Are High Proanthocyanidins Key to Cranberry Efficacy in the Prevention of Recurrent Urinary Tract Infection?

Abstract

Most research on American cranberry in the prevention of urinary tract infection (UTI) has used juices. The spectrum of components in juice is limited. This study tested whether whole cranberry fruit powder (proanthocyanidin content 0.56%) could prevent recurrent UTI in 182 women with two or more UTI episodes in the last year. Participants were randomized to a cranberry (n = 89) or a placebo group (n = 93) and received daily 500 mg of cranberry for 6 months. The number of UTI diagnoses was counted. The intent-to-treat analyses showed that in the cranberry group, the UTIs were significantly fewer [10.8% vs. 25.8%, p = 0.04, with an age-standardized 12-month UTI history (p = 0.01)]. The Kaplan-Meier survival curves showed that the cranberry group experienced a longer time to first UTI than the placebo group (p = 0.04). Biochemical parameters were normal, and there was no significant difference in urinary phenolics between the groups at baseline or on day180. The results show that cranberry fruit powder (peel, seeds, pulp) may reduce the risk of symptomatic UTI in women with a history of recurrent UTIs. Copyright © 2015 John Wiley & Sons, Ltd.
Copyright © 2015 John Wiley & Sons, Ltd.

Full text

Are High Proanthocyanidins Key to Cranberry
Efficacy in the Prevention of Recurrent Urinary
Tract Infection?
Jitka Vostalova,
1
Ales Vidlar,
2
*Vilim Simanek,
1
Adela Galandakova,
1
Pavel Kosina,
1
Jan Vacek,
1
Jana Vrbkova,
3
Benno F. Zimmermann,
4,5
Jitka Ulrichova
1
and Vladimir Student
2
1
Department of Medical Chemistry and Biochemistry, Faculty of Medicine and Dentistry, Palacky University Olomouc, Hnevotinska 3,
77515, Olomouc, Czech Republic
2
Department of Urology, University Hospital, I.P. Pavlova 5, 77500, Olomouc, Czech Republic
3
Institute of Molecular and Translational Medicine Department, Faculty of Medicine and Dentistry, Palacky University Olomouc,
Hnevotinska 3, 77515, Olomouc, Czech Republic
4
Department of Nutritional and Food Sciences - Chair of Food and Technology and Food Biotechnology, University of Bonn,
Römmerstrasse 164, 53117, Bonn, Germany
5
Institut Prof. Dr. Georg Kurz GmbH, Stöckheimer Weg 1, 50829, Köln, Germany
Most research on American cranberry in the prevention of urinary tract infection (UTI) has used juices. The
spectrum of components in juice is limited. This study tested whether whole cranberry fruit powder
(proanthocyanidin content 0.56%) could prevent recurrent UTI in 182 women with two or more UTI episodes
in the last year. Participants were randomized to a cranberry (n= 89) or a placebo group (n= 93) and received
daily 500 mg of cranberry for 6 months. The number of UTI diagnoses was counted. The intent-to-treat analyses
showed that in the cranberry group, the UTIs were significantly fewer [10.8% vs. 25.8%, p= 0.04, with an
age-standardized 12-month UTI history ( p= 0.01)]. The Kaplan–Meier survival curves showed that the
cranberry group experienced a longer time to first UTI than the placebo group ( p= 0.04). Biochemical
parameters were normal, and there was no significant difference in urinary phenolics between the groups at
baseline or on day180. The results show that cranberry fruit powder (peel, seeds, pulp) may reduce the risk of
symptomatic UTI in women with a history of recurrent UTIs. Copyright © 2015 John Wiley & Sons, Ltd.
Keywords: Vaccinium macrocarpon; urinary tract infection; recurrent; haematology; clinical chemistry markers; urinary metabolites.
INTRODUCTION
Urinary tract infections (UTIs) are the most commonly
diagnosed bacterial infection in women, with more than
50% experiencing at least one UTI during their lifetime
(Foxman, 2003; Micali et al., 2014). The ingestion of
cranberries (Vaccinium macrocarpon Ait., V. oxycoccus
L., Ericaceae) has traditionally been associated with
the prevention of UTIs, which arise from colonization
and subsequent infection by uropathogenic Escherichia
coli (Guay, 2009). Major classes of constituents that
may contribute to the health benefits of cranberry are phe-
nolic acids, flavonoids, anthocyanins, proanthocyanidins
and triterpenoids (Pappas and Schaich, 2009). A-type
proanthocyanidins (PACs) seem to be responsible for
inhibiting the adhesion of E. coli and other uropathogens
to uroepithelial cells in vitro (Foo et al., 2000; Howell
et al., 2005) and ex vivo (Howell et al., 2010; Lavigne
et al., 2011). On the other hand, an ex vivo urine
antiadhesive effect has been ascribed to high concentra-
tions of hippuric and salicyluric acids in the urine of
healthy women consuming a daily dose of 1200 mg of
dried cranberry juice (Valentova et al., 2007). Another
possible mechanism of polyphenolic metabolite action
could be the selection of less adherent bacterial strains
in the gastrointestinal tract (Raz et al., 2004). As food,
and dietary supplements, cranberry is used in juice,
juice cocktail (approximately 26% to 33% pure cran-
berry juice), cranberry pills/capsules containing dried
juice or cranberry fruit powder (100% cranberry fruit
solids). The complex mixture of bioactive components
is found only in whole cranberry fruit (Grace et al.,
2012).
