Midlife women, bone health, vegetables, herbs and fruit study. The Scarborough Fair study protocol.

0.31 MB·English

by  Gunn, Caroline A

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STUDY PROTOCOL Open Access
Midlife women, bone health, vegetables, herbs
and fruit study. The Scarborough Fair study
protocol
Caroline A Gunn*, Janet L Weber and Marlena C Kruger
Abstract
Background: Bone loss is accelerated inmiddleaged women but increased fruit/vegetable intake positively affects
bonehealth byprovision of micronutrients essential for boneformation,buffer precursors which reduce acid load
and phytochemicals affecting inflammation and oxidative stress.Animal studies demonstratedbone resorption
inhibiting properties of specific vegetables, fruit and herbsa decadeago.
Objective: Toincreasefruit/vegetable intake inpost menopausal women to 9 servings/day using a food specific
approach to significantly reduce dietary acidload and include specific vegetables, fruit and herbs withbone
resorbing inhibitingpropertiesto assess effect onbone turnover, metabolic and inflammatory markers.
Methods/Design:The Scarborough Fair Studyis a randomised active comparator controlledmulti centre trial.It
aimed to increasefruit and vegetable intake in 100post menopausal women from ≤ 5servings/day to ≥ 9
servings/day for 3months. The women inthe dietary intervention were randomly assigned to one of thetwo arms
ofthe study. Both groups consumed ≥9 servings/day offruit/vegetables and selected herbsbut the diet of each
group emphasiseddifferent fruit/vegetables/herbswith one group (B) selecting from a range ofvegetables, fruit
and culinary herbs with bone resorbing inhibitingproperties.50 womenformed a negative control group (Group C
usual diet).
Primary outcomevariableswere plasma bone markers assessed atbaseline,6 weeks and 12 weeks. Secondary
outcome variables were plasma inflammationand metabolic markersand urinary electrolytes (calcium, magnesium,
potassium and sodium) assessed atbaselineand 12 weeks. Dietary intake and urine pH change also were outcome
variables. The dietary change was calculated with 3day diet diaries and a 24 hour recall. Interventionparticipants
kept a twiceweeklyrecord of fruit, vegetable and herb intake and urine pH.
Discussion: Thisstudy will provide information onmidlife women’s bonehealth and how a dietary intervention
increasing fruit and vegetable/herb intake affects bone, inflammatoryand metabolic markersand urinary electrolyte
excretion. It assesses changes innutrient intake, estimated dietary acid load and sodium: potassium ratios. The
study also explores whether specific fruit/vegetables and herbs withboneresorbing properties has aneffect on
bonemarkers.
Trial registration: ACTRN12611000763943
Keywords: Bone, Osteoporosis, Postmenopausal, Fruit, Vegetables and herbs, Net endogenous acid production,
Inflammation, Phytochemicals
*Correspondence:[email protected]
InstituteofFood,NutritionandHumanHealth,MasseyUniversity,PrivateBag
11222,PalmerstonNorth4442,NewZealand
©2013Gunnetal.;licenseeBioMedCentralLtd.ThisisanOpenAccessarticledistributedunderthetermsoftheCreative
CommonsAttributionLicense(http://creativecommons.org/licenses/by/2.0),whichpermitsunrestricteduse,distribution,and
reproductioninanymedium,providedtheoriginalworkisproperlycited.
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Background the age of 30 years, accelerating at menopause to lower
Osteoporosis meaning “porous bone” is the term for bone strength and mineral density [14,16-18]. F/V’s in-
inadequate bone mass. It is a global problem seen most fluence on acid–base balance is crucial as the sole die-
often in the elderly and in women (80%) [1] and is con- tary source of alkaline precursor constituents and is an
sidered one of the ten most important diseases affecting important reason to recommend increased consumption
theworld’spopulation [2]andisparticularly prevalent in duringageing toforestall bone loss[19,20].
