<clinical_study>
  <!-- This xml conforms to an XML Schema at:
    https://clinicaltrials.gov/ct2/html/images/info/public.xsd -->
  <required_header>
    <download_date>ClinicalTrials.gov processed this data on September 20, 2023</download_date>
    <link_text>Link to the current ClinicalTrials.gov record.</link_text>
    <url>https://clinicaltrials.gov/ct2/show/NCT02974491</url>
  </required_header>
  <id_info>
    <org_study_id>37090614.6.0000.0118</org_study_id>
    <nct_id>NCT02974491</nct_id>
  </id_info>
  <brief_title>Consumption of Apple Juice in Hemodialysis Patients</brief_title>
  <official_title>Acute Consumption of Fuji Apple Juice Does Not Affect Oxidative Stress Biomarkers in Hemodialysis Patients: A Pilot Intervention Study</official_title>
  <sponsors>
    <lead_sponsor>
      <agency>Universidade Federal de Santa Catarina</agency>
      <agency_class>Other</agency_class>
    </lead_sponsor>
  </sponsors>
  <source>Universidade Federal de Santa Catarina</source>
  <oversight_info>
    <has_dmc>Yes</has_dmc>
  </oversight_info>
  <brief_summary>
    <textblock>
      The investigators hypothesized that acute consumption of Fuji apple juice (AJ) could increase&#xD;
      the antioxidant status and/or decrease the oxidative stress (OS) biomarkers, without&#xD;
      increasing serum biochemical parameters in patients undergoing maintenance hemodialysis&#xD;
      (MHD). In this pre-post pilot feasibility study, patients served as their own controls,&#xD;
      received 300 and 150 mL AJ immediately after a dialysis section, on different days, with a 3&#xD;
      week-washout period. Blood was collected at the baseline period, after 30 and 60 min of AJ&#xD;
      consumption. OS biomarkers (total antioxidant status (TAS), total oxidant status (TOS),&#xD;
      ascorbic acid, catalase (CAT), glutathione peroxidase (Gpx), superoxide dismutase (SOD) and&#xD;
      reduced glutathione (GSH) and potassium, phosphorus, uric acid, and glucose concentrations&#xD;
      were analyzed.&#xD;
    </textblock>
  </brief_summary>
  <detailed_description>
    <textblock>
      Subjects and study design&#xD;
&#xD;
      In this pre-post pilot feasibility study, subjects served as their own controls. Participants&#xD;
      were recruited from a hemodialysis clinic, located in Florianópolis' metropolitan area, Santa&#xD;
      Catarina state (Southern Brazil), from June to August 2015 with the following inclusion&#xD;
      criteria: hemodialysis treatment ≥ 3 months, age ≥ 20 years, and body mass index (BMI) ≥ 23&#xD;
      kg/m2. Exclusion criteria were allergy to apple, presence of cancer or acquired&#xD;
      immunodeficiency syndrome, kidney transplant less than 6 months before enrolling in the&#xD;
      study, taking antioxidant or nutritional supplements during the 30 days before enrollment,&#xD;
      having been hospitalized within 6 weeks before the beginning of the study, or suffering from&#xD;
      an acute illness.&#xD;
&#xD;
      Subjects were instructed to keep their usual dietary habits for the duration of the study,&#xD;
      except for avoiding intake of polyphenol-rich foods (i.e., vegetables, fruits and&#xD;
      fruit-containing products, chocolate, tea, coffee, honey, and any alcoholic beverage) within&#xD;
      2 days before and during intervention. On two different days, each volunteer consumed 300 mL&#xD;
      Fuji AJ, immediately after a dialysis section. After a washout period of 3 weeks, the&#xD;
      volunteers drank 150 mL Fuji AJ in a similar manner as described above. Before and after 30&#xD;
      and 60 min of AJ consumption, blood samples were withdrawn for biochemical analysis. This&#xD;
      protocol was chosen taking into account the previous favorable effects of AJ consumption on&#xD;
      serum OS biomarkers described for healthy volunteers.&#xD;
&#xD;
      Baseline clinical and biochemical data were obtained from medical records. This study was&#xD;
      approved by the ethics committee of human research of the Federal University of Santa&#xD;
      Catarina (UFSC) (protocol number 37090614.6.0000.0118) and all participants gave written and&#xD;
      informed consent.&#xD;
&#xD;
      Apple juice and analysis&#xD;
&#xD;
      Fuji apples were obtained from the Experimental Seasonal Fields at the Empresa de Pesquisa&#xD;
      Agropecuária e Extensão Rural de Santa Catarina' Station in the city of São Joaquin, Santa&#xD;
      Catarina, Brazil (latitude 28º 17' 39&quot;, longitude 49º 55' 56&quot; and altitude 1.415 m),&#xD;
      harvested in their commercial maturation stage during 2015's harvest. After harvest, apples&#xD;
      were stored at 4 ± 1°C. To allow for a quick and easy apple intake in large amounts, 300 mL&#xD;
      of AJ, equivalent to 3.5 apples, and 150 mL AJ, equivalent to 1.5 apples were used. Before&#xD;
      preparing the juice, the fruits were sanitized in sodium hypochlorite. Apples with peel and&#xD;
      no seeds were blended in a centrifugal juice extractor without water addition. Each dose of&#xD;
      AJ was prepared and consumed immediately.&#xD;
&#xD;
      The contents of dry matter, total soluble solids, potential of hydrogen (pH) and titratable&#xD;