The growing concern over antibiotic resistance has
stimulated interest in cranberries in the prevention
of recurrent UTIs (rUTI). Over the years, a number
of randomized clinical trials has been conducted to
assess the efficacy of cranberry in reducing the risk of
rUTIs in women (Micali et al., 2014; Jepson et al.,
2012; Wang et al., 2012). Most clinical trials in adult
women with a history of rUTIs or acute bacteriuria
used pure cranberry juice/cocktails or capsules contain-
ing dried juice enriched with A-type PACs. A recent
randomized clinical study showed that 500/1000 mg of
cranberry fruit powder (CFP) used for 90 days signifi-
cantly reduced bacteriuria and symptoms of UTI in
women with symptomatic UTI at baseline (Sengupta
et al., 2011). Interestingly, this appears to be the only
study of cranberry being used in the treatment of
acutely infected subjects.
One weakness of clinical trials with cranberry prod-
ucts has been that the cranberry preparations were
not fully characterized by chemical composition. The
* Correspondence to: Ales Vidlar, Department of Urology, University
Hospital, I.P. Pavlova 5, 77500 Olomouc, Czech Republic.
E-mail: alevi@centrum.cz
PHYTOTHERAPY RESEARCH
Phytother. Res. 29: 1559–1567 (2015)
Published online 13 August 2015 in Wiley Online Library
(wileyonlinelibrary.com) DOI: 10.1002/ptr.5427
Copyright © 2015 John Wiley & Sons, Ltd.
Received 09 July 2015
Accepted 13 July 2015

efficacy of powdered whole cranberry fruit (quantified
using standardized methods) in reducing the risk of
rUTI in female subjects free of UTI at baseline
has not yet been assessed. The aim of this 6-month
randomized, double-blind and placebo-controlled trial
was to evaluate whether a daily dose of 500 mg of cran-
berry fruit powder could prevent rUTIs in otherwise
healthy women from 18 to 75 years old, with a history
of UTIs.
MATERIAL AND METHODS
Cranberry material. Cranberry fruit powder (100%
fruitofNorthAmericanVaccinium macrocarpon
Aiton, Ericaceae; Batch No. 090921) was purchased
from NATUREX-DBS (Sagamore, MA, USA).
Declared total PACs in CFP determined by the
4-dimethylaminocinnamaldehyde (DMAC) method
(Prior et al., 2010) was 0.56%, and this powder was ap-
plied for the clinical trial. The results of our secondary
metabolite determination of CFP by HPLC-ESI-MS/
MS are shown in Table 1; PACs have been analysed
according to Jungfer et al., 2012 (Table 2). Each cran-
berry capsule contained 250 mg of CFP (1.4mg of PACs
according to DMAC). Placebo capsules contained low-
density STAR-DRI® 1015A maltodextrin, canola oil,
Red 40 Lake, sodium aluminium silicate and Blue 1
Lake. CFP capsules were indistinguishable in appear-
ance from the placebo capsules.
Design and participants. The study was conducted ac-
cording to the guidelines of the Helsinki Declaration
(2008 revision), and all procedures involving human
subjects were approved by the Ethics Committee of
the University Hospital and Faculty of Medicine and
Dentistry, Palacky University, Czech Republic (refer-
ence 129/09). Written informed consent was obtained
from all participants. The study was a 6-month,
single-centre, randomized, double-blind and placebo-
controlled trial consisting of two parallel treatment
arms. It was conducted between January 2010 and
April 2011 at the Clinic of Urology of the University
Hospital.
