Pyrogallol Red Method Research Articles

Difference in urinary protein-to-creatinine ratio using the benzethonium chloride and pyrogallol red methods in children.

The appropriate reference value of the urinary protein-to-creatinine ratio (PCR) for proteinuria may change when the urinary pyrogallol red (PR) protein assay method is changed to the benzethonium chloride method (BC). This study aimed to evaluate the difference between BC-based PCR (BC-PCR) and PR-based PCR (PR-PCR) values in children. We compared the BC-PCR and PR-PCR values in the same first-morning urine samples without significant proteinuria in school urine screening settings. The upper limit of the reference values was set at the 97.5th percentile. Notably, 133 samples from 124 individuals (female: 62%, age: median 12.3 years, range 6.3-16.9 years) were collected between August 2020 and October 2022. The diagnoses included 34 normal individuals and 99 with asymptomatic hematuria. The urinary protein (UP) concentrations measured using the BC (BC-UP) and PR (PR-UP) methods were in a linear relationship; however, the BC-UP concentrations were higher than the PR-UP concentrations (mean of differences: 11.2, 95% confidence interval (CI): 11.0-13.4 mg/dL). Also, the BC-PCR values were higher than the PR-PCR values (mean of differences: 0.090, 95% CI: 0.082-0.098 g/gCr). The BC-PCR showed a body-size-related decrease, reflecting a body-size-related urinary creatinine increase. The suggested BC-PCR reference values for proteinuria were 0.25 and 0.17 g/gCr for elementary (6-12.4 years) and junior high school students (12.5-16 years), respectively. These values were higher than those of the PR-PCR and need further studies. When evaluating PCR results, the urinary protein assay should be stated.

HOW TO INTERPRET URINARY PROTEINURIA AS A MARKER OF HYPERTENSIVE NEPHROPATHY?

Objective: Proteinuria may be the first sign of underlying nephropathy. It is pathological when the value is higher than 0.15 g/24h. The gold standard of quantification is the determination of proteinuria on the collection of urine for 24 hours, however, in view of the errors related to the collection and storage, we adopted another mode of expression. The objective of our work is to evaluate the correlation between the results of the 24 hours proteinuria and the proteinuria calculated by ratio (proteinuria/creatinuria). Design and method: Our prospective comparative study was performed on 100 24-hour urine samples (100 patients). We recorded the 24h diuresis and we determined the proteinuria by the pyrogallol red method (Advia 1800) and the urinary creatinine by the compensated jaffa method (Cobas 6000). Statistical evaluation was performed using GraphPad Prism software7 Results: Our results show: the average proteinuria of 24 h is 309.7 mg/24H (the norms lower than 150 mg/24H) and the average proteinuria/creatinine ratio is 283.7 mg/g (the norms lower than 300 mg/g). Of the 100 samples, only 5 patients (5%) showed pathological 24-hour proteinuria with a proteinuria/creatinuria ratio within the norms, and one patient (1%) had 24-hour proteinuria within the norms and a high proteinuria/creatinuria ratio, 25 patients (25%) had a simultaneous increase in 24-hour proteinuria and in the proteinuria/creatinuria ratio The remaining 69 patients (69%) had 24-hour proteinuria and a proteinuria/creatinuria ratio within the physiological norms The comparative study found a significant difference p<0.0001 with a significant positive Pearson correlation R 2 = 0.92. The agreement of the 24 h proteinuria tests and the Proteinuria/creatinuria ratio is modelled by the Bland and Altman diagram which finds a bias of 1.02 and a Confidence interval 95%] 0.046-1.36 [. Conclusions: Therefore it is not necessary to carry out a 24-hour urine collection, preferring measurements on urine samples from the first morning micturition. In order to overcome the numerous false positives and negatives linked to the collection of 24-hour urine, this examination is a good alternative.

