Diagnostic utility of protein to creatinine ratio (P/C ratio) in spot urine sample within routine clinical practice

Abstract

The spot (random) urine protein to creatinine ratio (P/C ratio) is an alternative, fast and simple method of detecting and estimating the quantitative assessment of proteinuria. The aim of the work was to review the literature concerning the usefulness of spot urine P/C ratio evaluation in the diagnosis of proteinuria in the course of kidney disease, hypertension, gestational hypertension, preeclampsia, immunological diseases, diabetes mellitus, and multiple myeloma, and in the diagnosis of proteinuria in children. We searched the PubMed and Google Scholar databases using the following keywords: proteinuria, spot urine protein to creatinine ratio, spot urine P/C ratio, protein creatinine index, PCR (protein to creatinine ratio), P/C ratio and methods, Jaffe versus enzymatic creatinine methods, urine protein methods, spot urine protein to creatinine ratio versus ACR (albumin to creatinine ratio), proteinuria versus albuminuria, limitations of the P/C ratio. More weight was given to the articles published in the last 10–20 years. A spot urine P/C ratio >20 mg/mmol (0.2 mg/mg) is the most commonly reported cutoff value for detecting proteinuria, while a P/C ratio value >350 mg/mmol (3.5 mg/mg) confirms nephrotic proteinuria. The International Society for the Study of Hypertension in Pregnancy recommends a P/C ratio of 30 mg/mmol (0.3 mg/mg) for the classification of proteinuria in pregnant women at risk of preeclampsia. A high degree of correlation was observed between P/C ratio values and the protein concentration in 24-h urine collections. The spot urine P/C ratio is a quick and reliable test that can eliminate the need for a daily 24-h urine collection. However, in doubtful situations, it is still recommended to assess proteinuria in a 24-h urine collection. The literature review indicates the usefulness of the spot P/C ratio in various disease states; therefore, this test should be available in every laboratory. However, the challenge for the primary care physician is to know the limitations of the methods used to determine the protein and creatinine concentrations that are used to calculate the P/C ratio. Moreover, the P/C ratio cutoff used should be determined in individual laboratories because it depends on the patient population and the laboratory methodologies.

Keywords:

• P/C ratio
• proteinuria
• ACR - albumin to creatinine ratio
• spot urine sample
• 24-h urine collection

Background

Proteinuria is one of the main symptoms of kidney disease. Assessment of proteinuria is helpful in establishing a diagnosis, monitoring the course of the disease and assessing the effectiveness of applied treatment [1]. In states such as nephrotic syndrome, the amount of urinary protein excreted directly reflects disease activity [2]. In nephrotic syndrome, severe proteinuria ≥3.5 g/d (so-called nephrotic proteinuria) is associated with the presence of hypoalbuminemia, edema, sodium retention, and hyperlipidemia, as well as thromboembolic and infection complications [3].

The aim of the work was to review the literature on the suitability of the protein to creatinine ratio (P/C ratio) in a single urine sample in the diagnosis of proteinuria in the course of kidney diseases, hypertension, gestational hypertension, preeclampsia, immunological diseases, diabetes mellitus, MM, and in the diagnosis of proteinuria in children. Alternative names for the spot (random) urine P/C ratio are protein to creatinine ratio (PCR) and protein creatinine index (PCI). In addition, the review was aimed at answering the following questions:

• Are the reference values for the spot urine P/C ratio established?

• Can the spot urine P/C ratio be a fast and reliable test to confirm/exclude proteinuria?

• Can the spot urine P/C ratio be useful in everyday medical practice for the diagnosis and monitoring of patients with renal insufficiency and other disease states that are accompanied by proteinuria, thus eliminating the need for a 24-h urine collection?

• How does the value of a spot urine P/C ratio correlate with protein excretion in a 24-h urine collection?

• Do factors such as patient’s race, course and multiplicity of pregnancy, type of sample used, and method used affect the useful of the spot urine P/C ratio for assessing proteinuria?

Search strategy

We searched the PubMed and Google Scholar databases for articles publish from 1975 to the end of 2019. We began the search at 1975, because that year marked the start of studies concerning the analysis of P/C ratio, its application in comparison with the 24-h urine collection, and methods of urine protein measurement. No language restrictions were applied, but most of the cited papers were published in English. We have included 138 references in our work. More weight was given to the articles published in the last 10–20 years, which represented more than 85% of all publications.

We used the following keywords and terms: proteinuria, spot urine protein to creatinine ratio, spot urine P/C ratio, protein creatinine index, PCR (protein to creatinine ratio), P/C ratio and methods, Jaffe versus enzymatic creatinine methods, urine protein methods, spot urine protein to creatinine ratio versus ACR (albumin to creatinine ratio), proteinuria versus albuminuria, limitations of the P/C ratio.

We included studies if they covered any of the following aspects:

• ratio test: the spot urine protein to creatinine ratio (P/C ratio) and/or albumin to creatinine ratio (ACR)

• “gold standard” of method of protein estimation: the protein (total protein/albumin) excretion in 24-h urine collection

• diagnostics usefulness of spot urine P/C ratio in kidney disease, nephropathy, glomerulonephritis, hypertension, preeclampsia, eclampsia, diabetic nephropathy, lupus nephritis, MM, children with proteinuria

• spot urine P/C ratio for assessing proteinuria depending on selected factors: race, type of sample used, method used, course and multiplicity of pregnancy, the glomerular filtration rate (GFR) value, stage of chronic kidney disease

• threshold value: spot urine P/C ratio in physiological and pathological states

We excluded studies that evaluated the protein to creatinine ratio and ACR estimated with semiquantitative methods (dipstick methods) or that were calculated from the 24-h urine collection.

In the manuscript and tables, all values are given in SI units; conventional units are given in the brackets (e.g. mg/mg for the urine P/C ratio or mg/g for ACR).

Methodology aspects

Proteinuria evaluation

The “gold standard” method for the assessment of proteinuria is the quantitative determination of the protein concentration in a 24-h urine collection, the so-called “timed urine collection”. It allows protein fluctuations during the day and day-to-day readings to be taken into account. However, it is time-consuming and troublesome for the patients and sometimes it involves errors that can significantly affect its accuracy (Figures 1 and 2) [4–6].

Figure 1. Reasons of diagnostic inaccuracy using 24-h urine collection [4–6].

Figure 2. Advantages and disadvantages of 24-h urine collection [4–6].

An alternative, fast and simple method of quantitative evaluation of urinary protein excretion may be the assessment of the P/C ratio, which is obtained by dividing the protein concentration by the creatinine concentration in a spot urine sample.

More than 30 years ago Ginsberg et al. [7] used the spot urine P/C ratio to estimate proteinuria quantitatively on a daily basis. The authors assumed that in patients with stable renal function and reasonably constant excretion of creatinine [mg/dL] and protein [mg/dL] in the urine during the day, the calculation of the protein to creatinine ratio eliminated the need to measure the protein concentration in a 24-h urine collection. In their study, they observed high correlations of the P/C ratio results determined in three urine samples collected during the day with the quantitative protein excretion in the 24-h urine collection (r = 0.96, r = 0.93, r = 0.94, respectively) and slightly weaker correlations when using a sample taken at night and upon wakening (r = 0.78 and r = 0.83, respectively). The authors pointed out that these slight differences may have resulted from changes in the body position (orthostatic proteinuria) and from various physical activities during the day and night. They also emphasized that the spot urine P/C ratio results should be interpreted with caution and its possible limitations should be considered, especially in young, muscular men and in young, frail women/children (debilitated, weakened), in whom excretion of urinary creatinine may not be constant and may differ significantly from normal, as it depends on age, sex, weight, and muscle mass [7].

Numerous studies have investigated the use of the spot urine P/C ratio for the quantitative assessment of proteinuria not only in kidney disease but also in other disease states that are accompanied by proteinuria [1,7–13].

In 2002, the USA National Kidney Foundation (NKF), in its recommendations for the evaluation, classification and stratification of chronic kidney disease, indicated that the screening dipstick test should be used in a spot urine sample for screening and monitoring of proteinuria in children and adults. It is not usually necessary to collect timed samples (overnight or 24-h). In contrast, patients with a positive dipstick test (1+ or greater) should have proteinuria confirmed using a quantitative measurement: spot urine P/C ratio or ACR within a period of 3 months. For adults with an increased risk of chronic kidney disease, the P/C ratio can be used to quantify proteinuria if ACR is high (56.5 to 113 mg/mmol) [500–1000 mg/g] [14,15].