developed countries with ageing populations and longer Additional benefits on bone metabolism ensue from
life spans [3]. Bone loss is accelerated at early meno- bioactive constituentsfound predominantly invegetables
pause resulting in increasingly fragile bones prone to but also some herbs and fruit. Phytochemicals, antioxi-
breakage. Inflammation also increases with age and dants and other bioactive compounds influence bone
exacerbatesboneloss [4-6]. metabolism through a variety of mechanisms [21-25]
Osteoporosis poses a significant health and economic particularly in reducing inflammation and oxidative
burden for New Zealand families and the public health stress [26,27]. This pharmacological effect on bone re-
system. Thenumber ofolder (> 50years) New Zealanders sorption was first observed a decade ago by Muhlbauer
is increasing steadily and the cost of treating fractures [28,29] who, in precise and controlled conditions with
and secondary illnesses related to osteoporosis is animals, demonstrated specific vegetables, herbs and
expected to rise from $330 million in 2007 to $458 mil- fruit positively affected bone resorption quite apart from
lion by 2020 [3]. effects on diet acid load. Muhlbauer determined the
Fruit and vegetables (F/V) are positively associated effect was additive, therefore, the more of this range
with bone status. The beneficial effect is thought to be consumed, the more bone resorption reduced. This
through provision of micronutrients potassium, magne- effect has previously been shown only in the animal
sium, calcium, vitamins A, C, E and K, and potentially a model.
lowerdietary acidloadconferredbythefruitandvegeta- Intervention studies with mid life women assessing
bles food group [7-9]. Typical western diets are acidic acid load and bone health have been limited to modest
because predominantly acid (hydrogen ions) rather than increases in self selected fruit and vegetables [8,30], use
base (bicarbonate) is created during the metabolism of of supplements [8,31] or use of alkaline water [32,33],
the daily food intake. Acid forming grains and high pro- mimicking F/V alkali forming effect. No study has
tein food derived from animal origin (meat, fish and increased F/V intake to significantly affect NEAP or spe-
eggs) contain sulphur based amino acids, methionine cified daily intake of vegetables, herbs and fruit shown
and cysteine which create acid when metabolized. Alka- in the animal model to have bone resorption inhibiting
line forming foods contain potassium salts which can be properties. A diet high in F/V and including some from
broken down to make alkaline buffers [10]. Vegetables thisrangeof vegetables,herbsandfruitcouldbeauseful
and fruit are considered alkaline because of their high dietary strategy to ameliorate bone loss particularly at
mineral content in the form of salts of organic acids. criticaltimessuch asmenopause.
The salts, predominantly potassium based but also cal- Despite the numerous reports in the literature attri-
cium and magnesium, generate bicarbonate to balance buting health benefits with increased consumption of
theacidproducedfrom therestofthediet. F/V and improvement in chronic disease risk factors
Western diets are low in F/V and high in grains and [22] most New Zealanders don’t reach the Ministry of
animal protein compared to the typical diet of early Health (M.o.H.) target of 2 servings of fruit and 3 ser-
man. The change from plant based diets to modern, vings of vegetables every day[34,35].
western diets characterized by foods that are acid rather It is hypothesised that an increase in vegetable and
than alkaline forming results in a low grade systemic fruit consumption to ≥ 9 servings/day will reduce the
metabolic acidosis [11-13]. The level of acidity created estimated Net Endogenous Acid Production (NEAP) by
can be estimated from the dietary intake. A significant approximately 20 mEq/day and result in reduction in
change in estimated net endogenous acid production bonemarkersofresorption Ctelopeptideoftype1colla-
(est.NEAP)issaidtohaveoccurredfrompreagricultural gen (CTx) and bone formation marker Procollagen 1
times (−88mEq/d) to today (+ 48 mEq/d) [13]. The N-terminal peptide (P1NP) in post menopausal women,
chronic, low grade metabolic acidosis induced by the and those women who include 4–5 servings of vegeta-
modern, western diet is exacerbated during ageing when bles, herbs and fruit with bone resorption inhibitory
renal function begins to decline [14,15] requiring the properties (BRIPs) as half of the 9 servings/day will
body’s skeletal reserves to be called upon to relinquish reduce resorption marker CTx by a greater amount. It is
bicarbonate to produce alkaline buffers needed to con- alsohypothesised thatthisincreaseinfruitandvegetable
tinuously balance the acid load. This results in bone intake will significantly affect inflammatory and meta-
mass that is worn away gradually and indefinitely after bolic markers including: c-reactive protein (CRP),
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adiponectin, interleukin 6 (IL-6), interleukin 10 (IL10), Methods/Design
tumour necrosis factor (TNF), triglycerides, cholesterol, Figure 1 illustrates the study design. This study is a ran-
fibrinogen and plasminogen activator inhibitor-1 (PAI-1). domized active comparator controlled intervention to
This study therefore aims to investigate the effect of increase fruit and vegetable intake in healthy postmeno-
increased fruit, vegetables and herbs on bone, meta- pausalwomen overa3monthperiod.
bolic and inflammatory markers and whether including
specific fruit, vegetables and herbs with BRIPs [28] as Samplesize
part of an increased fruit/vegetable intake has any add- The number of subjects required in each group was cal-
itional effect. culated to be 32 (minimum). This was determined using
Scarborough Fair study design
150 women (≥5 yrs PM) recruited through advertisements/fliers
inlocal newspapers/magazineand workplaces.