FREQUENCY OF MICROALBUMINURIA IN HYPERTENSIVE PATIENTS WITH ACUTE ISCHEMIC STROKE

Background: Stroke, and its most common variant ischemic stroke, is one of the major causes of morbidity and mortality worldwide. Besides conventional risk factors, microalbuminuria (MA) is considered to be associated with atherosclerosis, hence ischemic stroke. Early detection of microalbuminuria may help in identifying hypertensive patients at high risk of ischemic stroke. The aim of current study was to determine the frequency of microalbuminuria in hypertensive patients with ischemic stroke. Methodology: This cross-sectional study was conducted at the Department of Medicine, Hayatabad Medical Complex Peshawar, Pakistan from July 2020 to June 2021. A total of 196 hypertensive patients of either sex with ischemic stroke were included in this study. They were tested for microalbuminuria in a 24-hours urine sample using Pyrogallol Red method. Urine albumin concentration of 30–300 mg/day was considered as microalbuminuria. The frequency of microalbuminuria was stratified according to age and gender using Microsoft Excel sheet. Results: The participants were aged 30–70 years and most of them (88, 45%) were in the age group 61–70 years. Majority (122, 62%) of the patients were male. Microalbuminuria was identified in 92 (47%) patients with slight male predominance (57, 62%). Conclusion: Given the high frequency of Microalbuminuria in hypertensive patients with acute ischemic stroke, microalbuminuria is a useful and inexpensive parameter in identifying hypertensive patients at increased risk of ischemic stroke. Pak J Physiol 2022;18(2):35?7

TOTAL PROTEIN TO CREATININE RATIO AS AN ALTERNATIVE TO ALBUMIN TO CREATININE RATIO TO PREDICT MICROALBUMINURIA IN TYPE 2 DIABETES MELLITUS

Background: Diabetic nephropathy is the most common complication of diabetes mellitus. Early detection of microalbumin in urine plays an im­portant role in preventing the progression to late stages of chronic kidney disease. For a country like Nepal, the cost of assessment of microalbumin in urine is unaffordable by many patients for regular monitoring. Total pro­tein to creatinine ratio (TPCR) might be a cheaper alternative for this. This study assessed the feasibility of using TPCR as an alternative method for predicting microalbuminuria in diabetic patients.
 Methods: Type 2 Diabetic patients with age ranging 30-80 years were in­cluded in the study after ethical clearance. Urinary total protein was deter­mined by pyrogallol red method and urinary creatinine by Jaffe’s method. Urine for microalbumin was determined by nephelometry. Relationships between variables were examined by Pearson correlation or Spearman’s correlation analysis as appropriate. Receiver operating characteristic (ROC) curve analysis was performed to obtain the cutoff value of TPCR for detection of albuminuria
 Results: The mean age of the study population was 55.9 ± 11.8 years. There was significant positive correlation between TPCR and urine albu­min (ρ = 0.56; p <0.01) and between TPCR and ACR (ρ = 0.47; p < 0.01). The regression equation for TPCR and ACR was, ACR = 0.82 TPCR = 70.5, r2 = 0.88. ROC curve analysis showed that the urine TPCR had a sensitivity and specificity of 76% and 58% respectively, for the detection of albuminuria with a cutoff value of 95 mg/g(AUC=0.74, p < 0.01).
 Conclusions: TPCR might be the cheaper alternative for the prediction of microalbuminuria in patients with type 2 diabetes mellitus.

Optimization of 25% Sulfosalicylic Acid Protein-to-Creatinine Ratio for Screening of Low-Grade Proteinuria.

Proteinuria is an important prognostic marker in the diagnosis and management of kidney diseases. Sulfosalicylic acid method (SSA) is a simple, low cost, qualitative test, widely used to assess proteinuria. The aim of this study was to optimize SSA test as a quantitative screening tool to assess proteinuria at lower excretory levels which would facilitate the screening and early diagnosis of renal impairment using protein-to-creatinine ratio (PCR). The study was conducted in two phases. In phase I, optimum SSA percentage to detect low-grade proteinuria was selected by comparing the performance of 3%, 6%, and 25% SSA methods in manual spectrophotometric analysis. In phase II, clinical applicability of the optimized method was evaluated using retained urine samples of patients with chronic kidney disease (CKD) assessed for urine protein by the pyrogallol red (PGR) method in a tertiary care hospital in Sri Lanka. Optimized 25% SSA protein-to-creatinine ratio (PCR) was compared with PGR PCR and albumin-to-creatinine ratio (ACR). Sensitivity, specificity, degree of agreement, correlation, and diagnostic accuracy were evaluated. Turbidimetric analysis using 25% SSA was linear in the range 3–50 mg/dL giving the highest analytical sensitivity. The test yielded a sensitivity of 86.5% and specificity of 96.5% and a degree of agreement of 5 mg/dL with the PGR method. Optimal cut-off for 25% SSA PCR in receiver operating characteristic analysis was 166 mg/g. Spearman's correlation coefficient for 25% SSA PCR versus ACR was r = 0.823, p < 0.0001, and for 25% SSA PCR versus PGR PCR was r = 0.913, p < 0.0001. The 25% SSA PCR has a sensitivity of 92% against ACR, the current prognostic marker for proteinuria in patients with CKD. The 25% SSA test is a simple method, and it performs satisfactorily as a screening test with a cut-off for PCR optimized at 166 mg/g. The test merits further evaluation due to its low cost.

