Two-hour Urine Collection for Evaluating RenalFunction Correlates With 24-hour Urine

Collection in Pregnant PatientsWayne Evans,* Jon P. Lensmeyer, Russell S. Kirby, Margaret E. Malnory,

and Fredrik F. Broekhuizen

University of Wisconsin Medical School Milwaukee Clinical CampusDepartment of Obstetrics and Gynecology, Milwaukee, Wisconsin

Objective: We sought to determine if 2-h urine collection for the assessment of urine protein excretion andcreatinine clearance correlates with 24-h urine collection in pregnant patients with renal disease.Methods: We enrolled patients of gestational ages ranging from 8–41 weeks, admitted as inpatients and havingundergone evaluation for renal function (n 5 59). We obtained the following samples: 1) 2-h urine, and 2) 24-hurine. We measured serum creatinine concentration, urinary protein, creatinine concentration, and creatinineclearance. The data was analyzed using descriptive analysis, two-way ANOVA, univariate linear regressionanalysis, and Bland-Altman plot comparing the efficacy of 2-h urine results with 24-h urine results.Results: We analyzed the data on 51 patients. Total proteinuria calculated by protein/creatinine (P:C) ratio inthe 2-h group correlated with the total protein measured in the 24-h group (1,840.8 6 786 and 1,944 6 1,060mg [mean 6 SE], respectively, r2 5 0.95, P , 0.0001). Creatinine clearance correlated in the 2- and 24-h groups(111 6 42 and 122.5 6 50 ml/min, respectively; r2 5 0.73, P , 0.001).Conclusion: Two-hour urine sampling offers the same clinical information as 24-h urine collection for the evaluationof renal function. Use of 2-h urine collection reduces the time of evaluation and diagnosis, whereby increasing patientcompliance and reducing errors in performance of the tests. J. Matern.-Fetal Med. 2000;9:233–237.© 2000 Wiley-Liss, Inc.

Key words: renal function; proteinuria; creatinine clearance; pregnancy

INTRODUCTION

Numerous conditions in pregnancy require measurementof urinary protein excretion and creatinine clearance. Stan-dard tests for renal function involve patients collectingurine for a 24-h period. Several studies demonstrated quan-titative measurement of protein excretion by a singlevoided specimen is accurate when compared to the stan-dard 24-h urine collection [1–4]. These studies also dem-onstrated single voided samples and 2-h urine collectionswere time and cost-effective, and subject to less errors incollection and handling when compared to 24-h sampling.Three studies evaluating renal function in critically illpatients showed measuring protein excretion and creati-nine clearance with 2-h urine samples correlated with 24-hsamples [5–7]. Three studies evaluating renal function inpregnant patients demonstrated the accuracy of measuringprotein excretion in a single voided specimen [8–10].

Twenty-four-hour urine sampling in evaluating protein-uria in pregnancy is still widely used despite the tested

accuracy of measuring protein excretion in a single voidedspecimen. We therefore sought to determine if measuringprotein excretion and creatinine clearance in a 2-h urinespecimen correlates with standard 24-h urine collection inobstetric patients evaluated for renal function.

MATERIALS AND METHODS

We conducted a prospective longitudinal study approvedby the Investigational Review Board (IRB) of the Univer-sity of Wisconsin Medical School-Milwaukee ClinicalCampus at Sinai Samaritan Medical Center, Milwaukee,Wisconsin. Each subject enrolled gave written consent.

*Correspondence to: Wayne Evans, Assistant Professor, Depart-ment of Obstetrics and Gynecology, University of Wisconsin MedicalSchool-Milwaukee Clinical Campus, Sinai Samaritan Medical Center,950 N. 12th Street, P.O. Box 342, Milwaukee, WI 53201-0342.

Received 15 October 1999; revised 3 February 2000; accepted 13March 2000

The Journal of Maternal-Fetal Medicine 9:233–237 (2000)

© 2000 Wiley-Liss, Inc.