Threshold value for spot urine P/C ratio

The spot urine P/C ratio may be expressed in different units, i.e. in mg/mg, mg/g, mg/mmol, and it is often reported simply as a numerical value (without units, especially when it is reported in mg/mg). The use of different units can lead to difficulties in interpreting the results for both the physician and the patient. Therefore, in this review, for comparison of the P/C ratio cutoffs in different disease states, all P/C ratio values were converted to SI units (mg/mmol), while conventional unit (mg/mg) values are given in brackets.

Ginsberg et al. and Kristal et al. indicated a value <20 mg/mol (0.2 mg/mg) as a reference value for the P/C ratio and >350 mg/mmol (3.5 mg/mg) for the differential diagnosis of nephrotic proteinuria [7,16]. Chitalia et al. gave slightly different cutoff values, i.e. 26 mg/mmol (0.26 mg/mg) for the detection of proteinuria and 320 mg/mmol (3.2 mg/mg) for nephrotic proteinuria [17]. Similar cutoff points ​​of P/C ratio were also indicated by Patil et al.: >20 mg/mmol (0.2 mg/mg) and >30 mg/mmol (0.3 mg/mg) as values ​​corresponding to proteinuria of >0.2 g/d and >0.3 g/d, respectively [5]. These authors also found that a P/C ratio value of 12 mg/mmol (0.12 mg/mg) (sensitivity 100%, specificity 98%) distinguished abnormal protein concentration from normal protein concentration in the general population. Patil et al. additionally established a cutoff value of 16 mg/mmol (0.16 mg/mg) to distinguish normal from elevated urine protein concentration in patients with low risk renal disease; this cutoff had almost ideal sensitivity and specificity (95% and 100%, respectively) and no false positive results. In order to differentiate nephrotic proteinuria, they used the value of 260 mg/mmol (2.6 mg/mg) (sensitivity 100%, specificity 96%). When clinical suspicion of nephrotic syndrome was low, they recommended using a cutoff of 320 mg/mmol (3.2 mg/mg) (sensitivity 80%, specificity 100%) [5].

In the 2004 Australian guidelines, Caring for Australians with Renal Impairment (CARI), proteinuria was classified as a spot urine P/C ratio ≥20 mg/mmol (0.2 mg/mg), which corresponded to an ACR of ≥3.4 mg/mmol (30 mg/g), and the P/C ratio was recommended as the initial test for evaluation of proteinuria in high risk populations (hypertension patients, and patients with known vascular disease and a family history of renal disease) [18].

Table 1 presents the 2012 National Kidney Foundation (USA) guidelines for defining the three categories of albuminuria and the corresponding values ​​of proteinuria measured by various methods: albumin excretion rate (AER), ACR, P/C ratio, and protein excretion rate (PER) [19]. However, such diversity in the reporting of proteinuria/albuminuria can be misleading and difficult to interpret.

Table 1. Albuminuria categories (A1, A2, A3) and relationship among proteinuria measured by different tests. Modification based on [19].

	Units	Categories
		A1 Normal to mildly increased	A2 Moderately increased	A3 Severely increased
AER	mg/24h	<30	30–300	>300
PER	mg/24h	<150	150–500	>500
ACR	mg/mmol	<3	3–30	>30
	mg/g	<30	30–300	>300
P/C ratio	mg/mg	<0.15	0.15–0.50	>0.50
	mg/mmol	<15	15–50	>50
	mg/g	<150	150–500	>500
Urine dipstick test		Negative to trace	Trace to 1+	1+ or greater

ACR: albumin-creatinine ratio; AER: albumin excretion rate; P/C ratio: protein-creatinine ratio; PER: protein excretion rate.

On the other hand, the 2014 National Institute for Health and Care Excellence Guidelines (NICE, UK) emphasized that there is no consistent definition of proteinuria. The upper limit is about 0.15 g/d, which is equivalent to a P/C ratio of 15 mg/mmol (0.15 mg/mg) [20].

Methods of protein and creatinine evaluation

To our knowledge, no studies in the literature have assessed creatinine and protein levels using different methods and comparing how these affect spot urine P/C ratio results. However, our review of the literature demonstrated the diversity of methods used to assess the spot urine P/C ratio, which may result in variable determinations between laboratories. Some publications lacked information on the protein and creatinine assessment methods used to calculate the P/C ratio [2,7,21].

Protein

For protein concentration evaluation, a large variety of methods have been used, largely because there is no reference method. It should be noted that dry chemistry, turbidimetric and dye-binding methods do not give comparable analytical sensitivity and specificity for all proteins (Table 2). Urine is a complex biological fluid, and there is high intra- and inter-individual variability in the excretion rate of different proteins [22]. High analytical variability was noted among the colorimetric and turbidimetric methods used for total urinary protein concentration evaluation [22]. The biuret method with ethanolic phosphotungstic acid as precipitant correlated well with the Coomassie Brilliant Blue method, but poor relationships were observed with the Ponceau S and biuret-trichloroacetic acid methods [23]. Most methods for total urine protein evaluation react more strongly with albumin than with globulins [24–26]. Moreover, in the normal physiological state, the protein concentration in the sample is low and some methods are not sensitive enough to detect them. In the case of pathological samples, where the protein can be very elevated, it may be necessary to dilute samples because the protein concentration is higher than the linearity of the method. Other factors that may affect the precision and accuracy of urine protein results include different amounts and composition of proteins from sample-to-sample, the presence of non-protein interfering substances, and the high content of inorganic ions [27,28]. An additional problem is the lack of a standard reference material (calibrator material) for total protein evaluation in the urine [25]. The study of Marshall and Williams [29] indicated that the use of urinary protein (rather than human albumin) as a calibrator significantly improved comparability between the Coomassie Brilliant Blue, pyrogallol red-molybdate, trichloroacetic acid, and benzethonium chloride protein methods and, to a lesser extent, comparability with the trichloroacetic acid, and Ponceau S methods.

Table 2. Limitations of methods used for urine protein and creatinine evaluation.

Protein evaluation methods	Limitations
Dry chemistry, semi-quantitative method Reaction with pyrocatechol violet-dye	Dedicated mainly to albumin High risk of false positive results in the case of: alkaline samples, samples with the presence of urease-positive bacteria alkalizing urine, when the patient takes the quinine derivatives
Turbidimetry Sulfosalicylic acid Trichloroacetic acid Benzethonium chloride Ammonium chloride	Lack of standardization of the method resulting from: using different precipitants with different concentrations lack of a constant reaction temperature no constant reaction time from the addition of a precipitating substance to the reading affected by turbidity of the samples or by xanthochromia
Dye-binding methods Pyrogallol red	Overestimated result in: the presence of high concentration of iron (II) ions hematuria the case of taking: acetaminophen (Paracetamol), certain antibiotics from the group of penicillin and aminoglycosides
Other methods Ponceau-S Coomassie Brilliant Blue G-250	Difficult to perform, requires decantation Incomplete precipitation of low molecular weight proteins by trichloroacetic acid
Creatinine evaluation methods	Limitations
Colorimetric method Enzymatic Jaffe	The analytical specificity, sensitivity and analytical precision is systematically better for the enzymatic assays compared to the Jaffe assays Picrate (Jaffe method) can also react with pseudochromogens: acetoacetate, pyruvate, ketoacids, proteins, glucose, ascorbic acid, bilirubin or even some drugs

Analysis of data from French laboratories showed that for the assessment of protein concentration in urine, the turbidimetric method with benzethonium chloride and colorimetric method with pyrogallol red were the most commonly used methods. The pyrogallol red method was characterized by a high coefficient of variation (CV) between laboratories. In turn, the pyrocatechol violet method gave urinary protein concentration results that were 10–15% higher compared to those obtained with the pyrogallol red method. In contrast, the pyrogallol red method gave results 10–20% higher than those obtained with the benzethonium chloride method [22]. These differences may be related to the limitations of existing pyrogallol red methods: variability of reagent composition between manufacturers (such as the presence or lack of sodium dodecyl sulfate), different calibration materials, or underestimation of the results when free light chains are present [30,31]. On the other hand, according to Pupkova and Prasolova [32] the pyrogallol red-molybdate method is superior to the sulfosalicylic acid method.

Data from the United Kingdom National External Quality Assessment Scheme of urinary total protein measurement indicated that the CV between laboratories for variety of methods used was 12.4% [27]. In practice, this means that a patient could have a total urine protein concentration result of 200–400 mg/L or a spot urine P/C ratio of 30–60 mg/mmol (0.3–0.6 mg/mg) [27]. Currently there is no reference method for urine protein evaluation; as well, calibrators need to be improved to reduce interlaboratory CVs. Therefore, it is not surprising then Labquality (out-of-laboratory control program, https://www.labquality.fi/en/eqas, http://sowa-med.pl/wp-content/uploads/2016/05/Zakresy-błędu-dopuszczalnego-3.pdf) specifies ± 50% as the maximum permissible error for total urine protein.