Abstract P031: Reduced Augmentation Of The Intrarenal Renin-angiotensin System (ras), Lower Systolic Blood Pressure, And Preserved Renal Function In Female Compared To Male Rats With Unilateral Renal Artery Stenosis

Despite growing evidence of sex differences in the progression of hypertension, there are no guidelines that differentiate treatment between men and women. Intrarenal renin-angiotensin system (RAS) activation and tissue injury in 2-kidney, 1-clip (2K1C) hypertensive rats have been characterized in previous studies of male but not female rats. To evaluate possible sex differences in response to renovascular hypertension, urinary angiotensinogen (uAGT) excretion, systolic blood pressure (BP), urinary protein excretion, and renal function were assessed in female rats.Female (n=8) and male (n=6) rats underwent placement of a 0.2 mm clip on the left renal artery to simulate unilateral renal artery stenosis. BP was measured by tail-cuff plethysmography, and clearance studies were conducted in anesthetized rats to assess renal function. Urine protein concentration was determined by pyrogallol red method. uAGT was measured by ELISA as an index of intrarenal RAS activity. Systolic BP increased from 120±1 to 176±8 mmHg, and urinary protein excretion reached 20.2±5.6 mg/day in female rats. Although uAGT excretion increased from 13.2±7.7 ng/day to 74.1±29.9 ng/day in female rats, male rats had a significantly higher uAGT excretion of 1572.6±750 ng/day. Nonclipped kidneys exhibited more uAGT excretion compared to clipped kidneys, consistent with previous findings in males. Although 2K1C female rats demonstrate significantly lower renal function than sham females, they show more preserved renal function than male rats. Female rats also demonstrate significantly lower increases in systolic BP and urinary protein excretion compared to male rats. The data support substantial sex-dependent differences in renal responses to unilateral renal artery stenosis. The results show substantial increases in systolic BP, uAGT, and urinary protein excretion and decreased renal function after renal artery clipping in females, but the magnitude of the changes is markedly lower than in males. Nonclipped kidneys of both sexes exhibit greater uAGT excretion than clipped kidneys. Notably, females show less augmentation of the intrarenal RAS compared to male rats in renovascular hypertension.

Sex Differences in Urinary Angiotensinogen (uAGT) Excretion, Renal Function, and Systolic Blood Pressure in 2‐Kidney, 1‐Clip Hypertensive Rats

Intrarenal renin‐angiotensin system (RAS) activation and tissue injury in nonclipped kidneys of 2‐kidney, 1‐clip (2K1C) hypertensive rats have been characterized in previous studies of male but not female rats. To evaluate possible sex differences in response to renovascular hypertension, urinary angiotensinogen (uAGT) excretion, arterial blood pressure, urinary protein excretion, and renal function were assessed in 2K1C female rats.Female (n=8) and male (n=6) Sprague Dawley rats underwent placement of a 0.2 mm silver clip on the left renal artery to simulate unilateral renal artery stenosis. BP was measured by tail‐cuff plethysmography, and 24‐hour urine volume and water intake were monitored via metabolic cages over 21 days. After this period, renal clearance studies were conducted in anesthetized rats, and urine was collected from each kidney separately over 30 minute intervals. Urine protein concentration was determined by pyrogallol red method, and uAGT was measured by ELISA as an index of intrarenal RAS activity. Inulin and PAH concentrations were measured by spectrophotometry for calculation of glomerular filtration rate (GFR), renal plasma flow (RPF), and renal blood flow (RBF).21 days after renal artery clipping, systolic BP had risen to 176±8 mmHg from a baseline pressure of 120±1 mmHg, and urinary protein excretion increased to 20±5 mg/day in females. uAGT excretion was significantly increased from the baseline of 13 ng/day to 74 ng/day (507±127% increase). The nonclipped kidney showed 90±29% higher uAGT excretion compared to the clipped kidney, consistent with our previous findings in males. GFR and RPF were higher in nonclipped kidneys than in clipped kidneys but were greater than in corresponding measurements in males. GFR, RPF, and RBF were significantly higher in control rats compared to both clipped and nonclipped kidneys. Male rats showed significantly higher increases in uAGT, BP, and protein excretion.The present study demonstrates that BP, uAGT, and urinary protein excretion substantially increase after renal artery clipping in females, but the magnitude of the changes is markedly lower than in males. Additionally, the nonclipped kidney showed higher uAGT excretion compared to the clipped kidney. Although GFR, RPF, and RBF were significantly lower than control values for both clipped and nonclipped kidneys, these values were higher for females than males. These results demonstrated that female rats may be protected against AngII‐mediated renal injury and hypertension.Support or Funding InformationSupported by the ASPiRE Program at Tulane Univ Sch Medicine, New Orleans, LA and the Warren R. Bourgeois III, MD and Usha Ramadhyani Bourgeois, MD Student Research Endowed Fund at Tulane Univ Sch Medicine, New Orleans, LA