Phone/email inquiry from potential participants/.Information
sheet emailed/posted.
Phone response from women -clarification of queries
Screening questionnaire administered
Participants eligible for study emailed/postedconsent formsand 3
Day Diet Diary informationand instructions for first visit.
Week 1 of study
1stvisit to Human Nutrition Research Unit for both intervention and control groups
Randomisation of intervention group into group A and B
Double check consent form signed and any queries answered
Fasted blood sample taken between 0700 and 1000hrs
( light breakfast provided)
Questionnaire regarding usual diet, lifestyle and nutritional knowledge
Dietary assessment (3 Day Diet Diary) reviewed with nutritionist (food portion size atlas)
Studydietary requirements reviewed with participants in intervention with demonstration
of serving sizes and how to fill in weekly diary.
Anthropometric tests: weight, height, blood pressure, spot urine pH.
DEXA scan performed (first or second visit)
Participants willing to provide a 24 hour urine collection are given container with
instructions (verbal and written). Pickup of24 hour urine specimen
Researcher emailed participants fortnightly with general answers to any queries, tips,
recipes etc appropriate to each group in the intervention arms of the study.Participants
could email the researcher with a query and receive a prompt response (within 24 hours).
Week 6
Participants in the intervention arms of the study attend the clinic again for a fasted blood
sample (0700 and 1000hrs) and for a 24 hour dietary recall with nutritionist.
Week 11
All participants contacted to complete second 3 Day diet diary tobring completed to theirclinic visit
the following week. Those who volunteered a first 24 hour urine collection reminded to commence
another one 24 hours prior to attending final clinic visit.
Week 12/13
Final visit to Human Nutrition Research Unitfor blood sample (fasted), 24 hr urine collection
(light breakfast provided).
Anthropometric tests: weight, blood pressure, spot urine (pH). 3 DDD reviewed with nutritionistand
final questionnaire for all participants.
Figure1ScarboroughFairstudydesign.
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a power calculation based on demonstrating a difference Choices medical centre in Hastings. Participants were
of ~8% in the primary outcome variable C-telopeptides recruited using 2 different fliers. One flier recruited 100
of collagen (CTx) with 80% power and alpha of 0.05 (2 women to form the intervention group and be rando-
sided test) and accepting 0.4μg/ml as mean CTx of this mised to one of two groups (A or B) within the inter-
population (26). To detect any differences between the 2 vention to increase intake of fruit and vegetables to
diets and allowing for withdrawals, non-compliance or 9 servings/day. The other flier recruited 50 women
maintenance (~ 25%) approximately 50 women were (Group C) who were willing to have their bone, inflam-
needed in each group. Since there were 2 different diets matory and metabolic markers tested on two occasions
emphasizing different vegetables and fruit and a control 3 months apart ( baseline and end of study) and who
group who consumed their usual diet (≤ 5 servings F/V/ would continue eating their usual diet. This negative
day), threegroups of50participantswere required. control group was called the Diet and Metabolic Markers
group(DMM)andreferredtointhisprotocolasGroupC.
Inclusion/Exclusioncriteria Because of the motivation and commitment involved, it
The target population were healthy, post menopausal was considered preferable to recruit a negative control
(≥ 5yrs) women between 50–70 years. Women were group of women separately rather than randomising
included if they were taking some medications e.g. womentoacontrolgroupwhentheywereattractedtothe
hypertensive tablets, thyroxine (if thyroid function study because of a conscious decision to participate in the
stable) and diuretics other than potassium sparing but dietary change. The same exclusion and inclusion criteria
excluded if on medication for diabetes, heart disease, applied to the control group apart from the requirement
osteoporosis (including hormone replacement therapy) fordietarychange.
or medication that could affect bone or calcium metab- The study was advertised in local newspapers in the 3
olism (oral corticosteroids, warfarin, dilantin. potassium centres, in a few workplaces and in a small advertise-
sparing diuretics and regular use of proton pump inhibi- ment on the health page of The Listener (a popular na-
tors). Regular use of NSAIDs including aspirin was not tional magazine) over July/August 2011.This advertising
permitted as they could interfere with anti-inflammatory and word of mouth returned a good response rate
markers. If participants had stopped use of a NSAID 1 (> 350 enquiries) with enquiries mainly to participate in
month prior to study commencing they were included. the dietary intervention rather than the negative control
Women were also excluded if they had any of the fol- study (Group C). Recruitment was completed within 6
lowing conditions: osteoporosis previously diagnosed, weeksoffirstadvertising.
