This study was a double-blinded randomized controlled clinical trial with a parallel design organized in Dr. Cipto Mangunkusumo Hospital, a national referral hospital in Jakarta, Indonesia, from August through December 2020.
Subjects included CKD patients on haemodialysis who met the inclusion criteria and were recruited using a consecutive sampling method.
The inclusion criteria were as follows: 1) patients over 18 years old who underwent standard haemodialysis treatment twice a week for five hours for at least three months and 2) patients with gastrointestinal complaints (i.e., difficulty defecating, faeces with hard consistency, or a bowel movement frequency of fewer than three times a week). Patients with a history of malignancy, chemotherapy or radiotherapy, patients with autoimmune disorders or receiving immunosuppressants, patients who underwent gut resection, patients with Crohn’s disease or ulcerative colitis, patients whose haemodialysis schedule was altered, patients consuming prebiotics/probiotics/synbiotics, and patients suffering from infection or consuming antibiotics were excluded from this study.
All included subjects underwent history taking, physical examination, and laboratory examination (blood) and provided consent to participate in the study. The subjects then underwent history taking for demographic data, comorbidities and medications as well as physical examination for vital signs, body weight, and body height. We tested the blood sample to evaluate haemoglobin, leucocytes, thrombocytes, urea, creatinine, and albumin to determine baseline characteristics.
After randomization, each subject received 60 capsules containing synbiotics (Lactobacillus acidophilus and Bifidobacterium longum 5x109 CFU and 60 mg of fructooligosaccharides (FOS)) or 60 capsules containing placebo (Saccharum lactis). The daily dosage was two capsules per day taken every morning before a meal. All subjects were given a compliance card that had to be completed every day after they took the drug. Medication adherence was assessed every 30 days, and the subjects returned any medicine left and showed the compliance card at the subsequent follow-up.
Food intake was assessed before and after the intervention was administered using 24-hour food recall and was carried out by a nutritionist. To evaluate caloric intake and carbohydrates, protein, fat, and fibres consumed by the patient throughout the study, a programme (nutriSurvey) was used. In addition, all participants were asked to continue their dietary habits and previous lifestyle habits and were prohibited from consuming any motility agents during the study period.
After 30 days, a follow-up was performed to evaluate side effects and compliance with the drug regimen. Each subject was then provided with another 60 synbiotic capsules or 60 placebo capsules based on their previous grouping.
The exclusion criteria included subjects who withdrew from the study, those who missed their dose of synbiotics or placebo for more than three consecutive days, those with infections that required antibiotics, those whose haemodialysis schedule changed from twice a week to three times a week or whose treatment modality changed from dialysis to peritoneal dialysis or to renal transplant and those who experienced gastrointestinal symptoms, such as diarrhoea or profuse vomiting requiring hospital admission. Patient adherence to the medication under investigation was expected to reach over 90%.
The primary outcome of this study was a decrease in IS levels. The examination was conducted twice, before and after the intervention was administered. Blood samples were taken predialysis and after each subject fasted for ±8-10 hours. Approximately 100 μL of each subject's serum was added to 900 L of acetonitrile to precipitate the protein. The supernatant was then added to 500 μL of 5 M NaCl for salting-out-assisted liquid–liquid extraction (SALLE) and centrifuged at 14000 rpm for 10 minutes. As a result, two phases were formed: the organic phase (IS in acetonitrile and internal standard) and the aqueous, NaCl and matrix constituent phases. The organic phase was then separated, and as much as 0.2 L was injected into the HPLC system with a fluorescence detector (Agilent Technologies with MassHunterChemStation Software version B04.03.E). IS was measured quantitatively using seven calibration levels with a calibration range of 0.02-100 mg/L. The concentration of IS is expressed in units of mg/L.
The secondary outcome of this study was improvement in constipation-related symptoms and quality of life. They were assessed using the Indonesian-validated Patient Assessment of Constipation: Symptoms (PAC-SYM) and Patient Assessment of Constipation Quality of Life (PAC-QOL) questionnaires [12, 13]. The examination was conducted twice, at the beginning and the end of the study. The PAC-SYM questionnaire includes a 0-4 scale and consists of three parts: abdominal symptoms (questions 1-4), rectal symptoms (questions 5-7), and stool symptoms (questions 8-12). Symptom improvement was defined as a decrease in the total PAC-SYM score > 1. The PAC-QOL questionnaire includes a 0-4 scale and consists of four parts: physical discomfort (questions 1-4), psychosocial discomfort (questions 5-12), worries/concern (questions 13-23), and satisfaction (question 24-28). Quality of life improvement was defined as a decrease in the total PAC-QOL score > 1.
To estimate the sample size, a formula for comparing two means was used. With a(n) alpha of 0.05, power of 0.80, and dropout of 20%, the minimum sample size for each group was 30.
We only enrolled subjects who gave consent. A third party (pharmacist) randomized the subjects into two study groups using a computer randomizer. Synbiotics and placebo were both packed on an identical clear gastroenteric coated capsule. Each subject was given one plastic pot containing either intervention or placebo drugs; each identical pot contained 60 capsules. A white paper with information on drug use and administration was placed on the cover of the pot. The drugs were distributed by a third party. None of the patients, researchers, or physicians in charge were aware of the treatment groups.
We performed the statistical analysis using SPSS version 20.0. Mean and standard deviation (SD) analyses were performed for numeric data with normal distributions, whereas medians and interquartile ranges (IQRs) were calculated for data with nonnormal distributions. This study utilized intention-to-treat analysis. Bivariate analysis using an independent t test was performed to analyse data with a normal distribution, and the Mann–Whitney U test was performed to analyse data with a nonnormal distribution. A p value of <0.05 was considered statistically significant.