The diagnostic performance of human urinary dipsticks to estimate urine pH, specific gravity (SpG), and protein in horses: are they reliable?

BackgroundUrinalysis is a critical diagnostic test which is performed in routine veterinary medicine practice. In this diagnostic test, semiquantitative measurement of urine biochemical substances is carried out using urinary dipstick. In the current study, we evaluated the diagnostic performance of human urinary dipsticks to estimate pH, specific gravity (SpG), and protein in 80 urine specimens collected from horses. These parameters were measured using two commercial human dipsticks (KP and MN in abbreviation) and quantitative reference methods. The reference methods for pH, SpG, and protein were pH meter, handheld refractometer, and pyrogallol red method, respectively. The correlation between the semiquantitative dipstick analysis and quantitative reference methods was determined using Spearman’s rank correlation coefficient.ResultsIn general, our results revealed that the both human urinary dipsticks are unreliable tests for urinary pH, SpG, and protein content in horses. The analysis indicated that there was a poor correlation between the urine dipsticks and reference method (KP: rS = 0.534 and MN: rs = 0.485, Ps < 0.001) for protein. Additionally, there was a weak correlation between the results of pH measured using the urine dipsticks and reference method (KP: rS = 0.445 and MN: rs = 0.370, Ps < 0.001). Similar findings were obtained for SpG (KP: rS = 0.285, MN: rs = 0.338, Ps < 0.001). The estimation of proteinuria using the human dipsticks in horses lacked specificity, as many false positive protein results were obtained.ConclusionWe observed that the human commercial urinary dipsticks used in this study were not reliable to correctly estimate urine protein, SpG, and pH in horses.

Total Protein to Creatinine Ratio as an Alternative to Albumin to Creatinine Ratio To Predict Microalbuminuria In Type 2 Diabetes Mellitus

Background: Diabetic nephropathy is the most common complication of diabetes mellitus. Early detection of microalbumin in urine plays an im­portant role in preventing the progression to late stages of chronic kidney disease. For a country like Nepal, the cost of assessment of microalbumin in urine is unaffordable by many patients for regular monitoring. Total pro­tein to creatinine ratio (TPCR) might be a cheaper alternative for this. This study assessed the feasibility of using TPCR as an alternative method for predicting microalbuminuria in diabetic patients.&#x0D; Methods: Type 2 Diabetic patients with age ranging 30-80 years were in­cluded in the study after ethical clearance. Urinary total protein was deter­mined by pyrogallol red method and urinary creatinine by Jaffe’s method. Urine for microalbumin was determined by nephelometry. Relationships between variables were examined by Pearson correlation or Spearman’s correlation analysis as appropriate. Receiver operating characteristic (ROC) curve analysis was performed to obtain the cutoff value of TPCR for detection of albuminuria&#x0D; Results: The mean age of the study population was 55.9 ± 11.8 years. There was significant positive correlation between TPCR and urine albu­min (ρ = 0.56; p &lt;0.01) and between TPCR and ACR (ρ = 0.47; p &lt; 0.01). The regression equation for TPCR and ACR was, ACR = 0.82 TPCR = 70.5, r2 = 0.88. ROC curve analysis showed that the urine TPCR had a sensitivity and specificity of 76% and 58% respectively, for the detection of albuminuria with a cutoff value of 95 mg/g(AUC=0.74, p &lt; 0.01).&#x0D; Conclusions: TPCR might be the cheaper alternative for the prediction of microalbuminuria in patients with type 2 diabetes mellitus.