      acidity of AJ were measured following official methods. The total phenolic content of the AJ&#xD;
      extracts was measured using the Folin-Ciocalteu method colorimetric. The total flavanol&#xD;
      content was estimated using p-dimethylaminocinnamaldehyde (DMACA) method. The total&#xD;
      antioxidant capacity was determined using the DPPH (2,2-diphenyl-1-picryl-hydrazyl-hydrate)&#xD;
      method. For total monomeric anthocyanin, a spectrophotometric pH differential method was&#xD;
      used. Analyses were conducted with each AJ used in the two intervention days, in triplicate.&#xD;
&#xD;
      Blood sampling and laboratory methods&#xD;
&#xD;
      In each experiment day, blood samples from participants were collected in three different&#xD;
      moments. Venous blood samples were collected using a vacuum system (Vacutainer Becton&#xD;
      Dickinson BD®, São Paulo, Brazil) into heparin, Ethylenediaminetetraacetic acid&#xD;
      (EDTA)-containing tubes or additive-free tubes. Plasma and serum were immediately obtained by&#xD;
      centrifugation (3,000 revolutions per minute (RPM), 10 min, room temperature) for the&#xD;
      measurement of uric acid, phosphorus, glucose, potassium, Total Antioxidant Status (TAS) and&#xD;
      Total Oxidant Status( TOS). In order to quantify the endogenous antioxidant enzymes, an&#xD;
      aliquot of whole blood was lysed. In order to measure reduced glutathione (GSH), an aliquot&#xD;
      of whole blood was preserved in N-ethylmaleimide (NEM). All samples were immediately stored&#xD;
      at -80°C for later analysis in duplicate.&#xD;
&#xD;
      Serum uric acid, phosphorus and glucose concentrations were determined using commercially&#xD;
      available kits (Labtest Diagnóstica SA, Lagoa Santa, Minas Gerais, Brazil) in an automated&#xD;
      multi-biochemical analyzer (Cobas Miras Plus, Roche®, Germany). Serum potassium levels were&#xD;
      measured in an automated Dimension Max System (Siemens Healthcare Diagnostics Products®,&#xD;
      Germany), according to the manufacturer's instructions.&#xD;
&#xD;
      TAS and TOS assays were measured spectrophotometrically, according to Erel's methods.&#xD;
      Ascorbic acid and GSH were determined using high-performance liquid chromatography (HPLC), as&#xD;
      previously described. The superoxide dismutase (SOD) activity was evaluated using SOD Assay&#xD;
      Kit (Sigma Aldrich®, St. Louis, Missouri, USA) according to the manufacturer's instructions.&#xD;
      Catalase (CAT) activity was determined spectrophotometrically according to the previously&#xD;
      described method. The glutathione peroxidase (GPx) activity was determined using the NADPH&#xD;
      oxidation rate method.&#xD;
    </textblock>
  </detailed_description>
  <overall_status>Completed</overall_status>
  <start_date>August 2015</start_date>
  <completion_date type="Actual">October 2015</completion_date>
  <primary_completion_date type="Actual">October 2015</primary_completion_date>
  <phase>N/A</phase>
  <study_type>Interventional</study_type>
  <has_expanded_access>No</has_expanded_access>
  <study_design_info>
    <allocation>N/A</allocation>
    <intervention_model>Single Group Assignment</intervention_model>
    <primary_purpose>Treatment</primary_purpose>
    <masking>None (Open Label)</masking>
  </study_design_info>
  <primary_outcome>
    <measure>Measuring oxidative stress biomarkers</measure>
    <time_frame>Change from baseline at 30 and 60 minutes</time_frame>
    <description>Total antioxidant status (TAS) (mmol/L) and total oxidant status (TOS) (mmol/L) was measured at the baseline period, after 30 and 60 min of acute consumption of 300 and 150 mL of Fuji apple (Malus domestica Borkh) juice. Patients were not fasted for the withdrawal of blood. This measure was taken to prevent discomfort among volunteers. Venous blood samples were collected through aseptic venipuncture on the opposite arm of the arteriovenous fistula by a qualified professional using a vacuum system into heparin or EDTA-containing tubes or tubes without additives according to established standards of clinical and biological safety.</description>
  </primary_outcome>
  <primary_outcome>
    <measure>Measuring endogenous antioxidant enzymes</measure>
    <time_frame>Change from baseline at 30 and 60 minutes</time_frame>
    <description>Catalase (U mg/Hb), glutathione peroxidase (U mg/Hb) and superoxide dismutase (U mg/Hb) was measured at the baseline period, after 30 and 60 min of acute consumption of 300 and 150 mL of Fuji apple (Malus domestica Borkh) juice. Patients were not fasted for the withdrawal of blood. This measure was taken to prevent discomfort among volunteers. Venous blood samples were collected through aseptic venipuncture on the opposite arm of the arteriovenous fistula by a qualified professional using a vacuum system into heparin or EDTA-containing tubes or tubes without additives according to established standards of clinical and biological safety.</description>
  </primary_outcome>
  <primary_outcome>
    <measure>Measuring oxidative stress biomarkers</measure>
    <time_frame>Change from baseline at 30 and 60 minutes</time_frame>
