017; Hsia et al., 2020; Istas et al., 2018; Istas et al., 2019; Mellen et al., 2010; Moazzen & Alizadeh, 2017; Paquette et al., 2017; Park et al., 2016; Pokimica et al., 2019; Quirós-Fernández et al., 2019; Richter et al., 2021; Tjelle et al., 2015), 500–1000 mg (Curtis et al., 2019; Erlund et al., 2008; Huang et al., 2021; Johnson et al., 2015; Kojadinovic et al., 2017; Nilsson et al., 2017; Puupponen‐Pimiä et al., 2013; Stockton et al., 2017; Stull et al., 2015), and > 1000 mg (Amani et al., 2014; Loo et al., 2016; Pokimica et al., 2019; Stote et al., 2017; Tjelle et al., 2015). Interventions included juices (Amani et al., 2014; Basu et al., 2011; Curtis et al., 2019; Feresin et al., 2017; Hsia et al., 2020; Huang et al., 2021; Johnson et al., 2015; Loo et al., 2016; Pokimica et al., 2019; Richter et al., 2021; Stull et al., 2015; Tjelle et al., 2015), supplements/extract (Istas et al., 2019; Mellen et al., 2010; Quirós-Fernández et al., 2019; Stockton et al., 2017), and fresh fruits (Erlund et al., 2008; Istas et al., 2018; Istas et al., 2019; Nilsson et al., 2017; Paquette et al., 2017; Puupponen‐Pimiä et al., 2013).

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