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Measurement of strontium in serum, urine, bone, and soft tissues by Zeeman atomic absorption spectrometry.

Sr, a member of the earth alkaline elements, resembles to a great extent Ca. Like the latter element, >99% of the total Sr body burden is localized in bone. Whether the element plays an essential role in bone metabolism is not yet fully understood. In rats the oral administration of low doses of Sr may stimulate bone formation. When given at high doses, however, Sr may induce a defective bone mineralization expressed by a decreased bone mineral density and decreased size of bone apatite [1, 2]. In this context our recent findings of increased bone Sr concentrations in dialysis patients with osteomalacia [3] are of particular interest. At present it is not clear if in these individuals Sr has played either a primary or contributive role in the development of this particular type of renal osteodystrophy, nor is it clear if the increased concentrations occurred secondary to the presence of the bone disorder. Also, no unequivocal data are available in the current literature as to what extent chronic renal failure or the dialysis treatment per se leads to accumulation of the element. In the latter population, the element's ability to interfere with Ca homeostasis and vitamin D metabolism might be of particular interest, since in dialysis patients both of these processes are disturbed [4,5].

In 1986, Canavese et al. [6] stated that a better identification of the causal factors of the Sr imbalance in end-stage renal failure patients and information on the biological nature of Sr in biological fluids and tissues are necessary to better understand the effects of the element on its target tissue, more particularly the bone. Also, the element has recently been proposed as a potential therapeutic agent in the prevention and treatment of osteopenic bone lesions [7]. This implies that biological monitoring of Sr may become important in the near future.

The beneficial as well as deleterious effects of Sr on bone have been investigated in several experimental and some human studies [1, 2, 8, 9]. In the great majority of studies on the biological effects of Sr, however, data on Sr concentrations are lacking. Remarkably, this goes along with a scarcity of reports describing analytical methods that are sensitive and accurate enough to determine the element in biological fluids, bone, and tissues. Techniques applicable for analyzing Sr are flame atomic emission spectroscopy with a filament vaporizer [10], inductively coupled plasma atomic emission spectrometry [11], direct-current plasma echelle spectrometry [12], neutron activation analysis [13], and x-ray fluorescence [14]; these techniques are not currently available in the clinical laboratory. Here analysts mostly have to rely on atomic absorption spectrometry (AAS), either based on flame (FAAS) or electrothermal atomization (ETAAS). (2) FAAS in general is not sensitive enough to directly determine the element at the relatively low (ppb-ppm) concentration ranges at which it occurs in biological fluids and tissues. Moreover, in the analysis of biological material, FAAS-based methods are known to be subject to several chemical interferences. In view of its potential analytical performance, ETAAS is at present the method of choice to determine Sr in biological samples. However, even in the presence of the latter technique, several difficulties have been reported in the analysis of serum and urine [15]. Furthermore, data in the literature related to the ETAAS determination of Sr in more complex matrices such as bone are extremely scarce and to the best of our knowledge do not exist for the measurement of the element's low concentrations in soft tissues.

In view of the above and to allow us to study the accumulation/ distribution of Sr in dialysis patients and to investigate whether in these subjects the presence of particular types of renal osteodystrophy are accompanied by an increased/ decreased Sr concentration, Zeeman ETAAS-based analytical methods for the element's determination in serum and urine were further optimized and were developed for bone and soft tissue analysis.

Materials and Methods

APPARATUS

Analyses were performed with a Zeeman 3030 AAS equipped with an HGA-600 graphite furnace, an AS-60 autosampler, and an Anadex DP-9500B silent scribe printer, all from Perkin-Elmer, Norwalk, CT. The light source was a hollow cathode lamp, which was used at 25 mA at 460.7 nm. Instrument settings for the various biological matrices are presented in Table 1. Atomization was performed from the wall of pyrolytically coated graphite tubes.

MATERIALS

Materials used were either 5- or 10-mL polystyrene tubes, 50-mL polypropylene volumetric flasks (Brand, Wertheim, Germany), Teflon tubes (VEL, Leuven, Belgium) equipped with loosely fitting Teflon caps, and polystyrene sample cups (Biolab, Limal, Belgium). All automated pipettes used were "Finnpipettes" (Labsystems, Helsinki, Finland) equipped with disposable polypropylene pipette tips. The use of glassware was avoided since this material might be contaminated with Sr [16].