Longer Term Effect of Pregnancy Induced Hypertension on Chronic Kidney Disease and Hypertension

Background: Pregnancy-Induced-Hypertension (PIH) is claimed to create greater risk of hypertension (HTN) and chronic-kidney- diseases (CKD) at later life of the women. Objective: To explore what proportion of women with history of PIH develop HTN and CKD in later stages after delivery and, to study the interrelation of HTN and CKD with socio-demographic, anthropometric and biochemical risk factors. Methods: Under an observational case-control design 133 women with previous history of PIH [PIH group; Age(yrs), Median(range) 31(25–45), and BMI [(kg/m2, (Mean ± SD)(25.2 ± 2.1)] were compared with 113 women without history of PIH (Non-PIH group), Age(yrs) Median(range) 34(25–45), and BMI [(kg/m2, Mean ± SD) (25.8 ± 2.9)] for the development of HTN (SBP > 130 mmHg; DBP > 90 mmHg; or MBP > 105 mmHg) and CKD [as measured by total-urinary-protein and elevated urinary-protein-creatinine ratio (UPCR)]. Clinical and anthropometric parameters were measured by standard techniques, lipids by enzymatic-colorimetric method, urinary total protein by pyrogallol-red method, urinary-protein by strip method and urinary-creatinine was measured by alkaline-picrate method. Results: Out of the 133 women in PIH group 43(32.3%) developed HTN and 41 (30.8%) developed CKD in the Non-PIH group 17(15%) developed HTN and 16 (14.2%) developed CKD. SBP, DBP and MBP, all were significantly higher in the PIH compared to the Non-PIH group. PIH group had 4 times higher chance of developing HTN (Odds Ratio 4.2). In parallel to the findings on HTN, PIH group showed a significantly higher proportion of CKD [Proteinuric 10(7.5%)]. Both urinary-total-protein and UPCR were significantly higher in PIH group as compared to Non-PIH group. The PIH group had almost 3 times chance of developing CKD compared to the Non-PIH group. A significant correlation of MBP was found with UPCR, Total-cholesterol and serum-uric-acid. UPCR was also correlated with SBP. On logistic regression, MBP had a positive association with UPCR and Uric Acid. Conclusions: PIH-group is more vulnerable compare to Non-PIH group.

Use of red vacutainer for collection of CSF causes falsely high estimation of proteins by pyrogallol red method

BackgroundThough plain plastic/glass tubes are recommended for CSF collection, many laboratories use the commercially available red topped evacuated tubes for CSF collection for biochemical analysis. These red vacutainers affect the assay of some serum parameters. Aim: We evaluated the effect of using red vacutainer for CSF collection on estimation of proteins. MethodsCSF samples of 50 patients were collected in plain plastic containers. One milliliter from these were transferred to red vacutainers and mixed gently. Protein by pyrogallol red method was estimated directly from plastic containers and from the sample that was poured to red vacutainers. We further prepared different concentrations of bovine serum albumin in normal saline and estimated the O.D. on a spectrophotometer and protein levels on a clinical chemistry analyzer, similarly before and after transferring 1ml to red vacutainer. ResultsThe CSF protein levels were significantly higher (p=<0.0001) when transferred to a red vacutainer (median: 81.5 and range:32–324mg/dl) than that estimated directly from a plain plastic container (Median: 50.5 and range: 20–300mg/dl) and affected the interpretations in 50% cases. The protein levels were 25, 50, 50 and 355% higher when BSA prepared at concentrations of 100, 50, 25 and 10mg/dl respectively were transferred to red vacutainers. But for 750, 1500 and 3000mg/dl of BSA concentrations, the increase in red vacutainer was 1.4, 2 and 0.7% respectively. ConclusionCollection of CSF in red vacutainer significantly affects CSF protein estimations, probably due to the presence of clot activator. This might alter the interpretation of result and thus management of the subject. So the red vacutainer should not be recommended for CSF collection.

The Cutoff Level for Urine Protein in Urine Immunofixation Electrophoresis.

