Effect of citrus-based products on urine profile: A systematic review and meta-analysis

Background . Urolithiasis is a disease with high recurrence rate, 30-50% within 5 years. The aim of the present study was to learn the effects of citrus-based products on the urine profile in healthy persons and people with urolithiasis compared to control diet and potassium citrate. Methods. A systematic review was performed, which included interventional, prospective observational and retrospective studies, comparing citrus-based therapy with standard diet therapy, mineral water, or potassium citrate. A literature search was conducted using PUBMED, COCHRANE, and Google Scholar with “citrus or lemonade or orange or grapefruit or lime or juice” and “urolithiasis” as search terms. For statistical analysis, a fixed-effects model was conducted when p > 0.05, and random-effects model was conducted when p < 0.05. Results. In total, 135 citations were found through database searching with 10 studies found to be consistent with our selection criteria. However, only 8 studies were included in quantitative analysis, due to data availability. The present study showed a higher increased in urine pH for citrus-based products (mean difference, 0.16; 95% CI 0.01-0.32) and urinary citrate (mean difference, 124.49; 95% CI 80.24-168.74) compared with a control group. However, no differences were found in urine volume, urinary calcium, urinary oxalate, and urinary uric acid. From subgroup analysis, we found that citrus-based products consistently increased urinary citrate level higher than controls in both healthy and urolithiasis populations. Furthermore, there was lower urinary calcium level among people with urolithiasis. Conclusions. Citrus-based products could increase urinary citrate level significantly higher than control. These results should encourage further research to explore citrus-based products as a urolithiasis treatment.

Humans have suffered urinary tract stones for centuries 1 . The incidence and prevalence of urolithiasis are different between geographic locations, depending on age and sex distribution, stone composition and stone location 2 . Risk of stone development has been shown to be 5-10% with a higher prevalence in men than women 3 . Urolithiasis is a common disease with significant morbidity and cost worldwide [4][5][6] . Based on National Health and Nutrition Examination survey, kidney stones affect 1 in 11 people in the United States, and an epidemiological increase was found in 2012 compared to 1994 7 . Additional data from Dr. Cipto Mangunkusumo National General Hospital, Indonesia's national referral hospital, showed an increase in stone disease prevalence from 182 patients in 1997 to 847 patients in 2002 8 . Moreover, it is further worsen by its high recurrence rate reaching 30-50% within 5 years 7 .
Calcium-based urinary tract stone is the most common stone composition found in urolithiasis 9,10 . Supersaturation is believed to be the mechanism behind calcium stone formation 11 . One factor in determining urine stone formation or stone recurrence is urine profile, which is defined as urine volume and its composition. Hypercalciuria and hypocitraturia are the most common urine abnormalities found among calcium stone-formers 12 . A high fluid intake could prevent stone formation by lowering supersaturation, whereas citrate could prevent stone formation by ionizing urinary calcium 13,14 . Food that is rich of citrate is citrus. There are wide variety of citrus fruits and derivate products that can be easily obtained, such as lemonade, grapefruit, orange, lime, and citrus-based juice. Several studies had already been conducted to learn the effect of citrus-based products on urine profile. However, the results between those studies were contradictive. Therefore, our study aimed to systematically review and quantify the available studies regarding the effects of citrus-based products on urine profile and its comparison to a control diet and potassium citrate.

Eligibility criteria
We included both healthy people and patients with urolithiasis history in our selection criteria. Study subjects must have consumed citrus fruits, such as orange, lime, grapefruit, or juices made from the fruits. Study designs could be interventional, prospective observational, or retrospective with standard diet therapy (any kind of mineral water), or potassium citrate, as a control group therapy. We included studies with urine profile as the outcome. We only included articles written in English or Indonesian, and those with full text article available. We excluded non-systematic review articles. We did not limit studies based on their year conducted.

Search strategy
A literature search was conducted using PUBMED, COCHRANE, and Google Scholar as search engines on August 2016. The terms "citrus OR lemonade OR orange OR grapefruit OR lime OR juice" AND "urolithiasis" were used as search terms. We also searched the list of references in included studies. We did not use any limitation in study searching.

Study selection and data extraction
All studies were screened for duplication using EndNote X6 software. Duplication-free articles underwent title and abstract examination using predetermined inclusion and exclusion criteria mentioned above. Selection of studies was selected by two authors independently. Discrepancies of opinion were resolved by discussion. All studies, which fulfilled the inclusion and exclusion criteria, underwent full text review. For every eligible full text, we extracted the following data, if available: subjects specific condition, citrusbased product used in the study, number of patients consuming citrus-based product, citrate content or its concentration, control intervention, number of individuals under control intervention. For the outcomes, we extracted urine profile data as follows: volume, pH, calcium level, citrate level, oxalate level, and uric acid level. Measurement units used in this study are L/day for urine volume and mg/day for urinary calcium level, urinary citrate level, urinary oxalate level, and urinary uric acid level. All data in the form of numbers were extracted manually as mean and standard deviation for variable measurement.

Assessment of bias and statistical methods
This study used Cochrane Risk of Bias assessment tools 15 and Newcastle-Ottawa scale 16 to assess interventional and retrospective study's quality, respectively. These assessments of study quality were done by two authors independently. Quantitative synthesis of included studies was analyzed using Review Manager (RevMan) 5.0 software and mean difference was used as its effects size measurement. Heterogeneity of studies was assessed using chisquare. A fixed-effects model was conducted when p > 0.05, whereas a random-effect model is conducted when p < 0.05. We also conducted subgroup analysis to differentiate between healthy and urolithiasis populations.
Studies which could not be included in quantitative analysis were described qualitatively.