Creatinine

Urine creatinine concentration is most often assessed by the colorimetric modified Jaffe method, and less frequently by the more expensive enzymatic method [33–38]. In the Jaffe method, creatinine reacts with picrate, giving a yellow-orange color that is proportional to the creatinine concentration. However, this reaction is not entirely specific because picrate also reacts with other compounds known as pseudochromogens (Table 2) [39–43]. Over the past decades, various technical improvements (kinetic and/or rate-blanked assay, compensated Jaffe assay, etc.) have improved the precision of creatinine evaluation using the Jaffe method [39]. Enzymatic assays measuring creatinine concentration use several different enzymes [41,42]. Studies indicated that the analytical specificity, sensitivity and precision are better for the enzymatic assays compared to the Jaffe assays [43,44]. Therefore, Delanaye et al. [39] highlighted that the enzymatic method should be used preferentially in pediatric patients or patients with hyperfiltration and also in specific clinical situations (e.g. jaundice, ketoacidosis) where the Jaffe method is known have interferences. Moreover, random error (analytical imprecision) due to the intrinsic performance of the measurement is lower for enzymatic (CV approx. 2%) than for Jaffe assays (CV approx. 5.5%) [43,45,46]. According to Myers et al. [45], if the creatinine method is calibrated to the isotope dilution mass spectrometry (IDMS) assay, a total analytical error goal for creatinine measurement should not exceed 10%.

It is well established that the most accurate and reproducible method for creatinine concentration measurement is the IDMS assay. Therefore, the Creatinine Standardization Program (https://www.niddk.nih.gov/health-information/communication-programs/nkdep/laboratory-evaluation/glomerular-filtration-rate/creatinine-standardization) has requested manufacturers to standardize their creatinine assays to an IDMS reference measurement procedure to ensure that the same sample will give the same result in any laboratory in the world, whatever method and manufacturer is used. This is possible if all the calibrators are “traceable” to IDMS [47,48]. It should be noted that most enzymatic assays are standardized to IDMS [44,49], but some dry chemistry enzymatic methods and especially Jaffe methods give results that are not standardized to the reference method [50,51]. It should be emphasized that the studies comparing different methods of creatinine concentration evaluation were carried out mainly in serum [39,41,46,48]. Interestingly, the study of Apple et al. [52] did not find that urine creatinine concentration results depended on the method used (enzymatic versus Jaffe versus high-performance liquid chromatography). The study of Srivastava et al. [53] also confirmed that differences in the evaluation of urine creatinine concentration using various methods (Jaffe versus enzymatic versus enzymatic method traceable to IDMS) were rather small, while more significant differences were observed for serum tests. The authors concluded that this may be due to the lower serum creatinine concentration compared to urine, because the majority of analytical methods for creatinine show higher inaccuracy at lower concentrations [53]. In summary, the recommendations for creatinine measurements are to use an enzymatic assay to decrease random errors and an IDMS traceable assay to decrease systematic errors [39,44,49].

Given the above, the variety of methods employed, which are subject to certain limitations, do not allow comparison of P/C ratio results between laboratories. Therefore, for the diagnosis of proteinuria, a physician using the P/C ratio should use results obtained in a single laboratory using the same analytical conditions. Understanding the methodological limitations and the use of a single laboratory for analysis should ensure the suitability of P/C ratio measurements in everyday medical practice and its application to diagnosis, monitoring and evaluation of the effectiveness of the treatment undertaken [24–28,31].

Clinical findings

Spot urine P/C ratio and kidney diseases

Proteinuria is the main symptom of kidney disease, and proteinuria monitoring is necessary for patients with chronic kidney disease (CKD). It is worth noting that not all patients with CKD have proteinuria, whereas its presence allows the identification of patients who are at significantly more risk of disease progression. Literature data confirms that the evaluation of the P/C ratio in a single urine sample is a good alternative to a 24-h urine collection for the assessment of proteinuria, especially in patients with renal insufficiency [17,21,27,54].

Chitalia et al. [17] in patients with glomerular renal failure evaluated the P/C ratio in random urine samples taken at two times during the day and observed an almost perfect correlation in both samples with the protein concentration in 24-h urine collections (r = 0.97 and 0.99, p < 0.05) [17]. Studies of Montero et al. also indicated a positive correlation of the results of the spot urine P/C ratio with the protein concentration in 24-h urine collections in patients with different degrees of kidney failure. The best correlation of spot urine P/C ratio results with protein concentrations in 24-h urine collections (r = 0.828, p < 0.001) was observed when proteinuria ranged from 0.3–3.5 g/d (300–3449 mg/d). Interestingly, for protein concentrations below 0.3 g/d (300 mg/d), the correlation was lower (r = 0.498, p < 0.001), while in the course of nephrotic syndrome, no correlation was observed [54]. Also, Jyoti et al. observed excellent correlation (r = 0.93, p < 0.001) of the spot urine P/C ratio (557 mg/mmol, 5.57 mg/mg) with the protein concentration in 24-h urine collections (5.71 g/d) in patients with different types of glomerulonephritis [55].

In turn, Kristal et al. found a high correlation of spot urine P/C ratio results with protein excretion in 24-h urine collections and observed that it did not depend on the severity of proteinuria, while it was slightly dependent on the glomerular filtration rate [16]. In a study conducted by Ahmed et al., in the group of patients with different types of glomerulonephritis, positive correlations of spot urine P/C ratio results with protein concentrations in 24-h urine collections were found in relation to GFR values [21]. A high degree of correlation (r = 0.987) between these two results was observed in patients with GFR in the range of 30–59 ml/min/1.73m², while moderate correlation (r = 0.535) was seen in patients with advanced renal failure (GFR < 15 ml/min/1.73m²). In addition, the authors found the following correlations of the spot urine P/C ratio with protein concentration in 24-h urine collections in the following ranges: <0.2 g/d (r = 0.721); 0.2–1.0 g/d (r = 0.788); 1.0–3.4 g/d (r = 0.688); 3.5–6.0 g/d (r = 0.728). Ahmed et al. emphasized that they did not observe a correlation between these tests when the protein excretion reached values above 6 g/d [21]. Ali et al. divided the patients whose protein concentrations exceeded 0.5 g/d into five subgroups depending on their GFR values based on the ranges used to classify CKD; they found a significant positive correlation between the spot urine P/C ratios and the protein concentrations in 24-h urine collections regardless of the GFR value (r = 0.80 to r = 0.96). They emphasized that the mean protein concentration in the 24-h urine collections (3.49 ± 3.0 g/d) and the P/C ratio values 536 ± 465 mg/mmol (5.36 ± 4.65 mg/mg) were not similar only in the group of patients with CKD stage 5, and they hypothesized that the spot urine P/C ratio may overestimate protein excretion at lower levels of GFR [33]. On the other hand, Price et al. conducted a meta-analysis of the results of 16 studies (1,781 patients) concerning the usefulness of the spot urine P/C ratio and protein evaluation in 24-h urine collections in patients with renal disease or with preeclampsia. The authors concluded that the spot urine P/C ratio was useful primarily for the exclusion of the presence of significant proteinuria and its use should reduce the number of unnecessary 24-h urine collections. However, when the P/C ratio results were above the cutoff value, they stated that the quantitative assessment of proteinuria in 24-h urine collections should be performed [56].

The usefulness of the spot urine P/C ratio to assess proteinuria compared to protein concentration in 24-h urine collections was also analyzed in patients who underwent renal transplantation. In the majority of these studies, a strong or very strong positive correlation between these two tests was observed (r = 0.772 to r = 0.998) [34,57–61]. Wahbeh et al. emphasized that the spot urine P/C ratio was a precise, convenient and reliable method for estimating urinary protein excretion in patients after kidney transplantation, especially those with low proteinuria [34]. The authors also pointed to the excellent diagnostic usefulness of a cutoff >43 mg/mmol (0.43 mg/mg) to detect proteinuria in these patients (area under the receiver operating characteristic [ROC] curve, AUC = 0.967, p < 0.001, sensitivity 100%, specificity 90%, accuracy 90%). Additionally, using the Bland-Altman plot, they demonstrated that the limits of agreement for both proteinuria evaluation methods was +0.77 to −1.06 g/d. At the same time, they emphasized that their study focused on assessing the correlation of both tests, and not determining the limits of agreement between them [34].

In contrast, Talreja et al. indicated that ACR, P/C ratio and albumin and protein excretion in 24-h urine collections were similar predictors of doubling serum creatinine, kidney transplant rejection and patient death [60]. Therefore, a random urine sample was an appropriate alternative to a 24-h urine collection to measure protein excretion in patients after kidney transplantation [60]. On the other hand, Akbari et al. recommended the verification of P/C ratio and ACR measurements by means of protein concentration in 24-h urine collections before important diagnostic and therapeutic decisions were taken (e.g. before biopsy or before changes of immunosuppressive agents) [61].