The invitation to participate was through the refer-
ring physician treating the UTIs. Women aged over
18 years old and with a medical history of at least
two episodes of symptomatic UTIs in the previous
12 months were eligible. Participants meeting all of the
Table 1. Content of selected secondary metabolites in cranberry fruit powder
Compound (mg/100 g CFP)
Phenolic acids Benzoic acid and benzaldehyde derivatives
3,4-Dihydroxycinnamic acid 4.4 ± 0.1 Benzoic acid 167.6 ± 19.4
Chlorogenic acid 12.4 ± 0.4 3,4-Dihydroxybenzaldehyde 2.7 ± 0.2
4-Hydroxy-3-methoxycinnamic acid 2.7 ± 0.1 p-Hydroxybenzaldehyde 0.03 ± 0.01
3,4,5-Trihydroxybenzoic acid 5.8 ± 0.2 Vanillin 0.4 ± 0.0
4-Hydroxycinnamic acid 21.1 ± 0.5
4-Hydroxybenzoic acid 0.3 ± 0.0 Anthocyanins/Anthocyanidins
3,4-Dihydroxybenzoic acid 24.3 ± 0.5 Cyanidin 3-O-arabinoside 62.7 ± 1.0
2-Hydroxybenzoic acid 0.3 ± 0.0 Cyanidin 3-O-galactoside 42.4 ± 1.9
4-Hydroxy-3,5-dimethoxycinnamic acid 2.8 ± 0.1 Cyanidin 31.6 ± 1.3
4-Hydroxy-3,5-dimethoxybenzoic acid 2.8 ± 0.1 Delphinidin 3-O-glucoside 2.7 ± 0.2
4-Hydroxy-3-methoxybenzoic acid 5.8 ± 0.1 Delphinidin 24.4 ± 0.8
Flavonoids Malvidin 3-O-galactoside 5.4 ± 0.2
Apigenin 0.4 ± 0.0 Pelargonidin 3-O-glucoside 7.6 ± 0.2
Catechin 2.7 ± 0.1 Peonidin 3-O-glucoside 82.1 ± 1.3
Epicatechin 13.4 ± 0.1 Peonidin 3-O-rutinoside 1.3 ± 0.2
Epigallocatechin 24.4 ± 0.2 Peonidin 13.5 ± 1.1
Hesperidin 2.6 ± 0.1
Hyperoside 1408.0 ± 36.7 Pentacyclic triterpenoid
Isorhamnetin 188.2 ± 22.4 Ursolic acid 921.6 ± 74.2
Kaempferol 25.2 ± 4.6
Myricetin 482.8 ± 21.7
Isoquercitrin 504.0 ± 14.9
Quercetin 1138.8 ± 32.5
Rutin 3.0 ± 0.2
Values are expressed as mean ± SD, n=5.
Table 2. Content of proanthocyanidins in cranberry fruit powder
Proanthocyanidin (mg/100 g CFP)
B-dimer B1 0.54 ± 0.01 A-trimer 3 3.06 ± 0.08
B-dimer B2 2.29 ± 0.08 A-trimer 4 2.86 ± 0.17
B-dimer B5 0.72 ± 0.04 A-trimer 7 1.92 ± 0.08
Procyanidin A2 24.30 ± 0.43 A-trimer 8 4.44 ± 0.04
Values are expressed as mean ± SD, n= 2. Concentrations are cal-
culated as procyanidin A2 equivalents. Compound nomenclature is
according to Jungfer et al., 2012.
Copyright © 2015 John Wiley & Sons, Ltd. Phytother. Res. 29: 1559–1567 (2015)
1560 J. VOSTALOVA ET AL.

inclusion criteria and none of the exclusion criteria
(Table 3) and consenting to study participation were
randomly divided into two groups: cranberry and placebo
groups (Fig. 1). The randomization plan for treatment as-
signment to subjects was generated using online software
QuickCalcs (GraphPad Software Inc., USA). The cran-
berry group was given 500 mg CFP (two times 250 mg
CFP capsules) to be taken once a day after breakfast for
the 6-month period. The daily dose of CFP was based
on the findings of McMurdo et al. (2009). The placebo
Table 3. Eligibility criteria
Inclusion criteria Exclusion criteria
•Women from 18 to 75 years •Symptomatic UTI at baseline
•A history of recurrent symptomatic UTIs (defined as a medical
history of at least two symptomatic UTI episodes treated with
antibiotics in the previous 12 months)
•Antibiotic treatment during the study for reasons others
than UTI
a
•Pregnant and/or breast feeding women
•Clinical laboratory tests (haematology, clinical chemistry,
urinalysis) within normal reference ranges or if outside the
normal reference ranges, clinically insignificant
•Anatomical anomalies or other pathological findings with a
possible effect on the recurrence of UTIs (stricture of urethra,
nephrolithiasis, cystolithiasis, neurogenic bladder dysfunction)
•Insulin-dependent diabetes mellitus
•Subjects with a history of medical or surgical events that could
affect the study outcome or place the subject at risk, including
cardiovascular disease, gastrointestinal problems, metabolic,
renal, hepatic, neurological, sexually transmitted diseases or
active musculoskeletal disorders
•Immunocompromised individuals or individuals receiving
immunosuppressive medication
•Intermittent or indwelling urinary tract catheterization
•Subjects with a history of surgery within the last 6 months
•Use of narcotics
•Heavy episodic drinking of alcohol
•Participation in a clinical research trial within 30 days prior to
randomization
•Simultaneous participation in another clinical trial
a
Patients with occurrence of a symptomatic UTI during the study were treated immediately with antibiotics. At the end of antibiotics treatment,
urine samples were collected to confirm the absence of bacteriuria, and patients resumed study treatment.
Figure 1. Flow chart of the clinical trial.
Copyright © 2015 John Wiley & Sons, Ltd. Phytother. Res. 29: 1559–1567 (2015)
1561ARE HIGH PAC KEY TO CRANBERRY EFFICACY IN THE PREVENTION RUTI?