Results
We found 135 citations through database searching. Literature searching from the list of references found similar studies that were all already included in this study. Ten studies were found to be consistent with our selection criteria ( Figure 1).
Two of ten studies had to be excluded from quantitative analysis because of the following reasons: (1) Penniston et al. 17 only published baseline data and its maximal change following intervention; and (2) Tosukhowong et al. 19 used medians as their outcome measurement, and due to its non-uniform distribution, we were unable to convert these to means. Therefore, eight studies were analyzed to find the effect of citrus-based products on urine profile compared to controls. However, not all of the eight studies were included in urine profile outcome measurement, due to data availability. Characteristics of the included studies and their risk of bias assessment can be seen in  Figure 4).
Subgroup analysis showed a significantly higher urinary citrate level in both the healthy population and the population with history of urolithiasis who received citrus-based therapy compared to control. However, urine pH, which showed a statistically significant increase in urine pH compared to controls, did not demonstrate any differences in a subgroup analysis. On the other hand, urinary calcium was lower after consumption of citrusbased products compared to controls in the urolithiasis population. Furthermore, this study demonstrated that there was a lower urine volume in the healthy population after drinking citrus-based products compared to controls ( Figure 5). We did not find any differences in other urine profile variables, either in the healthy population or the population with history of urolithiasis (Supplementary Figure 1 and Supplementary Figure 2  RS -retrospective study; RCT -randomized controlled trial; CBAS -controlled before-after study; CS -crossover study. *Also included in qualitative synthesis for comparison between citrus-based product and potassium citrate.

Comparison between citrus-based products and potassium citrate in urine profile
Due to the reasons stated above, we decided to discuss the comparisons between citrus-based product and potassium citrate in a qualitative manner.
Three studies showed both citrus-based products (lemon juice and lime powder) and potassium citrate increased the level of urinary citrate significantly 17,19,20 . Even though no significant difference in post treatment urine profile was found between citrus-based products and potassium citrate, post-treatment citrate level in the potassium citrate group showed a 3.5 times increase from pre-treatment level, while it was only 2.5 times in the lemon juice group 20 .

Discussion
This study showed that citrus-based products, such as lemonade, orange juice and grapefruit juice, could increase urinary citrate levels and urine pH. Low citrate excretion, as in type I tubular acidosis, shows an increase in nephrolithiasis incidence 27 . Therefore, existence of citrate in urine is important since it is a well-known preventive factor in calcium stone formation, with an increase in   calcium salt solubility and calcium oxalate crystal growth inhibition as its primary mechanism. It also can reduce bone resorption and increase calcium reabsorption in kidneys. Furthermore, citrate fixes the inhibitory properties of Tamm-Horsfall protein 28 . Citrate and Tamm-Horsfall protein are related to inhibition of calcium oxalate agglomeration 29 . An increase in urine pH is due to metabolism of citrate into bicarbonate 13 . Moreover, an increase in urine pH could reduce reabsorption of citrate 30 . Thus, it could induce more citrate excretion. A study conducted by Curhan et al. 31 found an increased risk of stone formation associated with grapefruit juice consumption; although, the exact mechanism is still unclear. One theory suggests that grapefruit juice contains sugar, which can increase calcium excretion 31 . However, this study proved that citrus-based products could increase urinary citrate level, which could be a protective factor for urinary tract stone formation.
Potassium citrate has been used as urolithiasis stone treatment for more than two decades. Its effectiveness in urolithiasis treatment has been established from several studies 32,33 . From one metaanalysis conducted by Phillip et al., potassium citrate significantly reduced stone size, reduced new stone formation, and increased citrate levels 34 . The stone prevention mechanism of potassium citrate is thought to be due to alkali loading and its citrate-uric effect 35 . In this study, potassium citrate showed a significant increase in urinary citrate and is superior to citrus-based products in elevating urinary citrate. However, the use of potassium citrate has a limitation due to its side effect if used for a long term period, such as epigastric discomfort and frequent large bowel movement, and it requires the consumption of many tablets daily to reach sufficient therapeutic doses, which could dramatically decrease patient compliance 36 . Therefore, citrus-based products could be an alternative therapy with lower cost and better urinary citrate level than control therapy. This is the first systematic review and meta-analysis that focuses on citrus-based product and its effect towards urine profile compared to standard therapy. However, this study only searched for published article which could lead into publication bias. Moreover, most of the included studies were not conducted using the best method for interventional studies, which is randomized controlled trials. Therefore, from the positive results this study has shown, we encourage other researchers to conduct randomized controlled trials to provide stronger evidence the beneficial effects of citrus-based products on urinary stone disease.

Conclusions
Citrus-based products increase urinary citrate and urine pH significantly compared to control treatments. Compared to standard potassium citrate therapy, there was a smaller increase in urine pH and urine citrate using citrus-based products. However, due to potassium citrate side effects and patient's poor compliance, citrus-based products could be alternative therapy in preventing stone formation. These study's results should encourage further research to explore citrus-based product as a urolithiasis treatment. Author contributions PB and NR developed the concept of this study. IW designed the research methodology. FR did the literature searching. FR and PB did the selection of studies. FR prepared the draft of manuscript. All author contributed in the revision of the draft manuscript and have agreed to the final content.

Competing interests
No competing interests were disclosed. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Supplementary material
Supplementary Table 1. Newcastle-Ottawa scale for retrospective study's risk of bias assessment. Click here to access the data.
Supplementary Table 2: PRISMA checklist. Click here to access the data.
Supplementary Figure 1. Urine pH, urinary calcium, urinary oxalate, and urinary uric acid in healthy subject population. Click here to access the data.
Supplementary Figure 2. Urine volume, urine pH, urinary oxalate, and urinary uric acid in population with a history of urolithiasis. Click here to access the data.