Over an 18-month period, we enrolled 59 pregnant pa-tients admitted as inpatients and evaluated for renalfunction. Each patient was instructed to collect twourine samples: 1) a voided sample 2 h after a discardedfirst void, and 2) a 24-h voided urine sample. We alsoobtained from each patient venous blood samples tomeasure serum creatinine concentration. After admis-sion to the inpatient unit, the patient voided. Thisspecimen was discarded. We then instructed the patientto remain at bed rest in the left lateral decubitus positionand a voided sample was obtained after 2 h. The 24-hurine sample was obtained with the patient at bed rest inthe left lateral decubitus position. Some patients wereambulatory ad libitum but were instructed to remain inthe left lateral decubitus position when recumbent dur-ing the 24-h collection period.

Urinary and serum creatinine concentrations were mea-sured by the colorimetric picric acid method [11]. Weemployed the Coomassie Blue Technique using the DadeACA Start (Dade International, Newark, NJ) to deter-mine urinary protein concentration [12]. We determinedadequate urine samples quantitatively for the 2-h and 24-hsamples as $15 ml/h since this is reported to be the phys-iologic and clinical standard for the low end normal rangefor urine output [13]. We also determined qualitatively thatthe timed urine samples were adequate if the creatininelevel was at least 10 mg/kg of prepregnancy weight [12]. Wecalculated creatinine clearance for both the 2-h and 24-hsamples using the standard formula:

Urine creatinine ~mg/dl!Serum creatinine ~mg/dl!

3volume ~ml!

collection time ~min!3

1.73 m2

sa ~m2!

where 1.73 m2 is normal surface area and sa is the patient’ssurface area. We also calculated protein excretion by twomethods: 1) direct quantitation of protein in the 24-hsamples using the following formula:

urine protein (mg/dl 3 volume (ml)100

and 2) calculating the urinary protein/creatinine ratio inthe 2-h samples:

urine protein/urine creatinineestimated 24 hour creatinine clearance

where estimated creatinine clearance was determined byusing the Cockcroft-Gault (CG) formula specific forwomen:

(140 2 age [years]) 3 weight (kg) 3 0.8572 3 serum creatinine (mg/dl)

We used two-way ANOVA to determine the differencesin creatinine clearance calculated by the CG formula andstandard formula in both 2-h and 24-h collections. A pairedt-test was used to compare the results of protein excretionmeasured in 24 h and that estimated by P:C ratio. We thenused linear regression analysis and the correlation coeffi-cient (r2 value) to compare creatinine clearance and quan-titative proteinuria in the 2-h and 24-h groups. We per-formed this analysis looking at results by trimester and inthe total subject population.

Finally, we studied the data using a method of analysisdeveloped by Bland and Altman [14] specifically designedto mathematically assess the agreement between two meth-ods of clinical measurements. This method also confirms ifand how much two methods of measurements differ. In allmethods of analyses we determined a P , 0.05 as statisti-cally significant.

RESULTS

Of the total enrollment (n 5 59), we analyzed data on 51patients. We excluded eight patients from the study becauseour laboratory reported that the 2-h samples in these eightpatients were considered inadequate either due to low vol-ume or to a creatinine level less than 10 mg/kg. The mean

TABLE 1. Mean Values of Creatinine Clearance by Cockcroft-Gault Formula, 2-H Measurement and 24-H Measurement

N

Creatinine clearanceby CG formula 2-hsample (ml/min)1

Creatinine clearanceCG formula 24-hsample (ml/min)1

Creatinine clearancestandard formula 2-h

sample (ml/min)1

Creatinine clearancestandard formula

24-h sample(ml/min)1 P1 r2 2 P2

TrimesterFirst 7 104.2 6 41 97.3 6 36 106.4 6 50 116.4 6 47 NS 0.72 ,0.001Second 14 115.4 6 32 110.6 6 39 118.6 6 49 121.8 6 56 NS 0.84 ,0.001Third 30 109.1 6 52 104.1 6 48 112.7 6 56 118 6 46 NS 0.83 ,0.001All 51 119.0 6 60 101.8 6 39 111.0 6 52 122.5 6 50 NS 0.76 ,0.001

Values are means 6 standard error.N 5 number of subjects in each category; NS 5 not statistically significant.1ANOVA; P , 0.05 as statistical significance.2Linear regression analysis, squared correlation; P , 0.05 as statistical significance.