Spot urine P/C ratio and lupus nephritis

Lupus nephritis may occur in systemic lupus erythematosus at the onset of the disease or many years after the diagnosis. Proteinuria, in addition to abnormalities in the urine sediment (granular casts, cellular casts, hematuria) and impaired renal function, are the main findings in lupus nephropathy. Therefore, the assessment of proteinuria in patients with lupus nephritis is a screening test, but it is also important to monitor the treatment and progression of kidney disease [8,62,63].

Leung et al. recommended the use of the spot urine P/C ratio both as a screening test and for proteinuria monitoring in patients with lupus nephritis [64]. Medina-Rosas et al. conducted a meta-analysis of the results of 13 studies on the use of the spot urine P/C ratio in the course of lupus nephropathy [63]. Based on 8 studies, they noticed a high overall correlation (r = 0.8) between the P/C ratio and protein concentration in 24-h urine collections; however, the correlation decreased significantly with severe proteinuria. A study by Birmingham et al. [65], which was included in the meta-analysis [63], found especially poor correlation between these two tests (r = 0.5, p = 0.001) if the proteinuria ranged from 0.5–3.0 g/d. In clinical practice, a high degree of correlation of results for these two tests was found when proteinuria exceeded 1 g/d. The meta-analysis emphasized that several pitfalls that could lead to erroneous conclusions regarding the suitability of the P/C ratio. Firstly, the spot urine P/C ratio was validated to differentiate nephrotic and non-nephrotic proteinuria, not to assess accurately the urine protein concentration [63]. Secondly, the P/C ratio does not take into account changes in protein excretion during the day and between-day. Thirdly, the size of the patient groups in many studies was mostly below 50, which did not allow the formulation of unambiguous conclusions. Fourthly, the studies analyzed only the correlation between the spot urine P/C ratio and protein concentration in 24-h urine collections and did not analyze the agreement of the two methods; the Bland-Altman plot results showed the P/C ratio was not a useful tool for accurate proteinuria measurement during the day [63].

On the other hand, Solorzano et al. [66] observed a very strong correlation of protein concentrations in 24-h urine collections with spot urine P/C ratio results (r = 0.901) in patients with lupus nephropathy. However, the authors emphasized that the absolute values ​​differed between these two methods (80 mg/mmol (0.8 mg/mg) for P/C ratio vs. 1.0 g/d), which did not allow the interchangeability of the tests, especially when they were to be used to determine the activity of disease. They suggested repeating the protein urine evaluation two to three times using both methods if it is necessary to select one test for ongoing assessment of proteinuria and disease activity in a given patient [66]. Choi et al. [67] indicated that the spot urine P/C ratio was an excellent alternative to 24-h urine collections in the case of clinically significant proteinuria in patients with lupus nephropathy. In their opinion, when the spot urine P/C ratio was >100 mg/mmol (1.0 mg/mg), then the patient could directly qualify for renal biopsy, whereas when the P/C ratio was between 50–100 mg/mmol (0.5–1.0 mg/mg), the proteinuria should be confirmed in a 24-h urine collection before any important decision was made [67].

The Kidney Disease Improving Global Outcomes and the American College of Rheumatology and the European League Against Rheumatism/European Dialysis and Transplant Association recommend the use of spot urine P/C ratio to assess proteinuria in clinical trials and to monitor the course of lupus nephropathy [68–70].

Spot urine P/C ratio and diabetic nephropathy

Diabetic nephropathy, which accounts for about 20% of cases of chronic renal failure, is the most common cause of end-stage renal failure [71]. Therefore, early detection and quantitative assessment of proteinuria in the course of diabetes is important both in diagnosis and in treatment. Only single studies on the usefulness of the spot urine P/C ratio in the estimation of proteinuria in patients with diabetes are available in the literature [72], and in most cases the evaluation of albumin concentration is used [73–75]. Biradar et al. [72] observed a strong correlation (r = 0.925, p < 0.001) between the P/C ratio (130 ± 155 mg/mmol, 1.3 ± 1.55 mg/mg) and protein concentration in 24-h urine collections (1.6 ± 1.7 g/d). ROC curve analysis showed that a cutoff of 30 mg/mmol (0.3 mg/mg) for the P/C ratio was an excellent predictor of significant proteinuria in diabetic nephropathy (AUC = 0.947, sensitivity 81%, specificity 100%), and thus the P/C ratio might be a reasonable and faster alternative for the detection of proteinuria in these patients [72]. Yadav et al. also observed a strong correlation of the spot urine P/C ratio values with proteinuria results from 24-h urine collections in patients with diabetes (r = 0.892, p < 0.001). They indicated the value of 15 mg/mmol (0.15 mg/mg) as the P/C ratio cutoff point to detect significant proteinuria in these patients (AUC = 0.88, sensitivity 97%, specificity 74%) [10].

Interestingly, Ren et al. [76] conducted a meta-analysis that included 32 studies and over 2,500 patients with diabetic nephropathy that assessed proteinuria in 24-h urine collections; they showed that the use of the spot urine P/C ratio in this group of patients was rare and suggested the need for research in large groups of patients to determine acceptable threshold values of the P/C ratio for diabetic patients to detect proteinuria [76].

Spot urine P/C ratio and hypertension

The most common causes of proteinuria in patients with primary and secondary hypertension are lack of diagnosis and treatment and inadequate therapy, but also the use of antihypertensive drugs may increase proteinuria, e.g. angiotensin converting enzyme inhibitor administered with an angiotensin receptor antagonist. Persistent non-compensated hypertension can lead to hypertensive nephropathy and consequently end stage renal disease (ESRD) [77].

Studies of Toto et al. [78] in a population of African-Americans with longstanding hypertension who received one of three hypertensive drugs (metoprolol, amlodipine, ramipril) show that two-thirds of participants had spot urine P/C ratios below 22 mg/mmol (0.22 mg/mg). Urinary protein excretion was increased regardless of blood pressure correction and was higher in patients with hypertension receiving amlodipine. The authors did not notice differences in the rate of GFR decline in the group where the P/C ratio was ≤22 mg/mmol (0.22 mg/mg) regardless of the treatment involved. In contrast, in the group with P/C ratios >22 mg/mmol (0.22 mg/mg) receiving amlodipine, a decrease in GFR was 2-times faster compared to patients receiving ramipril and about 1.2 times faster compared to patients treated with metoprolol. The authors emphasized that the urine protein excretion in hypertensive patients was an independent prognostic factor and should be controlled along with blood pressure, because only a reduction in both parameters would result in a slower rate of decrease in renal function and a reduced risk of ESRD [78].

Ahsan et al. evaluated the diagnostic usefulness of the spot urine P/C ratio in predicting proteinuria and nephropathy in hypertensive patients in India. They observed a strong positive correlation between the spot urine P/C ratio and daily proteinuria and showed that the values ​​of the spot urine P/C ratio >20 mg/mmol (0.2 mg/mg) and >30 mg/mmol (0.3 mg/mg) allowed accurate estimation of the occurrence of proteinuria above >0.15 g/d and >0.3 g/d in these patients. In addition, they recommended the cutoff value of the P/C ratio of 80 mg/mmol (0.8 mg/mg) as a good screening tool (sensitivity 95%) for predicting nephropathy in Indian hypertensive patients. In their opinion, the spot urine P/C ratio should be used routinely as a screening test in all patients with hypertension for longer than 5 years, circumventing the inconvenient 24-h urine collection, so that the doctor could make a diagnosis and start treatment earlier [35,36].

Forlemu et al. [13] assessed left ventricular hypertrophy (LVH) in hypertensive patients. They noted increased spot urine P/C ratios in hypertensive patients with LVH (86 mg/mmol, 0.86 mg/mg) compared to individuals with hypertension but without LVH (50 mg/mmol, 0.50 mg/mg). The authors suggested that P/C ratio may be a useful routine screening test in patients with hypertension and LVH, and a useful test for monitoring proteinuria in high-risk patients with untreated hypertension. Ravikumar et al. also emphasized the relationship of the spot urine P/C ratio value >30 mg/mmol (0.3 mg/mg) with the occurrence of LVH, especially in patients with stage 2 hypertension (blood pressure ≥160/100 mmHg) [79].

Spot urine P/C ratio and preeclampsia

Hypertension in or after the 20^th week of pregnancy with proteinuria and organ symptoms may indicate preeclampsia, which is associated with the risk of developing eclampsia [80]. According to the guidelines of the International Society for Hypertension in Pregnancy (ISSHP), diagnosis of pregnancy proteinuria should follow the pattern shown in Table 3.