    <description>Ascorbic acid (micromole/L) was measured at the baseline period, after 30 and 60 min of acute consumption of 300 and 150 mL of Fuji apple (Malus domestica Borkh) juice. Patients were not fasted for the withdrawal of blood. This measure was taken to prevent discomfort among volunteers. Venous blood samples were collected through aseptic venipuncture on the opposite arm of the arteriovenous fistula by a qualified professional using a vacuum system into heparin or EDTA-containing tubes or tubes without additives according to established standards of clinical and biological safety.</description>
  </primary_outcome>
  <primary_outcome>
    <measure>Measuring oxidative stress biomarkers</measure>
    <time_frame>Change from baseline at 30 and 60 minutes</time_frame>
    <description>Reduced glutathione (micromole mg/Hb) was measured at the baseline period, after 30 and 60 min of acute consumption of 300 and 150 mL of Fuji apple (Malus domestica Borkh) juice. Patients were not fasted for the withdrawal of blood. This measure was taken to prevent discomfort among volunteers. Venous blood samples were collected through aseptic venipuncture on the opposite arm of the arteriovenous fistula by a qualified professional using a vacuum system into heparin or EDTA-containing tubes or tubes without additives according to established standards of clinical and biological safety.</description>
  </primary_outcome>
  <primary_outcome>
    <measure>Measuring serum biochemical parameters</measure>
    <time_frame>Change from baseline at 30 and 60 minutes</time_frame>
    <description>Potassium (mmol/L), phosphorus (mmol/L) and glucose (mmol/L) was measured at the baseline period, after 30 and 60 min of acute consumption of 300 and 150 mL of Fuji apple (Malus domestica Borkh) juice. Patients were not fasted for the withdrawal of blood. This measure was taken to prevent discomfort among volunteers. Venous blood samples were collected through aseptic venipuncture on the opposite arm of the arteriovenous fistula by a qualified professional using a vacuum system into heparin or EDTA-containing tubes or tubes without additives according to established standards of clinical and biological safety.</description>
  </primary_outcome>
  <primary_outcome>
    <measure>Measuring serum biochemical parameters</measure>
    <time_frame>Change from baseline at 30 and 60 minutes</time_frame>
    <description>Uric acid (micromole/L) was measured at the baseline period, after 30 and 60 min of acute consumption of 300 and 150 mL of Fuji apple (Malus domestica Borkh) juice. Patients were not fasted for the withdrawal of blood. This measure was taken to prevent discomfort among volunteers. Venous blood samples were collected through aseptic venipuncture on the opposite arm of the arteriovenous fistula by a qualified professional using a vacuum system into heparin or EDTA-containing tubes or tubes without additives according to established standards of clinical and biological safety.</description>
  </primary_outcome>
  <primary_outcome>
    <measure>Evaluating heigth</measure>
    <time_frame>at baseline</time_frame>
    <description>Height (meters) data were obtained from the patient's files.</description>
  </primary_outcome>
  <primary_outcome>
    <measure>Evaluating weight</measure>
    <time_frame>At baseline</time_frame>
    <description>Weight (kilograms) data were obtained from the patient's files.</description>
  </primary_outcome>
  <primary_outcome>
    <measure>Evaluating body mass index</measure>
    <time_frame>At baseline</time_frame>
    <description>Body mass index (BMI) (Kg/m2) data were obtained from the patient's files.</description>
  </primary_outcome>
  <primary_outcome>
    <measure>Evaluating medication</measure>
    <time_frame>At baseline</time_frame>
    <description>Medication data were obtained from the patient's files.</description>
  </primary_outcome>
  <primary_outcome>
    <measure>Evaluating adequacy of the dialysis treatment</measure>
    <time_frame>At baseline</time_frame>
    <description>Adequacy of the dialysis treatment was captured by extracting the routine Kt/V from the medical record.</description>
  </primary_outcome>
  <number_of_arms>1</number_of_arms>
  <enrollment type="Actual">6</enrollment>
  <condition>Oxidative Stress</condition>
  <arm_group>
    <arm_group_label>Apple Juice</arm_group_label>
    <arm_group_type>Experimental</arm_group_type>
  </arm_group>
  <intervention>
    <intervention_type>Other</intervention_type>
    <intervention_name>Fuji apple juice</intervention_name>
    <description>On two different days, each volunteer consumed 300 mL Fuji apple juice (AJ), immediately after a dialysis section. After a washout period of 3 weeks, the volunteers drank 150 mL Fuji AJ in a similar manner as described above. Before and after 30 and 60 min of AJ consumption, blood samples were withdrawn for biochemical analysis.</description>
    <arm_group_label>Apple Juice</arm_group_label>
  </intervention>
  <eligibility>
    <criteria>
      <textblock>
        Inclusion Criteria:&#xD;
&#xD;
          -  hemodialysis treatment ≥ 3 months, age ≥ 20 years, and body mass index ≥ 23 kg/m2&#xD;
&#xD;
        Exclusion Criteria:&#xD;
&#xD;