group received the same instructions as the cranberry
group. Subjects were asked to refrain from consuming
foods rich in phenolics, especially colour-pigment-
containing fruit (berries), or vitamin supplements or
to make any other dietary or lifestyle changes during
the study. After randomization (baseline), the women
returned to the clinic at 3 and 6 months and whenever
they experienced symptoms of a UTI. The clinical
report form included (i) a detailed medical history,
(ii) assessment of all concurrent medication and treat-
ment, (iii) dietary habits, (iv) kidney and bladder ultra-
sound and (v) complete laboratory analysis, including
haematology, clinical chemistry and urinalysis. Urine
samples were collected at baseline and at 3 and 6months
for analysis of the urine and urine sedimentation. If a
UTI was confirmed (bacteriuria≥10
5
cfu/mL plus symp-
toms of a UTI; see Urinalysis, microbial examination
and clinical diagnosis of UTI), the subject was treated
with antibiotics, culture-directed antibiotic treatment for
1–3 days. Once the course of antibiotics was completed,
urine samples were collected to confirm that the UTI
had resolved, and the subject resumed taking the product.
Vital signs (heart rate, systolic and diastolic blood pres-
sure) were assessed at baseline and at 3 and 6months.
The clinical diagnosis of a UTI was based on bacteriuria
plus the manifestation of at least one of the following
symptoms: pollakiuria (strong, persistent urge to urinate
and passing frequent, small amounts of urine), burning
sensation on micturition, hematuria, turbid or malodor-
ous urine, subpelvic pain, pruritus, fever and dysuria.
Haematology and clinical chemistry. Blood samples
were collected at baseline and at 6 months. These were
drawn under aseptic conditions from the vena cubiti,
after a several-minute rest in the half-sitting position.
Serum/plasma samples were separated in a cooled
centrifuge at 3000 ×g for 20 min. Basic haematolog ical
parameters (haemoglobin, erythrocytes, leukocytes,
platelets and haematocrit) were measured in Na
2
EDTA
blood. Routine clinical chemistry parameters were
determined in all samples: low-density lipoprotein
(LDL), high-density lipoprotein (HDL) cholesterol,
triacylglycerol, C-reactive protein, alanine aminotrans-
ferase, aspartate aminotransferase, gamma-glutamyl
transpeptidase, urea, creatinine, bilirubin and glucose
were quantified in serum using a HITACHI Modular
Evo P analyser (Hitachi, Japan).
Urinalysis, microbial examination and clinical diagnosis
of UTI. The urine had to be a midstream early morning
sample. The complete analysis of urine was performed on
the IQ200 Automated Urinanalysis System (IRIS Inter-
national, Inc., USA). The microbiological analysis was
performed at the Laboratory of Clinical Microbiology,
University Hospital. The laboratory diagnosis of UTI
was based on a significant isolate of a single organism,
and a UTI was culture-confirmed when the growth of a
single bacterial strain was ≥10
5
cfu/mL in a midstream
urine sample. Phenolic metabolites in urine were deter-
mined at baseline and at 6months. Analysis of free and to-
tal phenolics in urine was carried out using HPLC-ESI-ion
trap MS according to our protocol (Heinrich et al., 2013).
Statistical analysis. The sample size was estimated based
on the assumption that at least 30% of women would
experience an rUTI within 6 months in the placebo
group and that the rate of UTI recurrence would be re-
duced to 15% in the cranberry group. The primary end-
point of this clinical trial was the 50% reduction in
incidence of rUTI episodes in the cranberry group com-
pared to the placebo group. In order to detect this effect
with a power of 80% and a two-tailed alpha level of 5%,
80 women per group were needed (Kontiokari et al.,
2001). Thus, to account for subject attrition, a total of
182 women were recruited.
An intent-to-treat (ITT) analysis was performed.
This included any individual with at least one
postrandomization assessment. In order to examine
the relationship between the proportion of women
experiencing at least one UTI episode during the study
period and assignment to the active or placebo treatment
arms, a complementary log–log (CLL) binomial regres-
sion model was used. This generalized linear model
(GLM) specified a binomial distribution for the random
component and a complementary log–log link function.
The model was fit using the GLM function in R. Age
and age-adjusted history of UTI were associated with risk
of UTI, and for this reason, they were included in the
model. The log observation time (from randomization
date to the end-of-study or dropout dates) was included
as an offset term. A Kaplan–Meier estimate was used to
describe the distribution of time to first UTI. In order to
compare time to first UTI between the two treatment
arms, the Cox proportional hazards model was fitted with
treatment arm, age and age-adjusted prior 12-month his-
tory of UTI in the model. The count of UTIs found during
the observation period was compared between groups
using Poisson regression including age, age-adjusted prior
12-month history and an offset variable (log observation
time) in the model. Continuous variables were described
as means ± standard deviation or first quartile/median/
third quartile and compared using a repeated-measures
ANOVA at baseline and after 6 months.