Table 3. Pregnant proteinuria [80].

Pregnant proteinuria
1. Proteinuria should be pre-evaluated during routine urinalysis by means of a strip test Positive result if ≥1 + [∼300 mg/L (30 mg/dL)]* Negative result**
2.Proteinuria evaluation using the spot urine P/C ratio Abnormal result ≥30 mg/mmol (0.3 mg/mg)
3.Heavy proteinuria >5 g/day***

Note: Proteinuria is not required to recognize preeclampsia.

* Evaluate the P/C ratio.

** Further diagnosis of proteinuria with the P/C ratio is not necessary.

*** It is associated with a severe condition of the newborn.

In severe preeclampsia, hypertension of ≥160/110 mm Hg, proteinuria in the test strip + 3/massive proteinuria >5 g/d and organ symptoms are often found. In such cases, the diagnosis is clear, and treatment should start immediately. The assessment of protein concentration in a 24-h urine collection is difficult to justify due to the long wait time for the result in relation to the complications that may occur at any time. In less severe states, it is often recommended to assess the proteinuria in 24-h urine collections to confirm preeclampsia [81–84].

Since 2001, the ISSHP has recommended the use of a spot urine P/C ratio cutoff point at 30 mg/mmol (0.3 mg/mg) for the classification of proteinuria in women at risk of preeclampsia [82].

In the last decade, there have been many studies on the usefulness of the spot urine P/C ratio as an easy and fast test for the diagnosis of proteinuria accompanying preeclampsia [11,37,84–91]. Lokhande et al. [84] observed a significantly higher P/C ratio in women with mild preeclampsia (68 ± 24 mg/mmol. 0.68 ± 0.24 mg/mg) compared to healthy women (22 ± 7 mg/mmol, 0.22 ± 0.07 mg/mg). In addition, using the ROC curve, they determined the AUC = 0.95 for the P/C ratio cutoff of 30 mg/mmol (0.3 mg/mg), confirming its diagnostic usefulness for classifying proteinuria in women at risk of preeclampsia. However, the authors emphasized that the use of ISSHP-compatible P/C ratio cutoff resulted in failure to identify significant proteinuria in approximately 6% of women (sensitivity and specificity equal to 94%) [84].

Research by Cade et al. [11] also indicated that a spot urine P/C ratio equal to 30 mg/mmol (0.3 mg/mg) allowed the prediction of significant proteinuria in women with preeclampsia (95% sensitivity, 67% specificity). They observed a strong positive correlation of the spot urine P/C ratio with the protein concentration in 24-h urine collections (r = 0.98, p < 0.001). Interestingly, among the women who were assessed by a spot urine P/C ratio and not a 24-h urine collection, they did not find a significant increase in the diagnosis of severe hypertension and the need for magnesium sulfate infusion [11]. A strong correlation between the results of the P/C ratio and the protein concentration in 24-h urine collections (r = 0.896) in women with preeclampsia was also observed by Basharat et al. [85].

There is divergent data in the literature on the threshold value of the spot urine P/C ratio in the diagnosis of preeclampsia. Shreya et al. [86] and Rathindranath et al. [87] indicated that a P/C ratio of 20 mg/mmol (0.2 mg/mg) corresponded to proteinuria at 0.3 g/d and a P/C ratio <19 mg/mmol (0.19 mg/mg) allowed the exclusion of preeclampsia with 100% sensitivity. Another study showed that a P/C ratio ≤20 mg/mmol (0.2 mg/mg) could exclude significant proteinuria (≥0.3 g/d) with 100% specificity, and a P/C ratio ≥40 mg/mmol (0.4 mg/mg) confirmed significant proteinuria in preeclampsia with 100% sensitivity; the authors proposed a P/C ratio of 33 mg/mmol (0.33 mg/mg) as the optimal cutoff [37].

Valdés et al., based on ROC curve analysis, determined that a spot urine P/C ratio of 36 mg/mmol (0.36 mg/mg) was the appropriate limit for the evaluation of proteinuria in the diagnosis of preeclampsia (sensitivity 73%, specificity 91%, positive predictive value [PPV] 95%). In contrast, a high spot urine P/C ratio threshold of 458 mg/mmol (4.58 mg/mg) corresponding to severe proteinuria (sensitivity 100%, negative predictive value [NPV] 100%) was in their opinion an excellent diagnostic tool to rule out severe disease without the need to assess other laboratory parameters, and it also avoided unjustified therapy with magnesium sulfate or urinary catheter placement. The authors also showed that a spot urine P/C ratio of 36 mg/mmol (0.36 mg/mg) was a more diagnostically reliable cutoff (sensitivity 86%, specificity 100%, PPV 100%, NPV 50%) in the assessment of preeclampsia in the course of pregnancies terminated before 34 weeks compared to those above 34 weeks (sensitivity 63%, specificity 90%, PPV 89%, NPV 64%) [88]. In contrast to the above-mentioned examples, Kucukgoz Gulec et al. [89] proposed a threshold value for the spot urine P/C ratio of 53 mg/mmol (0.53 mg/mg) for proteinuria >0.3 g/d (sensitivity 81%, specificity 93%).

Interesting data was presented by Bhatti et al. [90] who emphasized the dependence of P/C ratio on ethnicity. The authors indicated that a spot urine P/C ratio cutoff of 30 mg/mmol (0.3 mg/mg) gave unsatisfactory proteinuria detection results in 41% of African-American and 23% of Caucasians pregnant women. In the Caucasian population, they proposed a cutoff of 21 mg/mmol (0.21 mg/mg) as a value that allowed detection of proteinuria with 100% sensitivity. In addition, they emphasized that the use of a spot urine P/C ratio in these women significantly reduced the cost of diagnostics [90]. In contrast, in the population of pregnant African-American women, they suggested that quantitative assessment of protein in 24-h urine collection, and not the spot urine P/C ratio, should be used for the assessment of proteinuria. This is because African-American women excrete more creatinine in the urine compared to Caucasian women with similar height and weight [92,93].

The study of Gaddy-Dubac et al. [94] showed that the use of a P/C ratio cutoff of >30 mg/mmol (0.3 mg/mg), according to ISSHP, resulted in less than acceptable accuracy of proteinuria estimation at 0.3 g/d (sensitivity 33%, specificity 85%). The authors suggested that ethnic diversity and different courses of pregnancy might contribute to unsatisfactory results in the use of the P/C ratio [94].

Various reports in the literature have shown that the values of the P/C ratio depended on the time of day at which the urine samples were collected [91,95]. In women with primary hypertension and the risk of developing preeclampsia, Vardonk et al. [95] observed the lowest P/C ratio of 60 mg/mmol (0.6 mg/mg) at 08:00 h, the highest value of 79 mg/mmol (0.79 mg/mg) at 12:00 h, and a value of 73 mg/mmol (0.73 mg/mg) at 17:00 h. In order to detect proteinuria, they determined the best cutoff for the P/C ratio depending on the time of urine collection: 50 mg/mmol (0.5 mg/mg) at 08:00 h, 42 mg/mmol (0.42 mg/mg) at 12:00 h, and 35 mg/mmol (0.35 mg/mg) at 17:00 h. They observed that fluctuations in the P/C ratio values based on the time of urine collection were explained by differences in the physical activity of the women during the day, variable blood pressure, emotional state and posture. They emphasized that for these women, P/C ratio tests were important for the rapid identification of clinically important proteinuria at any time of the day [95]. Chandramathy et al. [91] also assessed the time of day for P/C ratio measurements that best correlated with the protein concentration in 24-h urine collections. They observed P/C ratios of 47 mg/mmol (0.47 mg/mg) for samples taken in the morning (sensitivity 75%, specificity 75%); 40 mg/mmol (0.4 mg/mg) for samples taken at noon (sensitivity 89%, specificity 91%); and 45 mg/mmol (0.45 mg/mg) for samples taken at night (sensitivity 86%, specificity 80%). Surprisingly, based on the findings of Chandramathy et al., the best time for urine collection for a P/C ratio evaluation was 12:00 h [91].