          -  allergy to apple, presence of cancer or acquired immunodeficiency syndrome, kidney&#xD;
             transplant less than 6 months before enrolling in the study, taking antioxidant or&#xD;
             nutritional supplements during the 30 days before enrollment, having been hospitalized&#xD;
             within 6 weeks before the beginning of the study, or suffering from an acute illness&#xD;
      </textblock>
    </criteria>
    <gender>All</gender>
    <minimum_age>20 Years</minimum_age>
    <maximum_age>N/A</maximum_age>
    <healthy_volunteers>No</healthy_volunteers>
  </eligibility>
  <reference>
    <citation>Ruskovska T, Jansen EH, Antarorov R. Evaluation of assays for measurement of serum (anti)oxidants in hemodialysis patients. Biomed Res Int. 2014;2014:843157. doi: 10.1155/2014/843157. Epub 2014 May 19.</citation>
    <PMID>24982909</PMID>
  </reference>
  <reference>
    <citation>Halliwell, B, Gutteridge, JMC. Cellular responses to oxidative stress: adaptation, damage, repair, senescence and death. In: Halliwell, B, Gutteridge, JMC. Free Radical in Biology and Medicine. 4th ed. Oxford, UK: Oxford University Press; 2007, p. 87-267.</citation>
  </reference>
  <reference>
    <citation>McDonald CI, Fraser JF, Coombes JS, Fung YL. Oxidative stress during extracorporeal circulation. Eur J Cardiothorac Surg. 2014 Dec;46(6):937-43. doi: 10.1093/ejcts/ezt637. Epub 2014 Jan 30.</citation>
    <PMID>24482384</PMID>
  </reference>
  <reference>
    <citation>Melidou M, Riganakos K, Galaris D. Protection against nuclear DNA damage offered by flavonoids in cells exposed to hydrogen peroxide: the role of iron chelation. Free Radic Biol Med. 2005 Dec 15;39(12):1591-600. doi: 10.1016/j.freeradbiomed.2005.08.009. Epub 2005 Aug 29.</citation>
    <PMID>16298684</PMID>
  </reference>
  <reference>
    <citation>Spormann TM, Albert FW, Rath T, Dietrich H, Will F, Stockis JP, Eisenbrand G, Janzowski C. Anthocyanin/polyphenolic-rich fruit juice reduces oxidative cell damage in an intervention study with patients on hemodialysis. Cancer Epidemiol Biomarkers Prev. 2008 Dec;17(12):3372-80. doi: 10.1158/1055-9965.EPI-08-0364.</citation>
    <PMID>19064553</PMID>
  </reference>
  <reference>
    <citation>Castilla P, Echarri R, Davalos A, Cerrato F, Ortega H, Teruel JL, Lucas MF, Gomez-Coronado D, Ortuno J, Lasuncion MA. Concentrated red grape juice exerts antioxidant, hypolipidemic, and antiinflammatory effects in both hemodialysis patients and healthy subjects. Am J Clin Nutr. 2006 Jul;84(1):252-62. doi: 10.1093/ajcn/84.1.252.</citation>
    <PMID>16825703</PMID>
  </reference>
  <reference>
    <citation>Shema-Didi L, Sela S, Ore L, Shapiro G, Geron R, Moshe G, Kristal B. One year of pomegranate juice intake decreases oxidative stress, inflammation, and incidence of infections in hemodialysis patients: a randomized placebo-controlled trial. Free Radic Biol Med. 2012 Jul 15;53(2):297-304. doi: 10.1016/j.freeradbiomed.2012.05.013. Epub 2012 May 17.</citation>
    <PMID>22609423</PMID>
  </reference>
  <reference>
    <citation>Fouque D, Kalantar-Zadeh K, Kopple J, Cano N, Chauveau P, Cuppari L, Franch H, Guarnieri G, Ikizler TA, Kaysen G, Lindholm B, Massy Z, Mitch W, Pineda E, Stenvinkel P, Trevino-Becerra A, Wanner C. A proposed nomenclature and diagnostic criteria for protein-energy wasting in acute and chronic kidney disease. Kidney Int. 2008 Feb;73(4):391-8. doi: 10.1038/sj.ki.5002585. Epub 2007 Dec 19. Erratum In: Kidney Int. 2008 Aug;74(3):393. Trevinho-Becerra, A [corrected to Trevino-Becerra, A].</citation>
    <PMID>18094682</PMID>
  </reference>
  <reference>
    <citation>Golding JB, McGlasson WB, Wyllie SG, Leach DN. Fate of apple peel phenolics during cool storage. J Agric Food Chem. 2001 May;49(5):2283-9. doi: 10.1021/jf0015266.</citation>
    <PMID>11368590</PMID>
  </reference>
  <reference>
    <citation>Dai X, Luo H, Jiang L, Ling L, Xue Y, Yu Z. Efficacy of different sanitizing agents and their combination on microbe population and quality of fresh-cut Chinese chives. J Food Sci. 2012 Jul;77(7):M348-53. doi: 10.1111/j.1750-3841.2012.02770.x. Epub 2012 Jun 18.</citation>
    <PMID>22708729</PMID>
  </reference>
  <reference>
    <citation>Association of Official Analytical Chemists (AOAC). Official Methods of Analysis. 18th ed. Washington, DC: Association of Official Analytical Chemists; 2005.</citation>
  </reference>
  <reference>
    <citation>Singleton, VL, Rossi, JA. Colorimetry of total phenolics with phosphomolybdic phosphotungstic acid reagents. Am J Enol Vitic 1965;16(3):144-58.</citation>
  </reference>
  <reference>
    <citation>Arnous, A, Markis, D, Kefalas, P. Correlation of pigment and flavonol content with antioxidant properties in selected aged regional wines from Greece. J Food Composit Ana 2002;15(6):655-65.</citation>
  </reference>
  <reference>
    <citation>Brand-Williams, W, Cuvelier, ME, Berset, C. Use of free radical method to evaluate antioxidant activity. LWT Food Sci Technol 1995;28(1):25-30.</citation>
  </reference>
  <reference>
    <citation>Giusti, MM, Wrolstad, RE. Anthocyanins: characterization and measurement with UV-visible spectroscopy. In: Wrolstald, RE, ed. Current Protocols in Food Analytical Chemistry. New York, NY: John Wiley and Sons; 2001, p. 1-13.</citation>