REAGENTS

Concentrated HN[O.sub.3] (Suprapur 441; Merck, Darmstadt, Germany) and concentrated Triton X-100 (BDH Chemicals, Poole, UK) were used to prepare the sample dilution reagents. Soft tissues were dissolved in a 100 g/L tetramethylammonium hydroxide (TMAH) (Janssen Chimica, Geel, Belgium) solution in water, whereas for bone digestion concentrated HN[O.sub.3] (Suprapur, Merck) was used. EDTA was from Merck (Art 8418, "Titriplex III"). A 1 g/L stock calibrator of SrC[O.sub.3] in 0.3 mol/L HN[O.sub.3] U.T. Baker, Philipsburg, NJ) was used to prepare intermediate and working calibrators.

SAMPLES AND SAMPLE TAKING

Serum, urine, bone, and tissue samples were taken and stored as described previously [17]. Optimization of the methodology for Sr in serum was done with samples that were taken from both dialysis patients and individuals with normal renal function. Bone samples were transiliac biopsies from dialysis patients that were taken in the frame of previous studies of our group on the diagnosis and prevalence of particular types of renal osteodystrophy [18,19]. For method development for Sr in urine, samples were taken from healthy humans, whereas for soft tissues rat specimens were used.

SAMPLE PREPARATION

Serum. The serum samples were diluted fourfold in a 0.5 mL/L Triton X-100-1 mL/L HN[O.sub.3] solution.

Urine. Urine samples were diluted 20-fold in a 2 mL/L HN[O.sub.3] solution.

Bone. Wet weighed bone biopsy samples (10-500 mg) were quantitatively transferred to 10-mL Teflon tubes to which either 1 mL or 2 mL (according to the sample weight) of concentrated HN[O.sub.3] was added. Tubes were closed by means of loosely fitting Teflon caps to allow digestion at atmospheric pressure. These were placed in an oven for 3 to 4 h at 80-90[degrees]C until a clear digest was obtained. The digestion liquid was then quantitatively transferred to either 25-mL or 50-mL polypropylene volumetric flasks and adjusted to the appropriate volume with deionized/reverse osmosis-treated water. This solution was then transferred to two 10-mL polystyrene tubes and stored at -20[degrees]C; the digestion liquids were diluted 40-fold in an aqueous 0.5 mL/L Triton X-100-0.5 mL/L HN[O.sub.3] solution before analysis.

Soft tissues. Wet weighed soft tissue samples (100-300 mg) were dissolved in 10-mL stoppered graduated polystyrene tubes after the addition of 1 to 3 mL (according to sample weight; [+ or -] 1 mL/100 mg sample) of a 100 g/L TMAH solution in water. Caps of tubes were perforated with an 18-gauge syringe needle to allow evaporation and make dissolution at atmospheric pressure possible. Solubilization was performed for at least 12 h (overnight) at 60[degrees]C. Samples were regularly mixed by means of a vortex-type mixer to accelerate the dissolution process. After completion of dissolution, samples were, according to the expected concentration or sample weight, adjusted to the appropriate volume (1:2 up to 1:5 dilution) with an aqueous 20 g/L TMAH-2 g/L EDTA solution in water.

CALIBRATION

From a 1 g/L stock calibrator solution of SrC[O.sub.3] in 0.3 mol/L HN[O.sub.3], an intermediate calibrator of 1 mg/L was prepared by diluting 50 [micro]L of the stock calibrator in 50 mL of a 20 mL/L HN[O.sub.3] solution. From this intermediate calibrator, working calibrators with appropriate concentrations were prepared in either aqueous, matrix-matched, or sample solutions for comparison of the direct calibration, matrix-matched calibration, and standard addition technique. Comparison of these yielded the following calibration procedures:

Serum. To 100 [micro]L of a serum pool with relatively low Sr content, 100 [micro]L of working calibrators of 0, 12.5, 25.0, 50.0, and 100.0 [micro]g/L in 0.5 mL/L Triton X-100-1 mL/L HN[O.sub.3] and 200 [micro]L of the blank 0.5 mL/L Triton X-100-1 mL/L HN[O.sub.3] solution were added, which yielded a matrix-matched calibration curve that contained Sr calibrators of 0, 3.13, 6.25, 12.5, and 25.0 [micro]g/L, respectively.

Urine. To 100 [micro]L of a urine pool with relatively low Sr content, 100 [micro]L of working calibrators of 0, 62.5, 125.0, and 250.0 [micro]g/L in 2 mL/L HN[O.sub.3] and 1800 [micro]L of the blank 2 mL/L HN[O.sub.3] solution were added. A matrix-matched calibration curve containing Sr calibrators of 0, 3.13, 6.25, and 12.5 [micro]g/L, respectively, was obtained.

Bone. For bone an aqueous calibration curve was constructed by preparing working Sr calibrators of 0, 3.13, 6.25, 12.5, and 25.0 [micro]g/L in 0.5 mL/L Triton X-100-0.5 mL/L HN[O.sub.3].