Immunofixation electrophoresis (IFE) maintains its importance in diagnosing monoclonal gammopathies. In particular, urine IFE detects free light chains (FLC) in urine samples even at low concentrations and offers higher sensitivity compared to serum electrophoresis and serum IFE. The aim of the present study was to determine the place and significance of quantitative urinary protein measurement before IFE in interpreting the results of subsequent IFE and to determine the most appropriate protein concentrations for the appearance of bands. The records of a total of 600 patients, who underwent screening for Bence Jones proteinuria using IFE on 24-hour urine, were retrospectively reviewed. Urine IFE was performed using Helena SAS-I and SAS-I devices. The total protein concentration in the urine was quantitatively determined by the Pyrogallol red method, and the urine albumin level was determined using the immunoturbidimetric method. These analyses were measured on an Olympus/Beckmann AU5800. The evaluation of IFE results revealed that 311 patients had normal results, 108 patients had monoclonal bands, five patients had biclonal bands, 28 had polyclonal bands, and 148 patients had various degrees of proteinuria. ROC curves were created in order to determine the most appropriate urinary protein and albumin levels to observe bands in IFE. Accordingly, urine baseline protein level (mg/dL) showed the highest AUC value (cutoff value: 19.4 mg/dL, sensitivity: 92%, specificity: 98.2%, AUC: 0.972). The present study showed that quantitative protein measurement before IFE eliminated the disadvantages associated with the IFE method and its interpretation.

Interference of cerebrospinal fluid total protein measurement by povidone–iodine contamination

BackgroundA falsely high cerebrospinal fluid (CSF) total protein (TP) result measured by pyrogallol red (PGR) method was suspected to be caused by preparation of the collection site with povidone–iodine (PVP–iodine) solution. MethodsCSF TP was evaluated for interference in samples with different final concentrations of PVP–iodine (up to 0.25% PVP and 0.025% iodine) or iodine alone (up to 0.025% iodine) using three methods: PGR, modified biuret and benzethonium chloride (BZTC). Interference exceeding ±20% of the baseline value is considered clinically significant according to the criterion defined by the College of American Pathologists. ResultsThere were positive interference with the PGR method and negative inference for the BZTC method in CSF samples spiked with PVP–iodine. The PVP–iodine (up to 0.25% PVP and 0.025% iodine) did not cause a clinically significant interference with the modified biuret method. PVP alone without iodine caused a positive interference with the PGR method but did not interfere with the modified biuret or the BZTC method. When the samples were spiked with iodine alone, none of the three methods was affected (change<20%) by iodine concentration up to 0.025%. ConclusionsContamination of CSF specimens with PVP–iodine can lead to interference with CSF TP measurements using PGR or BZTC methods.

English

There was a good correlation between Glomerular Filtration Rate (GFR) values estimated by any of the 3 methods. Crcl correlated better with Cockcroft-Gault (CG eGFR) than Modification of Diet in Renal Disease (MDRD eGFR) and there was also a very good correlation between CG eGFR and MDRD eGFR. Based on this result we feel that in advanced CKD all 3 methods give similar GFR estimates but in earlier stages of Chronic Kidney Disease (CKD) Creatinine clearance (Crcl) gives higher estimate of GFR. Thus use of Crcl may wrongly classify patients into less severe stages of CKD which might lead to delay in initiation of proper treatment. MATERIALS AND METHODS: CKD patients were selected on the basis of serum creatinine levels. S.C > 2.5 mg/dl values were considered to be having CKD. 75 Samples were tested. Both males & females patients are taken in this study. Serum Creatinine & Urinary Creatinine by Jaffe's method, Blood Urea by Urease method, Urinary Protein by Pyrogallol red method, estimation of GFR by three methods. Creatinine Clearance &eGFR by using Cockcroft- Gault& MDRD equations. RESULT: A total of 75 patients attending the nephrology department (P.E.S Medical College, Kuppam)for treatment and monitoring for chronic kidney disease where included in the present study. Serum creatinine, serum urea and creatinine clearance were measured in all these patients as per methods described under materials and methods. Estimated GFR (eGFR) using CG (Cock croft) and MDRD (Modification of Diet in renal disease) formulae were calculated. Majority of the cases were complications due to long standing diabetes mellitus or hypertension. CONCLUSION: Creatinine Clearance over estimates GFR in early stages of CKD. Use of Crcl to assess GFR may not be proper to classify CKD into different stages. Any of the 3 methods (Crcl, CG eGFR and MDRD eGFR) can be used as markers of GFR in advanced stages of CKD. MDRD eGFR is a better indicator of eGFR for staging CKD in early stages of CKD. All patients included in the present study had gross proteinuria even in early stages of CKD. There was no correlation between protein: creatinine ratio and CKD stages. We recommend a larger study reporting eGFR using MDRD equation along with serum creatinine for evaluation of renal function and confirm our findings. Estimated Glomerular Filtration Rate (eGFR), Glomerular Filtration Rate (GFR), Cockcroft-Gault (CG eGFR), Modification of Diet in Renal Disease (MDRD eGFR), Creatinine clearance (Crcl), Chronic Kidney Disease (CKD).