RESULTS
Patient recruitment is depicted in Fig. 1. Of the 803
women who were screened for participation in the
study, 621 patients did not meet the inclusion criteria
or met one or more of the exclusion criteria. The
remaining 182 eligible women were enrolled and were
randomized to the cranberry (n= 89) or placebo
(n= 93) groups. Seventeen women did not complete
the study, seven (7%) in the placebo group and ten
(11%) in the cranberry group. Reasons for not complet-
ing the study included loss to follow-up, voluntary with-
drawal (n= 14) or pregnancy (n= 3) (Fig. 1). These 17
women were included in the ITT analysis. Six other par-
ticipants, all randomized to the cranberry group, were
excluded from the ITT analysis. Three of these six study
participants had a UTI history of one in the previous
12 months and were excluded from the analysis because
they had less than two in the previous 12 months. The
other three study participants were enrolled into the
trial by the Principal Investigator following parental
consent but were excluded from the analysis because
they were younger than 18 years. Thus, the ITT sample
included a total of 176 women (n= 83 in the cranberry
group and n= 93 in the placebo group). The cranberry
and placebo groups were similar with regard to baseline
Copyright © 2015 John Wiley & Sons, Ltd. Phytother. Res. 29: 1559–1567 (2015)
1562 J. VOSTALOVA ET AL.

characteristics (Table 4, Fig. 2). There was a positive
quadratic association between age and prior 12-month
UTI history. The residuals from this quadratic model
were adopted as age-adjusted UTI history scores. Age
was also found to have a quadratic association with
occurrence of UTI during the intervention period and
hence age, centred around 40 years, and the square
were included in the following models. During the
6-month intervention, the proportion of women having
at least one UTI episode was significantly lower in the
cranberry group (9/83, 10.84%) than in the placebo
group (24/93, 25.81%) (p= 0.04), with age-adjusted
12 month U TI history ( p= 0.01), age ( p= 0.73), and
age-squared (p= 0.05) included in the model. This cor-
responds to a relative risk reduction of 58% in the cran-
berry group relative to the placebo group. The fitted
cumulative incidence (or cumulative rate) of UTI over
6 months for a women with average duration of obser-
vation, average age, and average UTI history was
0.085 (8.5%) in the cranberry group and 0.194 (19%)
in the placebo group (p= 0.04). The proportion of
women experiencing at least one UTI episode caused
specifically by E. coli was 7/83 women (8.43%) in the
cranberry group and 22/93 women (23.66%) in the pla-
cebo group ( p= 0.03 vs. placebo), with age-adjusted
prior 12-month UTI history (p= 0.007), age ( p= 0.74)
and age-squared (p= 0.12) included in the model.
The Kaplan–Meier curves for time to first UTI are
shown in Fig. 3. The time to first occurrence was differ-
ent for the two groups, with a significantly longer time to
first UTI observed in the cranberry group relative to the
placebo group (p= 0.04), with age-adjusted 12-month
UTI history (p= 0.02), age ( p= 0.69) and age-squared
(p= 0.04) included in the model. Of the women in the
cranberry group, 10% (Kaplan–Meier estimate) experi-
enced a UTI episode by 133 days while 10% of the
participants in the placebo group experienced a UTI
episode by 65 days.
During the study, there was a total of 40 UTIs that
occurred in 33 women. Thirty-three of the UTIs were
primary occurrences and seven were secondary occur-
rences (six women in the placebo group and one woman
in the cranberry group experienced two episodes of UTI
during the 6-month study). The average count of UTIs
per subject in the study period was 0.12 (10/83) for the
cranberry group and 0.32 (30/93) for the placebo group
20 30 40 50 60 70
0.0 0.2 0.4 0.6 0.8 1.0
Age (Year)
Probability
Placebo group
Cranberry group
Figure 2. Differences in age distribution between subjects in the
cranberry and placebo groups.
0 50 100 150 200
0.0 0.2 0.4 0.6 0.8 1.0
Time since randomization (Day)
1 − Cumulative probability of recurrent UTI
Placebo group
Cranberry group
Figure 3. Kaplan–Meier curves of survival to UTI recurrence for the
placebo and cranberry groups.
Table 4. Baseline parameters
Placebo group (n= 93) Cranberry group (n= 89) p
UTIs in the last 12 months 3.27 ± 1.33 2.93 ± 1.22 0.08
Age (years) 38.03 ± 13.40 35.61 ± 12.97 0.23
Height (cm) 167.70 ± 6.03 166.30 ± 6.73 0.14
Weight (kg) 66.44 ± 10.79 64.18 ± 12.52 0.20
Temperature (°C) 36.32 ± 0.21 36.35 ± 0.19 0.36
Pulse (beats per minute) 69.02 ± 6.89 70.47 ± 6.32 0.15
Systolic blood pressure (mmHg) 116.90 ± 11.03 115.8 ± 10.61 0.48
Diastolic blood pressure (mmHg) 77.31 ± 7.39 76.87 ± 6.87 0.68
Urine sediment (HPF) Negative Negative NA
Urine pH 5.76 ± 0.76 5.83 ± 0.67 0.64
Values are mean ± SD. UTIs, urinary tract infections.
Copyright © 2015 John Wiley & Sons, Ltd. Phytother. Res. 29: 1559–1567 (2015)
1563ARE HIGH PAC KEY TO CRANBERRY EFFICACY IN THE PREVENTION RUTI?

(p= 0.03, after adjusting for age-adjusted 12 month his-
tory, age and age-squared). All occurrences of UTI were
medically diagnosed and confirmed microbiologically.