both hips replaced, previous fractures ofthe lower verte-
bra or hip, severe osteoarthritis* of the lower spine or Screening
hips, gastrointestinal, liver or renal disease and any se- Prospective participants who phoned or emailed expres-
vere* disease including treatment for cancer within the sing an interest in the study were initially sent out the
last 3 years. Women who smoked, drank more than 20 appropriate detailed information sheet for perusal. If
standard drinks/week or were already consuming > 6 they replied (email/ phone) willing to participate, a
servings fruit and vegetables every day, or were taking screening questionnaire was completed over the phone.
calcium supplements and unwilling to stop a month be- This questionnaire included demographics, health status
fore the study for the duration of the study were (including medications) and biographic information.
excluded. Any participant who developed an illness du- Over 300 women were screened with over half being
ring the study that required treatment with steroids or declined due to a significant health issue or on medica-
medication that affected bone, inflammatory and other tion deemed incompatible with the study e.g. regular use
metabolic markers was also excluded. The intervention ofprotonpumpinhibitors.
group participants had to be willing to increase their in-
take of fruit and vegetables to 9 servings/day and the Randomisationandblinding
negative control group willing to continue their normal Participants were stratified according to the 3 cities and
diet. randomly allocated to either group (A or B) using block
*Severedefinedasrequiringdailypainrelief. randomization. The random allocation sequence was ge-
nerated by administrative personnel (not the researcher
Settingandrecruitment who recruited participants) and intervention group parti-
This was a multi-centre trial, with 50 participants at cipants were assigned to group A or B as they arrived for
each trial site in Hawke’s Bay, Palmerston North and theirfirstappointment.
Auckland.ThestudywasconductedatMasseyUniversity’s As intervention participants were required to source,
clinical nutrition research units in Palmerston North and store, prepare and consume specific vegetables, fruits
in Albany, Auckland. Hawke Bay participants attended and herbs they were not blinded to which diet (A, or B)
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they were on once randomisation had occurred, but vegetable in their diet every day. They were also asked to
were blinded to which group contained the fruit, vegeta- have at least 2 servings of dairy or calcium enriched soy
bles and herbs with BRIPs. The randomization codes milkorotheralternativetocontrolforcalciumintake[35].
were maintained by an alpha numeric added to the par- Twice a week the intervention group women recorded
ticipant’s unique identifier and laboratory staff involved their total vegetable, fruit and herb intake as well as their
inthe analysis of blood orurinesampleswereblinded to urinepH(fasting,secondvoid)to thenearest0.25phunit
participants’group. with dipsticks provided (PHion Diagnostic Test Strips,
ApexWellnessGroup,UC,Scottsdale,AZ85260,USA).
Theinterventionrationale
Women in the intervention groups A and B were asked GroupA≥9servingsoffruit and vegetables (≤3
to consume a minimum of 9 servings a day of fruit and servings fruitand ≥6servings of vegetables)Specific
vegetables with herbs additional. 9 servings per day was fruit/vegetables/herbs (allnonBRIPs)specified for
chosen as the target number of fruit and vegetable ser- overhalf theservings andavoidingF/V/Hwith BRIPs.
vings as it allows for a significant decrease in estimated GroupB≥9servings offruit and vegetables (≤3
net endogenous acid production (NEAP) of the diet servings fruitand ≥6servings of vegetables)Specific
(~20 mEq/day unpublished feasibility study) and was the fruit/vegetables/herbs (allBRIPs)specified foroverhalf
number of servings recommended by the USDA for theservingsand avoiding someofGroupA’sspecified
healthy, fit 50 year old women [36] as well the number fruit/vegetables/herbs.