  </reference>
  <reference>
    <citation>Erel O. A novel automated direct measurement method for total antioxidant capacity using a new generation, more stable ABTS radical cation. Clin Biochem. 2004 Apr;37(4):277-85. doi: 10.1016/j.clinbiochem.2003.11.015.</citation>
    <PMID>15003729</PMID>
  </reference>
  <reference>
    <citation>Erel O. A new automated colorimetric method for measuring total oxidant status. Clin Biochem. 2005 Dec;38(12):1103-11. doi: 10.1016/j.clinbiochem.2005.08.008. Epub 2005 Oct 7.</citation>
    <PMID>16214125</PMID>
  </reference>
  <reference>
    <citation>Chung WY, Chung JK, Szeto YT, Tomlinson B, Benzie IF. Plasma ascorbic acid: measurement, stability and clinical utility revisited. Clin Biochem. 2001 Nov;34(8):623-7. doi: 10.1016/s0009-9120(01)00270-3.</citation>
    <PMID>11849621</PMID>
  </reference>
  <reference>
    <citation>Giustarini D, Dalle-Donne I, Milzani A, Fanti P, Rossi R. Analysis of GSH and GSSG after derivatization with N-ethylmaleimide. Nat Protoc. 2013 Sep;8(9):1660-9. doi: 10.1038/nprot.2013.095. Epub 2013 Aug 1.</citation>
    <PMID>23928499</PMID>
  </reference>
  <reference>
    <citation>Johansson LH, Borg LA. A spectrophotometric method for determination of catalase activity in small tissue samples. Anal Biochem. 1988 Oct;174(1):331-6. doi: 10.1016/0003-2697(88)90554-4.</citation>
    <PMID>3064653</PMID>
  </reference>
  <reference>
    <citation>Wendel A. Glutathione peroxidase. Methods Enzymol. 1981;77:325-33. doi: 10.1016/s0076-6879(81)77046-0. No abstract available.</citation>
    <PMID>7329310</PMID>
  </reference>
  <reference>
    <citation>National Kidney Foundation. K/DOQI clinical practice guidelines for bone metabolism and disease in chronic kidney disease. Am J Kidney Dis. 2003 Oct;42(4 Suppl 3):S1-201. No abstract available.</citation>
    <PMID>14520607</PMID>
  </reference>
  <reference>
    <citation>International Diabetes Federation. Guideline for management of postmeal glucose in diabetes, http://www.idf.org/sites/default/files/postmeal%20glucose%20guidelines.pdf; 2011 [accessed 03.10.16].</citation>
  </reference>
  <verification_date>November 2016</verification_date>
  <study_first_submitted>November 10, 2016</study_first_submitted>
  <study_first_submitted_qc>November 22, 2016</study_first_submitted_qc>
  <study_first_posted type="Estimate">November 28, 2016</study_first_posted>
  <last_update_submitted>November 22, 2016</last_update_submitted>
  <last_update_submitted_qc>November 22, 2016</last_update_submitted_qc>
  <last_update_posted type="Estimate">November 28, 2016</last_update_posted>
  <responsible_party>
    <responsible_party_type>Principal Investigator</responsible_party_type>
    <investigator_affiliation>Universidade Federal de Santa Catarina</investigator_affiliation>
    <investigator_full_name>Andréia G. Giaretta</investigator_full_name>
    <investigator_title>principal investigator</investigator_title>
  </responsible_party>
  <keyword>hemodialysis</keyword>
  <keyword>diabetes mellitus</keyword>
  <keyword>apple juice</keyword>
  <keyword>potassium</keyword>
  <!-- Results have not yet been posted for this study                                          -->
</clinical_study>


download date: ClinicalTrials.gov processed this data on September 20, 2023
link text: Link to the current ClinicalTrials.gov record.
url: https://clinicaltrials.gov/ct2/show/NCT02974491
org study id: 37090614.6.0000.0118
nct id: NCT02974491
lead sponsor: 
has dmc: Yes
The investigators hypothesized that acute consumption of Fuji apple juice (AJ) could increase
      the antioxidant status and/or decrease the oxidative stress (OS) biomarkers, without
      increasing serum biochemical parameters in patients undergoing maintenance hemodialysis
      (MHD). In this pre-post pilot feasibility study, patients served as their own controls,
      received 300 and 150 mL AJ immediately after a dialysis section, on different days, with a 3
      week-washout period. Blood was collected at the baseline period, after 30 and 60 min of AJ
      consumption. OS biomarkers (total antioxidant status (TAS), total oxidant status (TOS),
      ascorbic acid, catalase (CAT), glutathione peroxidase (Gpx), superoxide dismutase (SOD) and
      reduced glutathione (GSH) and potassium, phosphorus, uric acid, and glucose concentrations
      were analyzed.
Subjects and study design

      In this pre-post pilot feasibility study, subjects served as their own controls. Participants
      were recruited from a hemodialysis clinic, located in Florianópolis' metropolitan area, Santa
      Catarina state (Southern Brazil), from June to August 2015 with the following inclusion
      criteria: hemodialysis treatment ≥ 3 months, age ≥ 20 years, and body mass index (BMI) ≥ 23
      kg/m2. Exclusion criteria were allergy to apple, presence of cancer or acquired
      immunodeficiency syndrome, kidney transplant less than 6 months before enrolling in the
      study, taking antioxidant or nutritional supplements during the 30 days before enrollment,
      having been hospitalized within 6 weeks before the beginning of the study, or suffering from
      an acute illness.