Haghighi et al. [96] also evaluated the P/C ratio in urine samples for the estimation of proteinuria in the risk assessment of preeclampsia at several different time points in the day. They found a positive correlation of P/C ratio determined in samples collected over 4 h, 8 h, 12 h (day) and 12 h (night) compared to the assessment of proteinuria in 24-h urine collections. Moreover they determined the best cutoff values for the P/C ratio in individual samples: the P/C ratios were: for a 4-h sample, 28 mg/mmol (0.28 mg/mg) (sensitivity 88%, specificity 98%); for an 8-h sample, 24 mg/mmol (0.24 mg/mg) (sensitivity 95%, specificity 95%); for a 12-h (day) sample, 25 mg/mmol (0.25 mg/mg) (sensitivity 97%, specificity 98%); and for a 12-h (night) sample, 23 mg/mmol (0.23 mg/mg) (sensitivity 95%, specificity 97%). According to the authors, the measurement of P/C ratio in several-hour urine samples might be a good alternative to 24-h urine collections for detecting proteinuria in pregnant women with suspected preeclampsia, especially when there is insufficient time for a 24-h urine collection [96].

Studies have also assessed whether proteinuria >0.3 g/d or a spot urine P/C ratio ∼30 mg/mmol (0.3 mg/mg) is a good criterion for the assessment of pathology during pregnancy and whether such proteinuria can be observed in a normal pregnancy. Tanamai et al. [97] found that in healthy pregnant women with normal, uncomplicated pregnancy, a P/C ratio ≥30 mg/mmol (0.3 mg/mg) is very common just before delivery, at the term and during the peripartum period. The highest values ​​of the P/C ratio were observed 6 h after delivery. Therefore, they emphasized that a P/C ratio of 30 mg/mmol (0.3 mg/mg) in healthy pregnant women with an uncomplicated pregnancy during this period may be an unreliable criterion for the assessment of proteinuria in the diagnosis of preeclampsia, and that separate cutoff values ​​for women examined just before delivery, during and immediately after delivery should be established [97]. In another study [98], post-delivery spot urine P/C ratio values ​​in healthy pregnant women were found to be significantly higher compared to pre-delivery values ​​and exceeded the 30 mg/mmol (0.3 mg/mg) threshold. The P/C ratio value 2 h after vaginal delivery was 46 mg/mmol (0.46 mg/mg), and 4 h after cesarean delivery, 54 mg/mmol (0.54 mg/mg). On the basis of these results, the authors emphasized that spot urine P/C ratio measurements were unreliable in the immediate postpartum period regardless of mode of delivery. Therefore, P/C ratio cutoff values other than the 30 mg/mmol (0.3 mg/mg) should be considered for suspected preeclampsia in close proximity to delivery. During delivery and in the postpartum period in patients who were administered magnesium sulfate prophylactically, the spot urine P/C ratio may not be reliable in assessing proteinuria [98].

Additionally, in the risk assessment of preeclampsia, it is important to know whether the spot urine P/C ratio is determined in the urine of patients who have singleton or multiple pregnancies. Osmundson et al. [99] emphasized that women pregnant with twins could excrete more protein daily and more often have proteinuria, but that they were not always hypertensive. Therefore, other criteria for the diagnosis of preeclampsia should be developed for women in twin pregnancies [99]. Other authors indicated that, while the values of the P/C ratio in singleton and twin pregnancies in the initial period may be similar, they were significantly higher in twin pregnancies at the end of the third trimester (34–38 weeks of pregnancy) [100].

Independently and in parallel with the P/C ratio, the ACR may be used to estimate proteinuria during pregnancy [101–103]. A strong positive correlation (r = 0.95, p = 0.001) was found between these two tests. An ACR of 13.4 mg/mmol has been shown to correspond to a P/C ratio of 30 mg/mmol (0.3 mg/mg). The authors suggested that clinicians should use the test that is less expensive, that has fewer methodological limitations, and that they are more familiar with [101]. Interestingly, Waugh et al. [103] observed that ACR had a better diagnostic efficacy (sensitivity 99%) when estimating proteinuria in predicting severe preeclampsia as compared to the P/C ratio (sensitivity >90%).

Spot urine P/C ratio and myeloma kidney

In the course of MM, proteinuria may be the result of the accumulation in the urine of Bence-Jones proteins, i.e. monoclonal immunoglobulin light chains (κ, λ), but also of albumin and other proteins [104]. The International Myeloma Working Group recommends the assessment of proteinuria in 24-h urine collection in the diagnosis of MM and periodically during treatment and monitoring of disease activity, especially because test strips do not detect monoclonal protein [105,106].

The usefulness of the spot urine P/C ratio in estimating proteinuria in patients with MM was first assessed by Wozney et al. [12], who showed a good positive correlation (r = 0.81, p < 0.001) of the P/C ratio with the protein concentration in 24-h urine collections. In patients with MM, for selected protein concentrations in 24-h urine collections, they determined P/C ratio cutoff values: 25 mg/mmol (0.25 mg/mg) for proteinuria ≥0.3 g/d (sensitivity 83%, specificity 81%); 41 mg/mmol (0.41 mg/mg) for proteinuria ≥0.5 g/d; 57 mg/mmol (0.57 mg/mg) for proteinuria ≥1.0 g/d; and 371 mg/mmol (3.71 mg/mg) for proteinuria ≥3.5 g/d. In addition, they noticed that the P/C ratio was significantly higher in patients with positive urine immunofixation (37 mg/mmol, 0.37 mg/mg) compared to patients with negative urine immunofixation (10 mg/mmol, 0.10 mg/mg, p = 0.00), especially in the group of patients with light chain MM. The authors emphasized that the spot urine P/C ratio was useful as a screening test for assessing proteinuria in the course of MM.

To assess proteinuria in three patients with MM, Talbot et al. [107] evaluated the P/C ratio together with routine urinalysis. In one patient, they obtained a P/C ratio of 71 mg/mmol (0.71 mg/mg), while in the other two cases, the P/C ratio was 166 mg/mmol (1.66 mg/mg) and 178 mg/mmol (1.78 mg/mg). High values ​​of the P/C ratio in these patients were associated with positive urine immunofixation in two cases, while in the third patient, urine immunofixation was not performed [107].

The above-mentioned studies demonstrate the possibility of using the spot urine P/C ratio to assess proteinuria in patients with MM, although it should be emphasized that these are single reports and that further evaluation in a larger group of patients is required.

Spot urine P/C ratio and children

Proteinuria in children is usually found accidentally during routine urinalysis and occurs in about 10% of school-age children; however only in 0.1% of them is it confirmed in subsequent laboratory tests [108,109]. The most typical causes are orthostatic proteinuria (induced by standing) or functional (transient) proteinuria [108].

According to Leung et al. [108,110], when the result of the test strip for protein is trace, the test strip for protein should be repeated using the first morning sample; when the result is negative or trace, the test strip for protein should be repeated within a year to check that proteinuria has not recurred; and when the result is positive ≥1+, the test strip for protein should be repeated on a first morning sample and the P/C ratio should be calculated. If the spot urine P/C ratio is ≤20 mg/mmol (0.2 mg/mg), or ≤50 mg/mmol (0.5 mg/mg) for children under 2 years of age, and the other parameters in the urine screening test are normal, then orthostatic or functional proteinuria is very likely, and a checkup should be carried out within a year. However, if spot urine P/C ratio is >20 mg/mmol (0.2 mg/mg), or >50 mg/mmol (0.5 mg/mg) for children under 2 years of age, or the urine test results are abnormal (e.g. hematuria, leukocyturia, active urinary sediment), persistent proteinuria or proteinuria of clinical significance is more likely and testing should be extended to exclude or confirm kidney disease.

Quantitative evaluation of proteinuria in 24-h urine collections in children is very rarely used due to the great difficulty of complete collection [108,110]. Values for urinary protein excretion in children are given in Table 4.

Table 4. Urinary protein excretion in children. Modification based on [108,110].

Type of test	Age/Condition	Values
Spot urine P/C ratio	Children from 6 months to 2 years	>50 mg/mmol or >500 mg/g or >0.5 mg/mg
	Children >2 years	>20 mg/mmol or >200 mg/g or >0.2 mg/mg
	Nephrotic syndrome	>220 mg/mmol or >2200 mg/g or >2.2 mg/mg
Protein concentration in 24-h urine collection	Children <6 month of age	>8 mg/m^2/h or >0.24 g/1.73m^2/d
	Children >6 month of age	>4 mg/m^2/h or >0.15 g/1.73m^2/d
	Nephrotic syndrome	>40 mg/m^2/h or >3 g/1.73m^2/d
	Children Newborns	100–150 mg/m^2/d >0.3 g/m^2/d

NKF KDOQI (Kidney Disease Outcomes Quality Initiative) defined proteinuria in children as normal/minimal when the P/C ratio is <50 mg/mmol (0.5 mg/mg), increased when it is 50–200 mg/mmol (0.5–2.0 mg/mg), and nephrotic when it is >200 mg/mmol (2.0 mg/mg) [19,68]. Newborns are more likely to lose protein in the urine due to reduced reabsorption in the proximal tubules. In contrast, in children above 2 years of age, it is recommended to adopt adult standards for protein in the urine, i.e. <150 mg/L in a spot urine sample and <0.15 g/d [19]. Warady et al. [111] indicated that in children with persistent proteinuria, elevated spot urine P/C ratio values may be an important predictor of chronic renal disease progression (>200 mg/mmol (2.0 mg/mg) in children with non-glomerular kidney disease and >50 mg/mmol (0.5 mg/mg) in children with glomerulonephritis) [111]. Importantly, the use of the P/C ratio to assess proteinuria may be limited in children with low muscle mass and severe malnourishment because the P/C ratio depends on creatinine excretion [112].