Pathogens identified among the 33 primary UTIs were
E. coli (n= 28), Klebsiella species (n= 3), Staphylococcus
species (n= 1) and Streptococcus species (n= 1). In the
placebo group, one woman who experienced an infec-
tion with E. coli also had Enterococcus sp. identified in
the urine. Pathogens identified among the seven second-
ary UTIs were all E. coli.
Changes from baseline in haematology and clinical
chemistry parameters (Table 5) were similar for both
groups, with the exception of bilirubin, which increased
from baseline by 1.0 μmol/L in the placebo group and
decreased from baseline by 0.53 μmol/L in the cranberry
group (p<0.05 vs. placebo). All values remained within
the normal ranges. After 6 months of trea tment, there
were significant increases in total cholesterol (4.97
± 0.99 to 5 .18 ± 1.02 mmol/L), HDL ch olesterol (1.63
± 0.35 to 1.77 ± 0.77 mmol/L) and LDL cholesterol
(2.78 ± 0.87 to 2.95 ± 0.89 mmol/L) in the placebo group.
All values were within the normal ranges. There was a
slight yet significant decrease in urine pH between base-
line (5.83 ± 0.67) and the 6- month time point (5.64
±0.55; p= 0.024) in the cranberry group while the pH
in the placebo group did not change over time.
No anthocyanins or proanthocyanidins were detected
in either the plasma or urine samples of either group (data
not shown). The free and total concentrations of phenolic
compounds were determined in urine samples collected
on days 0 and 180. There was no significant difference in
phenolics between the groups (Table 6).
DISCUSSION
A number of systematic reviews and meta-analyses of
human interventional clinical trials on the effects of
cranberry on UTIs has been published (Micali et al.,
2014; Guay, 2009; Jepson et al., 2012; Wang et al.,
2012). The results of individual studies have been
largely inconsistent. These inconsistencies could be
due to (i) the populations studied [history of UTI, life
stage (pregnancy, menopausal status, age)], (ii) the study
settings (free-living vs. institutionalized), (iii) subject com-
pliance and/or (iv) the effectiveness of the cranberry-
containing product consumption. In this regard, the
prophylactic efficacy of various cranberry products (cran-
berry juice, dried juice, diluted juice concentrate, juice
cocktails, cranberry juice powder enriched with PACs)
has been tested; however, these products have been
largely uncharacterized in terms of chemical composi-
tion, making it difficult to assess their true potential
bioefficacy. In addition, subject compliance is generally
better in studies using cranberry juice in tablet or
capsule form (Stothers, 2002; McMurdo et al., 2009;
Beerepoot et al., 2011).
In this trial, the proportion of women experiencing at
least one UTI episode was significantly lower in the
group using cranberry fruit powder (10.8%) relative to
the placebo group (25.8%), although the recurrence
rate in the placebo group was lower than the expected
30% UTI reported by Kontiokari et al. (2001). The dif-
ference in the proportion of women experiencing a
UTI episode in the placebo and cranberry arms was sig-
nificant (p= 0.04), likely owing to a greater magnitude
of effect than was predicted in the sample size calcula-
tion. Although other trials of cranberry supplements
have reported reductions in the risk of rUTIs in women
with a history of UTIs, these had various limitations, in-
cluding inappropriate study design, small sample size
(Bailey et al., 2007; Walker et al., 1997), failure to fully
characterize the cranberry supplement (Stothers, 2002;
Beerepoot et al., 2011; Walker et al., 1997), lack of defi-
nition of the criteria that were used to diagnose a UTI
(Bianco et al., 2012) and/or a high rate of subject attri-
tion (Bailey et al., 2007; Walker et al., 1997). In contrast,
Table 5. Haematology and clinical biochemistry markers in placebo and cranberry groups at day 0 and day 180
Placebo group (n= 86) Cranberry group (n= 79)
Day 0 Day 180 Day 0 Day 180
Haemoglobin (g/L) 126/133/138 129/134/140
a
126/134/140 129/134/141
Erythrocytes (10
12
/L) 4.16/4.42/4.61 4.17/4.46/4.60 4.21/4.52/4.68 4.28/4.43/4.65
Leukocytes (10
9
/L) 4.81/6.02/7.04 4.83/6.14/6.89 4.91/5.94/6.94 5.13/5.82/7.20
Haematocrit 0.37/0.39/0.41 0.37/0.40/0.42
a
0.38/0.40/0.41 0.39/0.40/0.41
Platelets (10
9
/L) 208.0/237.5/275.8 216.8/245.5/287.8 224.5/261.0/293.0 229.5/266.0/309.0
a
Urea (mmol/L) 3.63/4.20/5.08 3.40/4.15/5.10 3.50/4.10/5.20 3.55/4.20/5.00
Creatinine (μmol/L) 60.3/66.0/73.0 61.0/64.5/71.0 61.0/68.0/73.5 61.0/66.0/72.0
Bilirubin (μmol/L) 5/8/9 6/8/11 6/8/11 5/7/11
b
Alanine aminotransferase (μkat/L) 0.24/0.32/0.42 0.22/0.30/0.42 0.25/0.30/0.37 0.22/0.28/0.39
Aspartate aminotransferase (μkat/L) 0.34/0.40/0.48 0.34/0.39/0.46 0.36/0.40/0.45 0.34/0.37/0.42
a
γ-Glutamyl transferase (μkat/L) 0.19/0.25/0.35 0.20/0.25/0.36 0.20/0.25/0.37 0.19/0.25/0.37
C-reactive protein (mg/L) 0.8/1.5/3.0 0.6/1.3/3.4 0.7/1.6/3.8 0.8/1.7/4.0
Total cholesterol (mmol/L) 4.35/4.87/5.44 4.45/5.11/5.87
a
4.45/5/50.55 4.49/5.15/5.83
TAG (mmol/L) 0.78/1.11/1.49 0.78/1.07/1.39 0.86/1.12/1.45 0.88/1.15/1.41
HDL (mmol/L) 1.37/1.66/1.88 1.39/1.74/2.01
a
1.42/1.69/1.92 1.47/1.72/1.94
Cholesterol/HDL 2.43/3.10/3.55 2.56/3.04/3.67
a
2.59/2.96/3.65 2.53/2.89/3.83
LDL (mmol/L) 2.20/2.72/3.34 2.25/2.95/3.54
a
2.29/2.78/3.53 2.30/2.89/3.60
Glucose (mmol/L) 4.4/4.8/5.3 4.5/4.9/5.3 4.5/4.8/5.3 4.5/4.9/5.4
a
The value was significantly different from the value in day 0 (p<0.05). The data are expressed as first quartiles, medians and third quartiles.