      Subjects were instructed to keep their usual dietary habits for the duration of the study,
      except for avoiding intake of polyphenol-rich foods (i.e., vegetables, fruits and
      fruit-containing products, chocolate, tea, coffee, honey, and any alcoholic beverage) within
      2 days before and during intervention. On two different days, each volunteer consumed 300 mL
      Fuji AJ, immediately after a dialysis section. After a washout period of 3 weeks, the
      volunteers drank 150 mL Fuji AJ in a similar manner as described above. Before and after 30
      and 60 min of AJ consumption, blood samples were withdrawn for biochemical analysis. This
      protocol was chosen taking into account the previous favorable effects of AJ consumption on
      serum OS biomarkers described for healthy volunteers.

      Baseline clinical and biochemical data were obtained from medical records. This study was
      approved by the ethics committee of human research of the Federal University of Santa
      Catarina (UFSC) (protocol number 37090614.6.0000.0118) and all participants gave written and
      informed consent.

      Apple juice and analysis

      Fuji apples were obtained from the Experimental Seasonal Fields at the Empresa de Pesquisa
      Agropecuária e Extensão Rural de Santa Catarina' Station in the city of São Joaquin, Santa
      Catarina, Brazil (latitude 28º 17' 39", longitude 49º 55' 56" and altitude 1.415 m),
      harvested in their commercial maturation stage during 2015's harvest. After harvest, apples
      were stored at 4 ± 1°C. To allow for a quick and easy apple intake in large amounts, 300 mL
      of AJ, equivalent to 3.5 apples, and 150 mL AJ, equivalent to 1.5 apples were used. Before
      preparing the juice, the fruits were sanitized in sodium hypochlorite. Apples with peel and
      no seeds were blended in a centrifugal juice extractor without water addition. Each dose of
      AJ was prepared and consumed immediately.

      The contents of dry matter, total soluble solids, potential of hydrogen (pH) and titratable
      acidity of AJ were measured following official methods. The total phenolic content of the AJ
      extracts was measured using the Folin-Ciocalteu method colorimetric. The total flavanol
      content was estimated using p-dimethylaminocinnamaldehyde (DMACA) method. The total
      antioxidant capacity was determined using the DPPH (2,2-diphenyl-1-picryl-hydrazyl-hydrate)
      method. For total monomeric anthocyanin, a spectrophotometric pH differential method was
      used. Analyses were conducted with each AJ used in the two intervention days, in triplicate.

      Blood sampling and laboratory methods

      In each experiment day, blood samples from participants were collected in three different
      moments. Venous blood samples were collected using a vacuum system (Vacutainer Becton
      Dickinson BD®, São Paulo, Brazil) into heparin, Ethylenediaminetetraacetic acid
      (EDTA)-containing tubes or additive-free tubes. Plasma and serum were immediately obtained by
      centrifugation (3,000 revolutions per minute (RPM), 10 min, room temperature) for the
      measurement of uric acid, phosphorus, glucose, potassium, Total Antioxidant Status (TAS) and
      Total Oxidant Status( TOS). In order to quantify the endogenous antioxidant enzymes, an
      aliquot of whole blood was lysed. In order to measure reduced glutathione (GSH), an aliquot
      of whole blood was preserved in N-ethylmaleimide (NEM). All samples were immediately stored
      at -80°C for later analysis in duplicate.

      Serum uric acid, phosphorus and glucose concentrations were determined using commercially
      available kits (Labtest Diagnóstica SA, Lagoa Santa, Minas Gerais, Brazil) in an automated
      multi-biochemical analyzer (Cobas Miras Plus, Roche®, Germany). Serum potassium levels were
      measured in an automated Dimension Max System (Siemens Healthcare Diagnostics Products®,
      Germany), according to the manufacturer's instructions.

      TAS and TOS assays were measured spectrophotometrically, according to Erel's methods.
      Ascorbic acid and GSH were determined using high-performance liquid chromatography (HPLC), as
      previously described. The superoxide dismutase (SOD) activity was evaluated using SOD Assay
      Kit (Sigma Aldrich®, St. Louis, Missouri, USA) according to the manufacturer's instructions.
      Catalase (CAT) activity was determined spectrophotometrically according to the previously
      described method. The glutathione peroxidase (GPx) activity was determined using the NADPH
      oxidation rate method.
allocation: N/A
intervention model: Single Group Assignment
primary purpose: Treatment
masking: None (Open Label)
measure: Measuring oxidative stress biomarkers
time frame: Change from baseline at 30 and 60 minutes
description: Total antioxidant status (TAS) (mmol/L) and total oxidant status (TOS) (mmol/L) was measured at the baseline period, after 30 and 60 min of acute consumption of 300 and 150 mL of Fuji apple (Malus domestica Borkh) juice. Patients were not fasted for the withdrawal of blood. This measure was taken to prevent discomfort among volunteers. Venous blood samples were collected through aseptic venipuncture on the opposite arm of the arteriovenous fistula by a qualified professional using a vacuum system into heparin or EDTA-containing tubes or tubes without additives according to established standards of clinical and biological safety.
measure: Measuring endogenous antioxidant enzymes
time frame: Change from baseline at 30 and 60 minutes
description: Catalase (U mg/Hb), glutathione peroxidase (U mg/Hb) and superoxide dismutase (U mg/Hb) was measured at the baseline period, after 30 and 60 min of acute consumption of 300 and 150 mL of Fuji apple (Malus domestica Borkh) juice. Patients were not fasted for the withdrawal of blood. This measure was taken to prevent discomfort among volunteers. Venous blood samples were collected through aseptic venipuncture on the opposite arm of the arteriovenous fistula by a qualified professional using a vacuum system into heparin or EDTA-containing tubes or tubes without additives according to established standards of clinical and biological safety.