Limitations of the spot urine P/C ratio evaluation

The spot urine P/C ratio is a rapid and simple method for estimating the concentration of urinary protein excretion but is not free from limitations. Table 5 presents the most frequently reported limitations of the spot urine P/C ratio evaluation [5,21,29,33,39,45,54,92,93,101,113–116].

Table 5. Limitations of the spot urine P/C ratio evaluation [5,21,29,33,39,45,54,92,93,101,113–116].

Reference	Limitations
[5]	Spot urine P/C ratio does not take into account protein excretion in the urine during the day and from day-to-day
[21]	Lack of correlation between spot urine P/C ratio and protein concentration evaluated in 24-hour urine collection, if daily protein excretion exceed 6 g/day and GFR <15 mL/min/1.73m^2
[92,93]	Spot urine P/C ratio result depends on ethnicity (lower values are observed in the African-Americans, because they excrete more creatinine to urine compared to Caucasians with similar height and weight)
[101]	Spot urine P/C ratio result depends on muscle mass, which indicates its limited use in children/patients with low muscle mass and severely undernourished (the risk of false positive results when the urine sample is excessively diluted or creatinine in the urine ≤2.65 mmol/L (30 mg/dL))
[54,113]	Poor or lack of correlation between spot urine P/C ratio and protein concentration in 24-hour urine collection in patients with minimal change disease or membranous nephropathy with nephrotic-range proteinuria
[114]	The protein concentration may be overestimated or underestimated, especially when urine creatinine levels are ≤3.45 mmol/L (39 mg/dL) or ≥5.48 mmol/L (62 mg/dL)
[33,115]	The spot urine P/C ratio may be overestimated at GFR levels ≤10 mL/1.73m^2
[29,39,45,116]	The diversity of methods used to assess protein and creatine concentration make the comparison of spot urine P/C ratio results between laboratories impossible

GFR: glomerular filtration rate; P/C ratio: protein-creatinine ratio.

Conclusions

The spot urine P/C ratio can be useful in everyday medical practice for the rapid exclusion/confirmation of proteinuria as well as for the quantitative estimation of proteinuria in many clinical conditions associated with proteinuria, i.e. kidney disease, kidney transplantation, lupus nephropathy, hypertensive nephropathy, preeclampsia, diabetic nephropathy, myeloma kidney as well as in proteinuria in children (Table 6) [5,10,12–17,21,27,33–37,54,55,60,61,63,65,67,70,72,79,82,84,86,87,89,90,94,96,97,99,103,108,110,113,115,117–130].

Table 6. Key points in the evaluation of the spot urine P/C ratio in different states [5,10,12–17,21,27,33–37,54,55,60,61,63,65,67,70,72,79,82,84,86,87,89,90,94,96,97,99,103,108,110,113,115,117–130].

References	Key points
	Kidney disease
[16,33]	In patients with lower levels of GFR, especially those with CKD stage 5, urinary protein excretion may be overestimated, indicating spot urine P/C ratio results dependent on the GFR value
[21,54]	The spot urine P/C ratio stronger correlated with protein concentration in 24-h urine collection if the proteinuria ranged from 0.3–3.5 g/day (300–3449 mg/day) compared to patients with nephrotic syndrome >3.5 g/day (>3500 mg/day); no correlation was observed when the protein excretion exceeds 6 g/day
[34,60]	Spot urine P/C ratio is a precise, convenient and reliable method for estimating urinary protein excretion in patients after kidney transplantation, especially with low proteinuria
[61]	Verification of the P/C ratio by means of protein concentration in 24-h urine collection is needed before important diagnostic and therapeutic decisions e.g. Before biopsy or before the change of immunosuppressive agents
[17,21,27,54,55,113]	The evaluation of the P/C ratio in a single urine sample is a good alternative to 24-h urine collection for the assessment of proteinuria, especially in patients with renal insufficiency and with different types of glomerulonephritis (e.g. patients with immunoglobulin A nephropathy)
[118]	Spot urine P/C ratio was as useful as protein/day in predicting outcomes in chronic kidney disease patients and is more convenient for patients, clinicians, and laboratories
	Lupus nephritis (LN)
[67]	The patient with LN should be directly qualified for renal biopsy if the spot urine P/C ratio is >100 mg/mmol (1.0 mg/mg)
[65,117]	Clinical decisions in LN patients should not rely on the spot P/C ratio results, as they can yield false-positive or false-negative diagnoses of glomerulonephritis compared to 24-h urine collection results
[63,64,70,117]	Spot urine P/C ratio is both a screening test and proteinuria monitoring assay in patients with LN, if proteinuria exceeds 1 g/day
	Diabetic nephropathy
[10]	P/C ratio of 15 mg/mmol (0.15 mg/mg) predicted significant proteinuria in diabetic nephropathy (sensitivity 97%, specificity 74%)
[72]	P/C ratio of 30 mg/mmol (0.3 mg/mg) predicted significant proteinuria in diabetic nephropathy (sensitivity 81%, specificity 100%)
[119]	In diabetes mellitus protein evaluation in 24-h urine collection should be performed instead of the spot urine P/C ratio
	Hypertension
[35]	Spot urine P/C ratio of 80 mg/mmol (0.8 mg/mg) with 95% sensitivity predicted nephropathy in hypertensive patients
[36]	Spot urine P/C ratio should be used routinely as a screening test in all patients with hypertension duration >5 years
[13,79]	The relationship between the spot urine P/C ratio values >30 mg/mmol (0.3 mg/mg) and the presence of left ventricular hypertrophy in patients with second stage of primary hypertension was observed
	Preeclampsia
[90]	Spot urine P/C ratio should not be used to detect significant proteinuria in pregnant African-American women
[94]	Spot urine P/C ratio results were unsatisfactory to estimate proteinuria at different course of pregnancy, e.g. medical complication including: overweight, obesity, diabetes, and smoking, symptomatic or asymptomatic bacteriuria
[96]	Spot urine P/C ratio in a several-hour urine samples (4-h, 8-h, 12-h, daily/night sample) could be a good alternative to detect proteinuria in pregnant women with suspected preeclampsia, especially when there was insufficient time to 24-h urine collection
[97]	Spot urine P/C ratio of 30 mg/mmol (0.3 mg/mg) was unreliable for suspect preeclampsia in close proximity to delivery
[99]	In multiple pregnancies women the diagnosis of preeclampsia should be made on P/C ratio cutoff other than 30 mg/mmol (0.3 mg/mg)
[36,37,80,84,86, 87,89,90,103]	For the estimation of proteinuria in women at risk of preeclampsia different cutoff values of the spot urine P/C ratio were recommended: 20 mg/mmol (0.2 mg/mg); 21 mg/mmol (0.21 mg/mg); 33 mg/mmol (0.33 mg/mg); 36 mg/mmol (0.36 mg/mg); >40 mg/mmol (0.40 mg/mg);≥ 40 mg/mmol (0.40 mg/mg); 53 mg/mmol (0.53 mg/mg), however the most frequently recommended cutoff point was 30 mg/mmol (0.30 mg/mg)
[120]	Spot urine P/C ratio in preeclamptic women >900 mg/mmol (9.0 mg/mg) (∼9 g/day), or >500 mg/mmol (5.0 mg/mg) (∼5 g/day) in women over 35 years, was associated with a greatly increased probability of adverse maternal and fetal outcomes
[121]	Spot urine P/C ratio between 30–35 mg/mmol (0.30–0.35 mg/mg) is an optimal threshold value to detect proteinuria >0.3 g/day
[122]	Spot urine P/C ratio was highly predictive to detect proteinuria >1g/day and it could be used as a rapid, alternative test in preeclamptic women
[123]	Spot urine P/C ratio ≥75 mg/mmol (0.75 mg/mg) in normotensive pregnant women was sufficient to detect significant proteinuria
[124]	Spot urine P/C ratio is a better tool for estimation of proteinuria in preeclampsia than the dipstick test
	Myeloma kidney
[12]	Spot urine P/C ratio is a useful screening test for assessing proteinuria in the course of multiple myeloma, taking into consideration that the dipstick test does not detect Bence-Jones protein
	Children
[108,109]	In children the spot urine P/C ratio is a good alternative to 24-h urine collection for evaluation of proteinuria
	In physiology children <2 years of age should have the P/C ratio ≤50 mg/mmol (0.5 mg/mg), while >2 years of age the P/C ratio should be ≤20 mg/mmol (0.2 mg/mg)
	Type of sample
[14,15, 115,125,126]	NKF K/DOQI guidelines and other authors suggested that untimed (spot) urine samples should be used to detect and monitor proteinuria in children and adults; a first-morning sample is preferred, but when it is not available a random sample could be accepted taking into account that daily activity may affect the amount of excreted protein in the urine
[5,127]	The ability to use a random urine sample is an advantage of the P/C ratio evaluation
	Glomerular filtration rate
[16,21,33]	Spot urine P/C ratio results are influenced by the GFR: good correlation between spot urine P/C ratio and the protein excretion in 24-h urine collection was observed in patients with GFR in norm or slightly decreased; weak relationship was observed at GFR <15 mL/min/1.73m^2
	General
[56]	Spot urine P/C ratio is fast and simple method for the exclusion of significant proteinuria and may reduce the number of unnecessary 24-h urine collections the P/C ratio initially was validated to differentiate nephrotic from non-nephrotic proteinuria
[115]	Spot urine P/C ratio could be used as an alternative to urine protein evaluation in 24-h collection in patients with GFR >10 mL/min/1.73m^2
[119]	We still lacks recommendations which methods of urine protein and creatinine concentrations evaluation should be used to minimize interlaboratory differences
[54,90,128–130,135]	The use of spot urine P/C ratio is a convenient and competitively priced method in patients with proteinuria <3.5 g/day