Cell Membrane Integrity of Candida Albicans after Different Protocols of Microwave Irradiation

Purpose: To evaluate the ability of low time microwaveexposureto inactivate and damage cell membrane integrity of C. albicans. Materials and Methods: Two 200ml C. albicans suspensions were obtained. Sterile dentures were placed in a beaker containing Experimental (ES) or Control suspensions (CS). ES was microwaved at 650 W for 1, 2, 3, 4 or 5 min. Suspensions were optically counted using Methylene blue dye as indicative of membrane-damaged cells; spread on Agar Sabouraud dextrose (ASD) for viability assay; or spectrophotometrically measured at 550nm. Cell-free solutions were submitted to content analyses of protein (Bradford and Pyrogallol red methods); Ca++ (Cresolphthalein Complexone method); DNA (spectrophotometer measurements at 260nm) and K+ (selective electrode technique). Data were analyzed by Student-t test and linear regression (α=0.05). In addition, flowcytometry analysis of Candida cells in suspensionwas performed using propidium iodide. Results: All ES cells demonstrated cell membrane damage at 3, 4 and 5 min,viable cells were nonexistent at 3, 4 and 5 min ES ASD plates and optical density of ES and CS was not significantly differentfor all exposition times. ES cells released highcontents of protein, K+, Ca++ and DNA after 2 min exposition when compared to that of the CSs. Similar results were observed with flow cytometry analysiswith regard to the periodsof microwave exposure. Conclusions: Microwave irradiation inactivated C. albicansafter 3min and damaged cell membrane integrity after 2 min exposition.

Serum magnesium levels in patients with diabetic retinopathy

Background:Diabetic retinopathy is one of the leading causes of blindness in the world. Hypomagnesemia has been reported to occur at an increased frequency among patients with type 2 diabetes compared with their counterparts without diabetes. Hypomagnesemia has been linked to poor glycemic control. Many studies have been undergone to find out the precipitated factors of retinopathy such as duration and type of diabetes, hyperglycemia, hypomagnesemia and increased urinary total protein levels.Aim:This study was carried out to study the correlation between serum magnesium levels, glycosylated hemoglobin and urinary total protein levels in diabetic patients with retinopathy.Materials and Methods:The study population comprised of 30 type 2 diabetic patients without retinopathy as Group 2, 30 type 2 diabetic patients with retinopathy as Group 3 in the age group 45-75 years as cases and 60 age and sex matched healthy individuals as controls (Group 1). Determination of Serum Magnesium (photometric xylidyl blue method), glycosylated hemoglobin, Hb1C (IFCC), fasting blood glucose, postprandial blood glucose (glucose oxidase method) and urine total protein (Pyrogallol red method) was carried out. The statistical software SPSS 11.0 and Systat 8.0 were used for the analysis of the data.Results:Hypomagnesemia was observed in cases compared with both Group 2 and Group 3. FBS, PPBS, HbA1c, Urine total protein levels were increased in cases (without retinopathy and with retinopathy) compared with controls.Conclusion:Hypomagnesemia and albuminuria individually or in conjunction serve as indicators for dysglycemia and could be used as marker for the risk of development of diabetic retinopathy.

Candida albicans inactivation and cell membrane integrity damage by microwave irradiation

In indicating the microwave irradiation for disinfecting dentures it is necessary to see how this procedure influences Candida albicans integrity and viability. The aim of this study was to evaluate the ability of microwaves to inactivate C. albicans and damage cell membrane integrity. Two 200-ml C. albicans (ATCC 10231) suspensions were obtained. A sterile denture was placed in a beaker containing the Experimental (ES) or the Control suspension (CS). ES was microwaved at 650 W for 6 min. Suspensions were optically counted using methylene blue dye uptake as indicative of membrane-damaged cells; spread on Agar Sabouraud dextrose (ASD) for viability assay; or spectrophotometrically measured at 550 nm. Cell-free solutions were submitted to content analyses of protein (Bradford and Pyrogallol red methods); Ca++ (Cresolftaleine complexone method); DNA (spectrophotometer measurements at 260 nm) and K+ (selective electrode technique). Data were analysed by Student's t- or Wilcoxon z-tests (alpha = 0.05). All ES cells demonstrated cell membrane damage. Viable cells were non-existent in the ES ASD plates. No significant difference in optical density between ES and CS was observed (P=0.272). ES cells released significantly high protein (P<0.001, Bradford; P=0.005, Pyrogallol red), K+ (P<0.001), Ca++ (P=0.012) and DNA (P=0.046) contents. Microwaves inactivated C. albicans and damaged cell membrane integrity.