b
The change-from-baseline value is significantly different from that in the placebo group (p<0.05).
Copyright © 2015 John Wiley & Sons, Ltd. Phytother. Res. 29: 1559–1567 (2015)
1564 J. VOSTALOVA ET AL.

Table 6. Concentration of free and total phenolics in urine of the placebo and cranberry groups on day 0 and 180
Compound
Placebo group (n= 86) Cranberry group (n= 79)
Concentration of analysed compounds (μg/g creatinine)
Day 0 Day 180 Day 0 Day 180
Free Total Free Total Free Total Free Total
BA 0/0/147 194/809/2552 0/0/548 276/829/2225 0/0/452 192/682/2240 0/0/497 86/472/1415
2-HBA 0/0/0 0/114/418 0/0/0 0/111/313 0/0/0 0/77/583 0/0/0 0/75/290
3-HBA 0/0/318 0/0/360 0/0/176 0/0/251 0/0/205 0/0/138 0/55/164 0/8/175
4-HBA 188/519/987 927/2049/4047 164/335/834 735/1759/3812 144/307/781 783/1572/3228 106/248/685 561/1133/2529
3,4-DHA 88/232/543 228/484/895 57/245/465 182/448/757 34/160/404 216/388/821 92/172/319 153/335/543
3,4,5-THBA 0/0/440.6 0/103/556 0/253/694 0/56/502 0/33/419 0/56/481 0/124/352 0/16/391
4-HMBA 70/249/619 620/1961/4210 61/253/564 554/1382/3281 80/172/365 495/1378/3023 36/127/426 357/1058/2262
PA 0/0/0 1052/6739/21 614 0/0/0 1145/4717/17680 0/0/0 1025/3646/12 697 0/0/0 767/3283/14 550
2-HPA 310/732/1120 293/761/1100 281/625/1124 288/652/1020 251/577/966 288/602/881 201/576/1033 223/550/922
3-HPA 457/1115/2852 405/1199/3057 451/1004/2263 479/995/2938 407/992/2072 402/1139/3395 457/945/2008 370/875/2341
4-HPA 7077/11 913/21 062 9449/18 265/30 077 5490/11 307/16 897 8317/15 744/24 520 4790/9848/19 841 7965/16 515/27 960 4953/9276/16 407 6427/12 397/20920
3,4-DHPA 1324/2998/6284 1702/3612/6979 1182/2547/4594 1612/3198/5797 1084/2632/4725 1251/3414/6329 1321/2479/3992 1054/1964/4512
a
4-HMPA 2118/3575/7794 1962/3190/7136 1118/2920/5366 1419/3265/6413 1359/2845/4705 1454/3457/6273 1074/2189/4253 1435/2502/5171
2-HPPA 0/77/292 120/359/781 0/106/288 99/286/794 16/92/239 73/268/804 27/70/213 71/192/574
3-HPPA 0/0/0 0/0/803 0/0/1548 0/0/1072 0/0/1724 0/0/746 0/0/1172 0/0/1213
3,4-DHPPA 199/422/1060 385/877/1557 146/444/1038 296/780/1778 153/411/1026 339/662/1615 191/445/675 391/697/1169
4-HMPPA 0/293.8/1230 566/1282/2644 0/232/1066 480/1217/2440 0/148/702 367/1071/2250 0/141/435
a
265/808/2103
3-HCA 0/43/122 27/99/260 0/47/180 22/115/257 0/34/126 21/79/256 0/25/87 25/82/231
4-HCA 0/24/72 20/81/155 0/19/61 31/77/179 0/0/48 36/82/228 0/21/66 18/59/135
3-HMCA 0/66/199 138/414/944 0/55/166 127/353/950 0/61/151 111/463/1009 0/43/129 126/352/834
4-HMCA 0/0/121 311/954/2109 0/19/146 215/546/1595 0/0/85 230/845/1948 0/8/90 190/630/1684
HA 105/248/466 102/243/458 109/2274/443 979/221/416 903/203/397 101/210/396 108/198/388 968/1979/353
2-HHA 62/149/285 69/131/229 53/150/385 62/144/338 40/130/262 50/130/262 57/129/259 48/129/254
QUE 0/0/0 0/0/840 0/0/0 0/96/1092 0/0/0 0/0/919 0/0/0 0/183/878
The values are expressed as expressed as first quartiles, medians and third quartiles.