measure: Measuring oxidative stress biomarkers
time frame: Change from baseline at 30 and 60 minutes
description: Ascorbic acid (micromole/L) was measured at the baseline period, after 30 and 60 min of acute consumption of 300 and 150 mL of Fuji apple (Malus domestica Borkh) juice. Patients were not fasted for the withdrawal of blood. This measure was taken to prevent discomfort among volunteers. Venous blood samples were collected through aseptic venipuncture on the opposite arm of the arteriovenous fistula by a qualified professional using a vacuum system into heparin or EDTA-containing tubes or tubes without additives according to established standards of clinical and biological safety.
measure: Measuring oxidative stress biomarkers
time frame: Change from baseline at 30 and 60 minutes
description: Reduced glutathione (micromole mg/Hb) was measured at the baseline period, after 30 and 60 min of acute consumption of 300 and 150 mL of Fuji apple (Malus domestica Borkh) juice. Patients were not fasted for the withdrawal of blood. This measure was taken to prevent discomfort among volunteers. Venous blood samples were collected through aseptic venipuncture on the opposite arm of the arteriovenous fistula by a qualified professional using a vacuum system into heparin or EDTA-containing tubes or tubes without additives according to established standards of clinical and biological safety.
measure: Measuring serum biochemical parameters
time frame: Change from baseline at 30 and 60 minutes
description: Potassium (mmol/L), phosphorus (mmol/L) and glucose (mmol/L) was measured at the baseline period, after 30 and 60 min of acute consumption of 300 and 150 mL of Fuji apple (Malus domestica Borkh) juice. Patients were not fasted for the withdrawal of blood. This measure was taken to prevent discomfort among volunteers. Venous blood samples were collected through aseptic venipuncture on the opposite arm of the arteriovenous fistula by a qualified professional using a vacuum system into heparin or EDTA-containing tubes or tubes without additives according to established standards of clinical and biological safety.
measure: Measuring serum biochemical parameters
time frame: Change from baseline at 30 and 60 minutes
description: Uric acid (micromole/L) was measured at the baseline period, after 30 and 60 min of acute consumption of 300 and 150 mL of Fuji apple (Malus domestica Borkh) juice. Patients were not fasted for the withdrawal of blood. This measure was taken to prevent discomfort among volunteers. Venous blood samples were collected through aseptic venipuncture on the opposite arm of the arteriovenous fistula by a qualified professional using a vacuum system into heparin or EDTA-containing tubes or tubes without additives according to established standards of clinical and biological safety.
measure: Evaluating heigth
time frame: at baseline
description: Height (meters) data were obtained from the patient's files.
measure: Evaluating weight
time frame: At baseline
description: Weight (kilograms) data were obtained from the patient's files.
measure: Evaluating body mass index
time frame: At baseline
description: Body mass index (BMI) (Kg/m2) data were obtained from the patient's files.
measure: Evaluating medication
time frame: At baseline
description: Medication data were obtained from the patient's files.
measure: Evaluating adequacy of the dialysis treatment
time frame: At baseline
description: Adequacy of the dialysis treatment was captured by extracting the routine Kt/V from the medical record.
arm group label: Apple Juice
arm group type: Experimental
intervention type: Other
intervention name: Fuji apple juice
description: On two different days, each volunteer consumed 300 mL Fuji apple juice (AJ), immediately after a dialysis section. After a washout period of 3 weeks, the volunteers drank 150 mL Fuji AJ in a similar manner as described above. Before and after 30 and 60 min of AJ consumption, blood samples were withdrawn for biochemical analysis.
arm group label: Apple Juice
criteria: 
gender: All
minimum age: 20 Years
maximum age: N/A
healthy volunteers: No
citation: Ruskovska T, Jansen EH, Antarorov R. Evaluation of assays for measurement of serum (anti)oxidants in hemodialysis patients. Biomed Res Int. 2014;2014:843157. doi: 10.1155/2014/843157. Epub 2014 May 19.
PMID: 24982909
citation: Halliwell, B, Gutteridge, JMC. Cellular responses to oxidative stress: adaptation, damage, repair, senescence and death. In: Halliwell, B, Gutteridge, JMC. Free Radical in Biology and Medicine. 4th ed. Oxford, UK: Oxford University Press; 2007, p. 87-267.
citation: McDonald CI, Fraser JF, Coombes JS, Fung YL. Oxidative stress during extracorporeal circulation. Eur J Cardiothorac Surg. 2014 Dec;46(6):937-43. doi: 10.1093/ejcts/ezt637. Epub 2014 Jan 30.
PMID: 24482384
citation: Melidou M, Riganakos K, Galaris D. Protection against nuclear DNA damage offered by flavonoids in cells exposed to hydrogen peroxide: the role of iron chelation. Free Radic Biol Med. 2005 Dec 15;39(12):1591-600. doi: 10.1016/j.freeradbiomed.2005.08.009. Epub 2005 Aug 29.
PMID: 16298684
citation: Spormann TM, Albert FW, Rath T, Dietrich H, Will F, Stockis JP, Eisenbrand G, Janzowski C. Anthocyanin/polyphenolic-rich fruit juice reduces oxidative cell damage in an intervention study with patients on hemodialysis. Cancer Epidemiol Biomarkers Prev. 2008 Dec;17(12):3372-80. doi: 10.1158/1055-9965.EPI-08-0364.