CKD: chronic kidney disease; GFR: glomerular filtration rate; LN: lupus nephritis; NKF K/DOQI: National Kidney Foundation Kidney Disease Outcomes Quality Initiative; P/C ratio: protein-creatinine ratio.

A strong correlation of the spot urine P/C ratio with the protein concentration in 24-h urine collections was observed. The spot urine P/C ratio is a rapid and reliable test that can eliminate the need for a 24-h urine collection. On the other hand, the comparability of both methods has been rarely assessed. In doubtful situations, the assessment of proteinuria based on the P/C ratio should be supported by an evaluation of the protein concentration in a 24-h urine collection (Table 7) [103,131–138].

Table 7. The utility of ACR, the spot urine P/C ratio and the protein/day evaluation in different clinical conditions [103,131–138].

Reference	Utility
[103]	Study analyzed the accuracy of quantitative assessments of the spot urine P/C ratio and ACR at different cutoffs in predicting severe preeclampsia compared to protein/day measurement in pregnant women with hypertension and suspected proteinuria. Authors conclude that in this group of patients proteinuria should be initially checked in dipstick screening test and if the result is positive, the ACR should be performed; in the case of ACR being unavailable an alternative should be the spot urine P/C ratio. The recommended cutoffs for clinically significant proteinuria should be 8 mg/mmol for ACR and 30 mg/mmol (0.3 mg/mg) for the spot urine P/C ratio. Authors do not recommendthe evaluation of protein/day in hypertensive pregnant women.
[131]	Authors conclude that annual screening for dip-stick proteinuria, followed by ACR or the spot urine P/C ratio confirmation, in addition to GFR measurement and treatment with ACEI or ARB for albuminuria or proteinuria is cost-effective for preventing ESRD or death in diabetic patients or in non-diabetic subjects who have hypertension or are aged > 60 years.
[132]	Highlights that there is no consensus regarding the utility of the ACR and the spot urine P/C ratio evaluation, e.g.: the UK CKD guidelines, Scottish Intercollegiate Guidelines Network and Caring for Australians with Renal Impairment Guidelines recommend the use of ACR for diabetic patients, while spot urine P/C ratio should be applied for non-diabetic CKD the Kidney Disease Quality Outcomes Initiative Guidelines recommend ACR, whereas spot urine P/C ratio may be use if the ACR is high >56.5–113 mg/mmol creatinine (500–1000 mg/g)
[133]	Study analyzed 117 spot urine samples to evaluate the relationship between ACR and the spot urine P/C ratio at the cutoffs recommended by the NICE guidance on CKD. Authors did not found discordant results between ACR and the spot urine P/C ratio at cutoffs recommended by NICE: 30 mg/mmol and 50 mg/mmol (0.50 mg/mg), , respectively.
[134]	Based on the literature review authors conclude that ACR might be recommended for the diabetic while the spot urine P/C ratio for the non-diabetic individuals.
[135]	Authors retrospectively analyzed 6842 patients results and found that: the relationship between ACR and the spot urine P/C ratio is non-linear the spot urine P/C ratio correlated more with protein/day as compared to ACR (r = 0.91, p < 0.001 and r = 0.84, p < 0.001, respectively). Moreover, in the urine protein range 0.3–1 g/day there was a significantly greater scatter with ACR. the difference between the spot urine P/C ratio and ACR is less in the ACEi/ARB-treated patients the spot urine P/C ratio is a more sensitive and specific test as compared to ACR for detection of significant proteinuria (>0.5 or >1 g/day) in unselected patients attending a hospital kidney department
[136]	Authors retrospectively analyzed 602 patients with established diagnosis of CKD and found that: ACR strongly, significantly correlated with the spot urine P/C ratio (r = 0.94, p < 0.001) at all albuminuria ranges; on the other hand at normal albuminuria range such correlation was not observed univariate and multivariate linear regression analysis found that similar clinical features were associated with both ACR and the spot urine P/C ratio
[137]	The study analyzed 438 patients with IgAN and showed that: ACR strongly, significantly correlated with the spot urine P/C ratio (r = 0.87, p < 0.001); however better correlation was observed at lower ranges of proteinuria (r = 0.87, p < 0.001) both, ACR and the spot urine P/C ratio correlated with protein/day (r = 0.71, p < 0.001 and r = 0.74, p < 0.001, respectively) ACR had better usefulness in predicting IgAN progression as compared to the spot urine P/C ratio and protein/day
[138]	The CKD in children study showed that there is no difference in the ability of ACR and the spot urine P/C ratio in predicting time to GFR decline of 50%

ACEi: angiotensin-converting-enzyme inhibitors; ACR: albumin to creatinine ratio; ARBs: angiotensin receptor blockers; CKD: Chronic Kidney Disease; GFR: glomerular filtration rate; IgAN: IgA nephropathy; NICE: National Institute for Health and Clinical Excellence.

A spot urine P/C ratio value of 20 mg/mmol (0.2 mg/mg) is the most frequently reported cutoff point for determining the presence or absence of proteinuria, and a P/C ratio of 350 mg/mmol (3.5 mg/mg) is used for the differentiation of nephrotic proteinuria. The ISSHP recommends a spot urine P/C ratio value of 30 mg/mmol (0.3 mg/mg) for the classification of women at risk of preeclampsia.

This review demonstrates the usefulness of the spot urine P/C ratio in various disease states and recommends that this test should be available in every laboratory. The spot urine P/C ratio cutoff to be used should be determined in individual laboratories because it depends on the patient group and the laboratory methodology. The challenge for the primary care physician is to know the limitations of the methods used to determine the protein and creatinine concentrations that are used to calculate the P/C ratio. However, we still lack recommendations about which urine protein and creatinine methods should be used to minimize interlaboratory differences.

Author contributions

JK conceived of the idea for the review and evaluated the searched literature critically. OMK and JT searched the literature. JK and OMK drafted the manuscript, with all authors providing comments. All authors read and approved the final manuscript.

Abbreviations
ACR	=	albumin to creatinine ratio
AER	=	albumin excretion rate
AUC	=	area under the ROC curve
CARI	=	Caring for Australians with Renal Impairment
CKD	=	chronic kidney disease
CV	=	coefficient of variation
ESRD	=	end stage renal disease
GFR	=	glomerular filtration rate
IDMS	=	isotope dilution mass spectrometry
ISSHP	=	International Society for Hypertension in Pregnancy
KDOQI	=	Kidney Disease Outcomes Quality Initiative
LVH	=	left ventricular hypertrophy
MM	=	multiple myeloma
NICE	=	National Institute for Health and Care Excellence
NKF	=	National Kidney Foundation
NPV	=	negative predictive value
P/C ratio	=	protein to creatinine ratio
PCR	=	protein to creatinine ratio
PER	=	protein excretion rate
PPV	=	positive predictive value
ROC	=	receiver operating characteristic

Acknowledgements

We are grateful to Martin Lenkiewicz, MSc, for his language assistance.

Disclosure statement

No potential conflict of interest was reported by the author(s).