234 EVANS ET AL.

subject age was 26.3 years (range: 14–47) and the meangestational age was 33.2 weeks. There were seven patientsthat presented in the first trimester, 14 in the second, and30 in the third. The diagnosis most frequently listed waspregnancy-induced hypertension (PIH) (n 5 32; 62%),followed by diabetes mellitus (n 5 8; 16%), and chronichypertension (n 5 7; 16%).

Table 1 shows the mean creatinine clearance by CGformula and standard formula for both the 2-h and 24-hcollections. This data is expressed in ml/min for all subjectsand for subjects grouped by trimester. By using ANOVA wedemonstrated no statistically significant differences in cre-atinine clearance results by either method in any group.Regression analysis and squared correlation showed directcorrelation of creatinine clearance calculated by bothmethods in all groups.

Paired t-test analysis demonstrated no difference in meanprotein excretion—expressed in mg—estimated by calcu-lating P:C ratio in the 2-h group and measured proteinuriain the 24-h group. This is shown in Table 2. As seen withthe creatinine clearance data, linear regression analysisdemonstrated a correlation with protein excretion in eachtrimester.

We demonstrated that estimated protein excretion byP:C ratio correlated directly with measured 24-h urineprotein excretion. This is illustrated in Figure 1a with anr2 5 0.95, P , 0.0001. A Bland-Altman plot confirms thiscorrelation (Fig. 1b). P:C ratio calculated from the 2-hurine samples of #0.3 correlated directly with total pro-teinuria of #300 mg measured in 24-h samples with asensitivity of 90.9% and a specificity of 86.2%. The positiveand negative predictive values were 83.3% and 74%, re-spectively. The P:C ratio of .0.3 correlated with a proteinexcretion .300 mg/24 h. The sensitivity was 87.1%, thespecificity 100%, the positive predictive value 100%, andthe negative predictive value 76.7%. We also demonstratedthat the P:C ratio of $3.0 correlated with 24-h of protein-uria .3,000 mg with a sensitivity of 90%, a specificity of87.1%, a positive predictive value of 100%, and a negativepredictive value of 68%.

We analyzed the data using linear regression and Bland-Altman analyses from subjects with total 24-h proteinuriaof ,5,000 mg (Fig. 2a,b) and ,1,000 mg (Fig. 3a,b). Thesefigures clearly demonstrate the correlation of estimatedproteinuria by P:C ratio and measured 24-h protein excre-tion (Fig. 2a r2 5 0.79, P , 0.001; Fig. 3a r2 5 0.88, P ,0.0001).

DISCUSSIONOur study demonstrates that creatinine clearance and

protein excretion derived from a single-voided 2-h collec-

Fig. 1. Relationship between measured protein excretion in 24-h urineand estimated protein excretion by P:C ratio. There is a direct corre-lation between measured proteinuria in 24-h sampling and proteinuriaestimated by linear regression analysis (a: r2 5 0.95, P , 0.0001)and Bland-Altman analysis (b).