recommended in the DASH Trial [37]. This increase in GroupCNegative controlgroupwhoconsumetheir
fruit and vegetables could then be compared to a group usualdiet
of 50 women consuming their normal diet (≤5 servings/
day). Bloodandurinesampling
In order to assess the impact of BRIPs both groups Procedures for taking blood, urine and anthropometric
were asked to emphasise a different selection of vegeta- measurements were standardized to reduce errors and
bles, herbs and fruit and avoid others. Group B include variability. Bone markers show circadian variability [43]
4–5 servings of specific vegetables, fruit and herbs with therefore all fasted blood samples were taken between
bone resorption inhibitory properties (BRIPs), while 0700 and 1000hr with blood drawn by certified phlebo-
Group A avoided those specific vegetables, herbs and tomists. After their first appointment women were
fruit and emphasised other specific fruit, vegetables and advised to make their subsequent appointments as close
herbs. To allow for change in bone turnover markers as possible to the exact time of day as their first and
particularly P1NP, the minimum study period was deter- most were able to comply with this. Plasma was used for
minedtobe3months[38]. analysis of bone, inflammatory and metabolic markers
Boneresorptioninhibitingpropertieshavebeenascribed and drawn into vacutainers containing EDTA, citrate or
to a limited range of common fruit, vegetables and herbs. heparin.After centrifugation at 3000 rpm for 15 minutes
These include the herbs dill, sage, garlic, parsley, thyme (4°C), the plasma was dispensed in aliquots and frozen
androsemary.VegetablesandfruitwithBRIPsincludeto- at −80°C. Participants collected a 24 hour urine sample
matoes, green beans, cucumber, broccoli, lettuce, prunes the day prior to attending their clinic visits. At the clinic
and oranges [28]. The minimum effective dosage of fruit/ the sample quantity was measured and 3 aliquots of 100
vegetablesandherbs(F/V/H)withBRIPscalculatedinthe ml were frozen immediately at −20°C. All blood and
animal model is 170 mg/day. This corresponds with 6.2 urine samples were analysed at the end of the 3 month
grams of fresh F/V/H per kilogram human body weight. study period. To reduce inter-assay variability both base-
Thisamountisequivalentto3–5servings/day[35]ofany line and end of study samples were analysed in the same
F/V with BRIPs for a 60–70 kilo woman. Culinary herb assay run.
servingsweretobeadditionalandusualculinarymeasures
inmealswereadvisedtoallinterventionparticipants(2–3 Biochemicalanalysis
clovesgarlic, 0.5-1teaspoonfordried/freshculinaryherbs Refer Table1fordetails of biochemicalanalysis done.
and up to 0.25 cup of parsley (Group B-BRIPS) and basil
(Group A- non-BRIPS). The effect on bone resorption of Dietaryassessment
thefoodswithBRIPsissaidtobeadditive[39].VitaminK 3 Day Diet Diaries (3DDDs) were done at baseline and
is also known to affect bone health and fracture risk end ofstudy (week12). Participantsreceived writtenand
through its action on cytokines and the gamma carboxyl- verbal instruction on how to complete the 3DDD’s via
ation of the bone protein osteocalcin [40-42]. To control email and phone call prior to attending their first clinic
for vitamin K intake all participants in the intervention appointment. They were asked to record all food and
were asked to include one serving of a leafy green beverages consumed over 2 weekdays and 1 weekend
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Table1Outcomevariablesandanalysismethod
OutcomeVariables Baseline Mid6 End Method
wks (3mths)
CTxboneresorption I&C I I&C RocheElecsys2010.RocheDiagnostics,CanterburyHealthEndocrinelaboratory(CHL).
marker AccreditedwithIANZtotheISO15189
P1NPboneformation I&C I I&C RocheElecsys2010.RocheDiagnostics,CanterburyHealthEndocrinelaboratory.
marker
Cholesterol I&C I&C CHLEnzymaticCholesterolOxidaseAbbottc8000analyser.Abbottreagents
Triglycerides I&C I&C CHLEnzymatichydrolysisofTriglycerides.Abbottc8000/c16000analyser,Abbottreagents.
LDL I&C I&C CHLcalculatedasfollows:LDLChol=TotalChol-HDLChol-(Trigs/2.22)mmol/L.
HDL I&C I&C CHLEnzymaticassay.Abbottc8000analyser.Abbottreagents
Fibrinogen I&C I&C Fibrinolysisreferenceplasma(Cat.No.T6010)(Kordia,Leiden,TheNetherlands).