BA, benzoic acid; 2-HBA, 2-hydroxybenzoic (salicylic) acid; 3-HBA, 3-hydroxybenzoic acid; 4-HBA, 4-hydroxybenzoic acid; 3,4-DHA, 3,4-dihydroxybenzoic (protocatechuic) acid; 3,4,5-THBA,
3,4,5-trihydroxybenzoic (gallic) acid; 4-HMBA, 4-hydroxy-3-methoxybenzoic (vanilic) acid; PA, phenylacetic acid; 2-HPA, 2-hydroxyphenylacetic acid; 3-HPA, 3-hydroxyphenylacetic acid;
4-HPA, 4-hydroxyphenylacetic acid; 3,4-DHPA, 3,4-dihydroxyphenylacetic acid; 4-HMPA 4-hydroxy-3-methoxyphenylacetic (homovanilic) acid; 2-HPPA, 2-hydroxyphenylpropanoic acid;
3-HPPA, 3-hydroxyphenylpropanoic acid; 3,4-DHPPA, 3,4-dihydroxyphenylpropanoic (dihydrocaffeic) acid; 4-HMPPA, 4-hydroxy-3-methoxyphenylpropanoic (dihydroferulic) acid; 3-HCA,
3-hydroxycinnamic acid; 4-HCA, 4-hydroxycinnamic (p-coumaric) acid; 3-HMCA, 3-hydroxy-4-methoxycinnamic (isoferulic) acid; 4-HMCA, 4-hydroxy-3-methoxycinnamic (ferulic) acid; HA,
hippuric acid; 2-HHA, 2-hydroxyhippuric (salicyluric) acid; QUE, quercetin.
a
The value was statistically significant at p<0.05.
Copyright © 2015 John Wiley & Sons, Ltd. Phytother. Res. 29: 1559–1567 (2015)
1565ARE HIGH PAC KEY TO CRANBERRY EFFICACY IN THE PREVENTION RUTI?

in the current study, the design was robust (randomized,
double-blind, placebo-controlled), and the sample size
was sufficient and justified. There was a relatively low
rate of subject attrition, criteria appropriate for the diag-
nosis of a symptomatic UTI were applied and a well-
characterized cranberry product was used. To the best
of our knowledge, this is the first study demonstrating
the efficacy of a well-characterized whole cranberry
fruit in the prevention of rUTIs in women. We found
no PACs in the plasma or urine samples, and there
was no significant difference in the phenolic compound
profile or benzoic acid derivatives in the urine samples
of the women from either group on days 0 and 180. Of
the phenolics determined, hippuric acid dominated.
It can be speculated that the increased urinary
antiadherence and lower incidence of UTIs are con-
nected to other cranberry constituents apart from PACs,
anthocyanins, phenolic acids, flavonoids and their
microbial-derived metabolites (de Llano et al., 2015).
The pentacyclic triterpenoids, mainly ursolic acid, may
play a complementary or synergistic role together with
polyphenolic constituents in the antiadhesion activity
of cranberry fruit (Vasileiou et al., 2013). For example,
this compound caused differential gene expression in
E. coli and inhibited biofilm formation in several bacte-
rial species (Ren et al., 2005). Ursolic acid has been
shown to affect P fimbriae and the curli fibre morphol-
ogy of uropathogenic E. coli strains and their adhesion
to uroepithelial cells (Wojnicz et al., 2013). Also, some
metabolites are formed through the action of intestinal
microflora, which is unique for each individual
(Cardona et al., 2013). This might explain individual
sensitivity to the effects of cranberry.
CONCLUSION
In summary, results of this study showed that intake of
500 mg of cranberry fruit powder containing 2.8 mg of
PACs/day for 6 months was associated with a reduction
in incidence of recurrent UTIs. The compliance with
the study protocol was excellent and no adverse events
were recorded. From the results, it is not possible
to pinpoint which compound/compounds in CFP
protected the epithelium of the urinary tract against
the formation of bacterial biofilm. Our data nonethe-
less provide encouraging evidence for the protective
effect of whole cranberry (peel, seeds, pulp) in women
with a medical history of rUTIs. This effect is possibly
due to the synergy of all cranberry components and/or
its metabolites rather than just PACs. However, addi-
tional studies are needed to determine which cranberry
secondary metabolites in addition to PACs are respon-
sible for the effects found.
Acknowledgement
Financial support of Palacky University, Olomouc, is gratefully
acknowledged.
Conflict of Interest
The authors have declared that there is no conflict of interest.
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