A sensitive and economical modified method for estimation of cerebrospinal fluid proteins

Cerebrospinal fluid protein estimation is a routine diagnostic tool for various diseases. The proposed modified bromocresol green (BCG) method is applicable to automated as well as manual measurements. Protein concentrations and absorbance at 600 nm are linearly related throughout a wide range of concentrations from 10 mg % to 2000 mg %. Ninety C.S.F. samples were analyzed and results obtained by both the modified BCG method and routine pyrogallol red method were compared. An excellent co-relation was observed (n=90, g=0.996, P<0.001).

Urinary protein excretion profile: A contribution for subclinical renal damage identification among environmental heavy metals exposure in Southeast Brazil

In Southeast Brazil. Ribeira Valley region has been a major public health concern due to the environmental heavy metals contamination indexes of vegetation, rocks and aquifers. caused by local mining in the past. Human contamination by low levels of heavy metals doesn't cause acute intoxication but in chronic exposure. renal damage may occur with progressive tubulointerstitial changes evolving to glomerular lesion. In this study we investigated the relationship between the profile of urinary excreted proteins (glomerular or tubular origin) of arsenic and mercury and blood lead concentration in children and adults from highly exposed regions of the Ribeira Valley. The subjects were classified as GROUP 1 (G1; higher environmental risk, n=333) and GROUP 2 (G2; lower risk of contamination. n=104) In order to determine the urinary excretion of total protein, albumin (MA. glomerular marker) and alpha 1 microglobulin (A1M, tubular marker) and the blood lead concentrations, random urine and blood samples were obtained. Plasmatic lead levels were assessed by atomic absorption spectrometry with graphite furnace. Total protein concentration (PROT) was assessed on a biochemical analyzer (pyrogallol red method). MA and AIM were determined by nephelometric method. Group 1 showed a higher frequency of altered urinary excretion of PROT (G1=34%; G2=1.0%), MA (G1=9.0%; G2=51%) and AIM (G1=7.5%; G2=3.8%), without significant differences between both groups. Elevated arsenic levels were more prevalent among subjects from Group 1 (28.8%) and demonstrated a significant correlation with abnormal urinary excretion of albumin and alpha-1-micrglobulin (p -0.019). Lead and mercury levels showed no difference among the groups and no correlation with MA and/or Al M. Our data suggests that abnormal urinary protein excretion is relatively frequent in this population independently of the plasmatic or urinary heavy metal levels. The early detection of possible renal damage become necessary for effective measures can be taken to prevent clinical nephropothies.

French experience of quality assessment of quantitative urinary analysis

Since 1986, clinical biochemists from the Rhône alpes area, in collaboration with a non-profit-making association (Pro Bio Qual), have been conducting an inter-laboratory quality assurance program for quantitative urine analysis. We investigated the precision and accuracy of individual methods and measurement systems routinely used in the monthly control for 13 analytes: albumin, α-amylase, calcium, chloride, creatinine, glucose, magnesium, phosphorus, potassium, total protein, sodium, urea, and urate. The number of laboratories participating in the program increased from 60 in 1986 to 277 in 1999. In 1986, the greatest inter-laboratory imprecision occurred in the assay of urinary total protein, because the commonly used sulfosalicylic acid turbidimetric methods displayed poor precision. Since 1989, the increasing use of pyrogallol red or Coomasie Blue dye colorimetric methods has improved inter-laboratory precision markedly. The acceptable precision and good practicability have encouraged the general use of the pyrogallol red method. As for albumin analysis, which uses a specific immunologic method, the precision was reasonably good. Quality assessment of chloride and sodium decreased when the method performance improved for other analytes (calcium, creatinine, glucose, magnesium, phosphorus, potassium, urea, urate). This program has helped the laboratories to improve the quality of quantitative urine analysis, particularly for total protein.