PA1 I&C I&C PlasmaPAI-1 bio-immunoassay,TriniLIZEPAI-1activityassaykit(Cat.No.T6004)
act
CRP I&C I&C PlantandFoodResearchAucklandFlowCytomixkits(BenderMedsystems,Austria)FC500
MPLflowcytometer(BeckmanCoulter,Miami,FL
IL6 I&C I&C Asabove
IL10 I&C I&C ‘’
TNFalpha I&C I&C ‘’
OPG I&C I&C ‘’
Adiponectin I&C I&C ‘’
24hrCalcium I&C I&C E.I.TResearchLaboratory-ShimadzuAA-6200AtomicAbsorptionFlameEmission
Spectrophotometerwithair/acetyleneflame.(MethodadaptedfromGimbletEl)
24hrMagnesium I&C I&C AASasabove(MethodadaptedfromGimbletEl)
24hrSodium I&C I&C AASasabove(MethodadaptedfromCastenmillarJM)
24hrPotassium I&C I&C AASasabove(MethodadaptedfromCastenmillarJM)
24hrCreatinine I&C I&C MasseyUniversityNutritionlaboratory.JaffeMethodFlexorE,VitalScientificNV,6956AV
Spankeren/Dieren,TheNetherlands
Iodine I&C I&C HillsLaboratory-inductively-coupledplasmamassspectrometry.
Selenium I&C I&C HillsLaboratory-inductively-coupledplasmamassspectrometry
Dietaryassessment
3DDDmacroand I&C I&C 3dayfoodintake(2weekdaysand1weekend)Recipes/packaging.
micronutrientintake
FoodworksNZ,2009(Xyrissoftware)
24hourdietaryrecall I Previousday24hourrecallsecondclinicvisitnutritionist(NZreg.)
EstimatedNEAPand I&C I&C Derivedfromalgorithms([44])andusingFoodworks(Xyris,NZ)datafrom3DDDs
PRAL
No.serveseachfood I&C I&C Derivedfrom3DDDnutritionist(NZreg.)convertedquantitiesingrams/mls/cupsetc.to
groups servings([35]
F/V/HerbDiary Interventiongrouponly.Twiceweekly-selfreportedintake
Urineph(2/wk) InterventiongroupTwiceweeklyselfreportedurinepH(secondvoid/fasting)PhionDiagnosticTestStrips(0.25
graduations),ApexWellnessGroup,UC,Scottsdale,AZ85260,USA
Weight/heightBMI I&C I&C DigitalscalesUWEGilbarco,NZ/Stadiometer/BMI=weight/height2
Bloodpressure I&C I&C OmronDigitalHEM-907automaticbloodpressuremonitor(duplicates)highvaluesrepeated
manually
Waist/hip I&C I&C Anthropometrymethodsperformedbytrainednutritionstaffusingstandardisedprocedures
measurements andequipment.Tapemeasure,DouglasPharmaceuticalsLimited,Auckland.
Bodycompositionand I&C DEXAHologicDiscoveryQDR4500Adensitometer,HologicInc.Bedford,Massachusetts
BMD
Intervention(I)andcontrolgroup(C).
day, including types, brands and amounts (cups, table- registered nutritionist at the first visit. Prompting me-
spoons, etc.) of foods as well as recipes for homemade thodswereusedforincompletequantitiesortoascertain
dishes. Participants supplied nutritional information specific foodtypese.g.milk(skimorfullfat).
panels from processed food packets. All 3DDD’s were A24 hour recall (unexpected) wasdone with interven-
checked for accuracy and completeness by a NZ tion participant’s midway through the study at the
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second clinic visit. Dietary data were entered into Food- 2010) and was said by some participants to be a
works(version2009,NZ,Xyrissoftware)byaM.Sc(nutri- motivator.
tion) graduate, checked for accuracy and completenessby
aN.Z.registerednutritionistandthentransferredtoSPSS Adverseeventreporting
(20)viaMicrosoftAccess(2007)andExcel(2007). Participants were advised to report any side effects from
New Zealand dietary reference values were used to the increase in vegetables and fruit intake. They also
assess all nutrient intakes [35]. Net endogenous acid pro- recorded in their diaries any incidence of adverse side
duction(NEAP)wascalculatedaccordingtothealgorithm effects e.g. stomach upset, which was reviewed at each
fromRemerandManz[44,45]. clinic visit. The study’s clinical collaborator was available
NEAP = Potential Renal Acid Load (PRAL) + Organic to provide guidance on medical issues. Participants were
Acid(OA) advisedtoreportimmediatelytheirconcernswithanyser-
PRAL represents the average intestinal absorption ious adverse side effect from this dietary regimen e.g.
rates of ingested protein and additional minerals. OA chronic diarrhoea(>48hours). Any participant recording
is an anthropometry-based estimate for organic acid serious adverse effects due to the diet would have been
excretion. immediately advised to consult their medical practitioner
Thetwo componentsofNEAParegiven asfollows: for assessment if they had not already done so and with-
PRAL (mEq/d) = 0.49 protein (g/d) + 0.037 × phos- drawnfromthestudy.