TABLE 2. Protein Excretion by Protein:Creatinine Ratio and 24-H Urine Measurements

N

Protein excretion byP:C ratio

(mg/24 h)1

Protein excretion by24-h measurement

(mg/24 h)1 P1 r2 2 P3

TrimesterFirst 7 746.4 6 546 890.6 6 812 NS 0.92 ,0.0001Second 14 1,327.7 6 1,100 1,542.5 6 1,322 NS 0.89 ,0.0001Third 30 4,863.6 6 972 6,105.4 6 1,180 NS 0.86 ,0.0001All 51 1,840.8 6 786 1,944.0 6 1,060 NS 0.95 ,0.0001

Values are means 6 standard error.N 5 number of subjects in each category; NS 5 not statistically significant.1Paired t-test; P , 0.05 as statistical significance.2Linear regression analysis, squared correlation; P , 0.05 as statistical significance.

TWO-HOUR URINE SAMPLE AND RENAL FUNCTION 235

tion provide the same clinical information as results from astandard 24-h urine collection in pregnant patients. Thisconfirms what has been found in three studies involvingnonpregnant critically ill patients [5–7]. We established adirect correlation with proteinuria estimated by P:C ratio inthe 2-h samples and measured in the 24-h samples. Ourresults are in agreement with findings of other reports thatdemonstrated the accuracy of a single voided urine samplefor the estimation of proteinuria in both pregnant andnonpregnant patients [1–4,8–10]. One study in the litera-ture reported a significant variation (up to 65%) in levels ofprotein excretion between single-voided and 24-h samplesinpatients with .1 g / 24 h proteinuria [9]. We did notdemonstrate such a variation. We observed that at levels,5,000 mg / 24 h the average variation was 10%. Above5,000 mg the average variation was less than 20%. Thehighest variation we found in comparing P:C ratio estima-tion of proteinuria to 24-h quantitative measurement ofprotein excretion was a single case of a patient who ex-creted 29,749 mg of protein in a 24-h collection. Theestimated proteinuria by P:C ratio from a single voided 2-hsample in this patient was 14,748 mg—a discrepancy of50%. We have no explanation for this discrepancy in thisparticular case. Examination of the case records revealed nosignificant clinical variation in this patient outside of the

marked proteinuria when compared to the other cases inour series.

Two-hour urine collection presents several advantagesover 24-h urine collection in assessing renal function. Cer-tain conditions in pregnancy such as preeclampsia requiremeasurement of renal function, namely protein excretion,as an aid to determine the severity of disease. Twenty-four-hour urine collections may cause a delay in the diagnosisand classification of the disease, which in turn may delayappropriate treatment and follow-up. Two-hour collectionmay afford more timely diagnosis, allowing appropriatemonitoring and treatment. Two-hour urine sampling mayalso be beneficial in monitoring the acute changes in renalfunction.

There is less risk of error in collection, handling, andstorage by patients, nurses, and laboratory personnel with2-h sampling. In both the outpatient and inpatient settings,it is difficult to monitor urine collection for 24 h unless thepatient has a continuous urinary bladder drainage system.With 24-h sampling, collection and handling errors in-crease, which may alter the results. Two-hour collectioncan be performed in an outpatient setting where collectionmay be monitored and handling supervised more closely.Also, the time between collection and testing is reduced,minimizing the need for prolonged storage.

Fig. 2. Relationship between protein excretion in subjects with lessthan 5,000 mg of proteinuria. There is a direct correlation betweenmeasured 24-h urine proteinuria and estimated proteinuria by P:Cratio by linear regression analysis (a: r2 5 0.79, P , 0.001) andBland-Altman analysis (b).

Fig. 3. Relationship between protein excretion in subjects with lessthan 1,000 mg of proteinuria. There is direct correlation betweenmeasured 24-h proteinuria and estimated proteinuria by P:C ratio bylinear regression analysis (a: r2 5 0.88, P , 0.0001) and Bland-Altman analysis (b).

236 EVANS ET AL.

Two-hour urine samples offer the same diagnostic infor-mation as the standard 24-h urine samples. Given theaccuracy of results and the potential decrease in errors ofcollection, it is clinically advantageous to consider use of2-h urine samples in determining renal function in theobstetric patient.

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