PMID: 19064553
citation: Castilla P, Echarri R, Davalos A, Cerrato F, Ortega H, Teruel JL, Lucas MF, Gomez-Coronado D, Ortuno J, Lasuncion MA. Concentrated red grape juice exerts antioxidant, hypolipidemic, and antiinflammatory effects in both hemodialysis patients and healthy subjects. Am J Clin Nutr. 2006 Jul;84(1):252-62. doi: 10.1093/ajcn/84.1.252.
PMID: 16825703
citation: Shema-Didi L, Sela S, Ore L, Shapiro G, Geron R, Moshe G, Kristal B. One year of pomegranate juice intake decreases oxidative stress, inflammation, and incidence of infections in hemodialysis patients: a randomized placebo-controlled trial. Free Radic Biol Med. 2012 Jul 15;53(2):297-304. doi: 10.1016/j.freeradbiomed.2012.05.013. Epub 2012 May 17.
PMID: 22609423
citation: Fouque D, Kalantar-Zadeh K, Kopple J, Cano N, Chauveau P, Cuppari L, Franch H, Guarnieri G, Ikizler TA, Kaysen G, Lindholm B, Massy Z, Mitch W, Pineda E, Stenvinkel P, Trevino-Becerra A, Wanner C. A proposed nomenclature and diagnostic criteria for protein-energy wasting in acute and chronic kidney disease. Kidney Int. 2008 Feb;73(4):391-8. doi: 10.1038/sj.ki.5002585. Epub 2007 Dec 19. Erratum In: Kidney Int. 2008 Aug;74(3):393. Trevinho-Becerra, A [corrected to Trevino-Becerra, A].
PMID: 18094682
citation: Golding JB, McGlasson WB, Wyllie SG, Leach DN. Fate of apple peel phenolics during cool storage. J Agric Food Chem. 2001 May;49(5):2283-9. doi: 10.1021/jf0015266.
PMID: 11368590
citation: Dai X, Luo H, Jiang L, Ling L, Xue Y, Yu Z. Efficacy of different sanitizing agents and their combination on microbe population and quality of fresh-cut Chinese chives. J Food Sci. 2012 Jul;77(7):M348-53. doi: 10.1111/j.1750-3841.2012.02770.x. Epub 2012 Jun 18.
PMID: 22708729
citation: Association of Official Analytical Chemists (AOAC). Official Methods of Analysis. 18th ed. Washington, DC: Association of Official Analytical Chemists; 2005.
citation: Singleton, VL, Rossi, JA. Colorimetry of total phenolics with phosphomolybdic phosphotungstic acid reagents. Am J Enol Vitic 1965;16(3):144-58.
citation: Arnous, A, Markis, D, Kefalas, P. Correlation of pigment and flavonol content with antioxidant properties in selected aged regional wines from Greece. J Food Composit Ana 2002;15(6):655-65.
citation: Brand-Williams, W, Cuvelier, ME, Berset, C. Use of free radical method to evaluate antioxidant activity. LWT Food Sci Technol 1995;28(1):25-30.
citation: Giusti, MM, Wrolstad, RE. Anthocyanins: characterization and measurement with UV-visible spectroscopy. In: Wrolstald, RE, ed. Current Protocols in Food Analytical Chemistry. New York, NY: John Wiley and Sons; 2001, p. 1-13.
citation: Erel O. A novel automated direct measurement method for total antioxidant capacity using a new generation, more stable ABTS radical cation. Clin Biochem. 2004 Apr;37(4):277-85. doi: 10.1016/j.clinbiochem.2003.11.015.
PMID: 15003729
citation: Erel O. A new automated colorimetric method for measuring total oxidant status. Clin Biochem. 2005 Dec;38(12):1103-11. doi: 10.1016/j.clinbiochem.2005.08.008. Epub 2005 Oct 7.
PMID: 16214125
citation: Chung WY, Chung JK, Szeto YT, Tomlinson B, Benzie IF. Plasma ascorbic acid: measurement, stability and clinical utility revisited. Clin Biochem. 2001 Nov;34(8):623-7. doi: 10.1016/s0009-9120(01)00270-3.
PMID: 11849621
citation: Giustarini D, Dalle-Donne I, Milzani A, Fanti P, Rossi R. Analysis of GSH and GSSG after derivatization with N-ethylmaleimide. Nat Protoc. 2013 Sep;8(9):1660-9. doi: 10.1038/nprot.2013.095. Epub 2013 Aug 1.
PMID: 23928499
citation: Johansson LH, Borg LA. A spectrophotometric method for determination of catalase activity in small tissue samples. Anal Biochem. 1988 Oct;174(1):331-6. doi: 10.1016/0003-2697(88)90554-4.
PMID: 3064653
citation: Wendel A. Glutathione peroxidase. Methods Enzymol. 1981;77:325-33. doi: 10.1016/s0076-6879(81)77046-0. No abstract available.
PMID: 7329310
citation: National Kidney Foundation. K/DOQI clinical practice guidelines for bone metabolism and disease in chronic kidney disease. Am J Kidney Dis. 2003 Oct;42(4 Suppl 3):S1-201. No abstract available.
PMID: 14520607
citation: International Diabetes Federation. Guideline for management of postmeal glucose in diabetes, http://www.idf.org/sites/default/files/postmeal%20glucose%20guidelines.pdf; 2011 [accessed 03.10.16].
responsible party type: Principal Investigator
investigator affiliation: Universidade Federal de Santa Catarina
investigator full name: Andréia G. Giaretta
investigator title: principal investigator