phorous (mg/d) - 0.021 × potassium (mg/d) – 0.026 × Follow-up: The researcher/clinical collaborator would
magnesium(mg/day)-0.013×calcium(mg/day) follow-up with any participant that had a medical issue,
OAest (mEq/day) = individual body surface area × 41/ that they had received medical treatment and the issue
1.73 was resolved satisfactorily. In the unlikely event of a ser-
Body surface area was calculated according to the ious adverse side effect from participation in this inter-
methodofduBoisandduBois vention study, the individual after consultation with
BSA =(W0.425×H0.725)×0.007184 their medical practitioner would have been advised to
(Website http:/www-users.med.cornell.edu/~spon/picu/ contact ACC (Accident Compensation Corporation). No
calc/bsacalc.htm) significant adversesideeffectshowever,werereported.
NEAP and PRAL are expressed in milliequivalents
(mEq) as ions in solution interact according to their Statisticalmethods
charge. Milliequivalents is obtained by converting milli- SPSS20andMinitab16willbeused.Repeatedmeasures
grams (mg) to millimoles (mmol) and multiplying by and one way analysis of variance (ANOVA) will be done
valencestogive charge quantities[44] on serum bone markers (baseline, 6 and 12 weeks) to
determine changeoverthe study period,multipleregres-
Questionnaires sion and Pearson’s correlation will also be used for daily
At the first clinic visit a baseline questionnaire was F/V intakes and bone markers. Analysis of nutrient in-
administered with the intent of focusing participants on take before and during the intervention will be by paired
their motivations for this behaviour change and how t-tests and repeated measures, Main outcome variables
they would overcome common barriers to dietary will be checked for normal distribution (Shapiro-Wilk
change [46] and how they would incorporate the vegeta- test) and any data not normally distributed will be log
bles, fruit and herb increase required into their daily transformed before comparisons are made. A level of
dietary intake. Thefinal study questionnaire assessed the significance of0.05 willbeused
participant’s experience of the dietary change, both posi- The primary analysis will be the comparison of change
tive and negative aspects, including financial costs and in bone markers between the intervention groups and
lifestyle changestheymade. negative control group following the “intention to treat”
principleat6weeksand3months.Secondaryanalyseswill
Compliancediaries involve any intervention treatment effects on inflamma-
Intervention participants were provided with a weekly toryandmetabolicmarkers.Perprotocolanalyseswillalso
diary in which they recorded their intake of fruit, vege- be performed and compared with intention to treat to
tables, herbs (type and quantity). and urine pH (fasted assessbias.
second void) on 2 days each week for the duration of
the study. This diary was to aid compliance with the Participant’sresults
dietary requirements and also to see if urine pH All participants received individualised dietary assessment
increased with increased amounts of fruit and vegeta- analysisandtheirDEXAbodycompositionresultsbutnot
bles [47,48]. An increase of 0.68 pH units was previ- individual blood biochemistry or urinary test results
ously demonstrated (unpublished feasibility study, (requirementofMUHEC).
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Dietaryanalysis includes a combination of prescribed fruit, vegetables
Following input of 3DDD’s into Foodworks (2009), all and herbs previously shown to affect bone resorption in
participantsingroupC(negativecontrolgroup)received the animal model but not previously trialled in humans
an overview of their baseline three day diet diary [39], its primary emphasis on vegetables (≥ 6 servings)
analysis. This provided detailed information on macro- rather than fruit (≤ 3 servings) [49] and the inclusion of
nutrient and micronutrient intake. Due to this study herbs [24]. This differs from previous intervention stud-
involving dietary change, all intervention participants ies on bone health using the fruit and vegetable food
received both beginning and ending 3DDD results to group [8,30]. Limitations with the study include the
assess micronutrient changes associated with their element of selection bias introduced as a result of strin-
increased vegetable and fruit intake. Participants with gent inclusion and exclusion criteria. In some ethnic
queries about their nutrient analysis were invited to groups, health status is likely to be already compromised
email the nutritionist (NZ reg.) and received immediate by midlife due to chronic illness (diabetes and heart dis-
feedback. ease)andthese groups were under represented.Predom-
inantly European women formed the study population.
DEXAanalysis A high element of commitment to a behaviour/lifestyle
All participants received a letter informing them of their change was required with the need to purchase, store
DEXA bone scan result. This was reported as normal andprepareallfruitandvegetablesandherbsconsumed.
(including mild osteopenia), osteopenia or having a T There were no financial incentives and participants
score ≥ 2.5 SDs below normal. Participants with DEXA received a petrol voucher ($10/per visit) at conclusion of
results not within normal parameters for their age group thestudy.
(latter 2 groups) also received the radiologists report on Results from this study may provide further evidence
their bone scan with their DEXA t and z scores details to encourage increased consumption of fruit, vegetables
andaletter advisingthemtodiscusswith theirdoctor. and herbs with emphasis on those with bone resorption
inhibitingproperties.
Fundingandethics
Funding for this study was provided from the following Abbreviations
ACC:AccidentCompensationCorporation;AI:Adequateintake;BRIPs:Bone
organisations: Hawke’s Bay Medical Research Founda-
resorptioninhibitingproperties;BSA:Bodysurfacearea;CRP:C-reactive
tion, The National Heart Foundation, Massey University protein;CTx:Cteleopeptide(crosslinkedCterminalofαchainoftype1
Research Fund (MURF), and Amgen/GSK Clinical collagen);DEXA:DualenergyXrayabsorptiometry;EAR:Estimatedaverage
requirement;F/V:Fruitandvegetables;HRT:Hormonereplacementtherapy;
Grants Program administered by the OA-ANZBMS
IL-6:Interleukin6;IL-10:Interleukin10;MUHEC:MasseyUniversityHuman
ResearchFund. EthicsCommittee;MoH:MinistryofHealth;NEAP:Netendogenousacid
Thisstudywasapproved by Massey University Human production;NSAIDs:Nonsteroidalanti-inflammatorydrugs;NFκB:Nuclear
factorkappabinding;OA:Organicacid;PAI-1:Plasminogenactivator
Ethics Committee (MUHEC) (Southern A) Reference
inhibitor-1;P1NP:Procollagen1N-terminalpeptide;PRAL:Potentialrenal
11/11inJune2011.Itwasconductedincompliancewith acidload;RDI:Recommendeddailyintake;SDT:Suggesteddietarytarget;
the protocol approved by the Massey University Ethics TNF:Tumournecrosisfactor;3DDD:Threedaydietdiary;USDA:United
StatesDepartmentofAgriculture.
Committee (MUHEC) and will be reported according to
CONSORT guidelines. No deviation from the protocol Competinginterests
was implemented without the prior review and approval Theauthorsdeclarenocompetinginterests.
of MUHEC. Participants were informed they could with-
Authors’contributions
draw from the study at any point. All subjects for this
CGdesignedthestudywithhelpfromJWandMCK.CGdraftedthe
study were provided with a detailed information sheet manuscript.JWandMCKprovidedcommentsonandacceptedthefinal
describingthestudyandprovidingsufficientinformation versionofthemanuscript.Allauthorsreadandapprovedthefinal
manuscript.
to make an informed decision about participation before
they gavesignedconsent. Authors’information
CGisaregisterednutritionistcompletingaPhDinNutritionalScienceat
Discussion MasseyUniversity.JWisaseniorlecturerinnutritionatMasseyUniversity.
MKisProfessorandChairofNutritionalPhysiology.MKhasresearch
This study “The Scarborough Fair study” will provide expertiseinhumannutritionandhealthwithamajoremphasisonbone
detailed information about the influence of increased healthandspecialistexpertiseininvitroassessmentofbioactives,and
conductingnutritionaltrialsinhumanandanimalmodels.
fruit and vegetable intake on the bone health of middle
aged New Zealand women (50 -70years). The interven-
Acknowledgements
tion involves a significant behaviour change and this in- ThestudywasfinancedbygrantsfromHawke’sBayMedicalResearch
crease in fruit, vegetables and herbs may be expected to Foundation,TheNationalHeartFoundation,MasseyUniversityResearch
Fund(MURF)andAmgen/GSKClinicalGrantsProgramadministeredbythe
have an effect on inflammatory, metabolic and bone
OA-ANZBMSResearchFund.DrAnne-TheaMcGill,AucklandUniversity,acted
measures of health.Thestrengths ofthe studyarethat it asclinicaladvisorforthestudy.
Gunnetal.BMCPublicHealth2013,13:23 Page9of10
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Received:13December2012Accepted:7January2013 23. BasuS,MichaëlssonK,OlofssonH,JohanssonS,MelhusH:Association
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Commun2001,288(1):275–279.
24. MuhlbauerRC,LozanoA,PalacioS,ReinliA,FelixR:Commonherbs,
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doi:10.1186/1471-2458-13-23
Citethisarticleas:Gunnetal.:Midlifewomen,bonehealth,vegetables,
herbsandfruitstudy.TheScarboroughFairstudyprotocol.BMCPublic
Health201313:23.
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