Soft tissues. Here, an aqueous calibration curve was prepared by diluting the intermediate calibrator in a 20 g/L TMAH-2 g/L EDTA solution so that working calibrators of 0, 3.13, 6.25, and 12.5 [micro]g/L were obtained.

Results and Discussion

CHOICE OF GRAPHITE TUBE

In the presence of uncoated graphite tubes, atomization from the wall yielded no signal (Fig. 1). Insertion of a L'vov platform did not result in any improvement, either in the presence of an uncoated tube or when a pyrolytically coated graphite tube was used or when the atomization temperature was increased to 2700[degrees]C. A possible explanation for this lies in the fact that optimum atomization temperatures with the L'vov platform are typically somewhat higher than those for direct atomization off the tube wall. In the determination of the relatively involatile Sr, the required temperature is probably not achieved in the presence of the platform.

Acceptable atomization signals were only obtained after atomization from the wall in the presence of pyrolytically coated graphite tubes. For Sr, atomization from the wall of pyrolytically coated graphite tubes has also been recommended by Slavin et al. [20], suggesting that the element, like the other earth alkaline metals, gets intercalated in the graphite of uncoated tubes, resulting in poor atomization signals. Others have used metal microtube atomizers made from a small inner-diameter tungsten tubing to improve the sensitivity and detection limits of the ETAAS determination and that of other relatively involatile elements [21].

In view of the nonideal peak shape with even pyrolytically coated graphite tubes, integrated absorbance ([A.sub.i]) measurements were preferred to peak height despite the lower sensitivity in the latter measurement mode. However, because of the better precision that was noted when the [A.sub.i] signals were used, measurement in the peak area mode did not result in poorer detection limits.

[FIGURE 1 OMITTED]

INSTRUMENT SETTINGS

A typical char-atomization curve in the presence of pyrolytically coated graphite tubes is presented in Fig. 2, showing Sr to be lost at temperatures >1500[degrees]C, whereas the plateau of maximal absorbance is reached at atomization temperatures >2200[degrees]C. The internal gas flow had an important influence on the sensitivity (Fig. 3). In contrast to others using nitrogen [15], we used argon as purge gas, which recently was shown to have a pronounced effect on both the absorbance signal (50% increase) and the lifetime of the graphite tube [16]. Balancing the loss in sensitivity against the expected concentration in the biological matrices under study, reproducibility, and tube lifetime, a miniflow of 50 mL/min instead of gas-stop was used during atomization for the determination of Sr in serum, bone, and urine. In view of the extremely low Sr concentrations noted in soft tissues for the analysis of these samples, the gas-flow during atomization was reduced to 10 mL/min to increase the sensitivity.

[FIGURE 2 OMITTED]

We used a slit width of 0.2 nm, which had a significant effect on the linearity of the calibration curve as compared with when a 0.7-nm slit width was used (Fig. 4). An overview of the instrument parameters is provided in Table 1.

SAMPLE PREPARATION

For the determination of Sr in serum and bone digestion liquids, the addition of Triton X-100 resulted in a better reproducibility without sensitivity loss. Perhaps this was due to a better sample deposition into the graphite furnace, which in turn was accompanied by a more regular drying of the sample. The addition of Triton X-100 was not necessary for the ETAAS analysis of urine samples. HN[O.sub.3] acid had a beneficial effect on the signal stability; however, it was not necessary in the analysis of soft tissue samples.

Sr in bone was determined after simple decomposition in HN[O.sub.3] under atmospheric pressure in loosely stoppered Teflon test tubes. Here, a clear digest is already obtained after a 3-4-h digestion at 90[degrees]C. After diluting the digestion liquid to the appropriate concentration range, Sr could directly be measured in the diluted digestion liquid. With the proposed method, the use of expensive digestion bombs or decomposition instrumentation is not necessary and a considerable number of samples can be analyzed in a day. In our laboratory the procedure has also been used successfully for the determination of a series of other trace metals [17, 22, 23]. Hence it allows multielement determination in one bone biopsy, which in view of the rather complex sampling procedures of these specimens must be considered an important advantage.

[FIGURE 3 OMITTED]

[FIGURE 4 OMITTED]

In view of the ultralow concentration at which Sr occurs in soft tissues as compared with bone, HN[O.sub.3] digestion could not be used in the analysis of the former specimen. Indeed, in the presence of such low concentrations, further dilution of the digestion liquid is limited, yielding an unacceptable high HN[O.sub.3] concentration in the final solution, which in turn would have a deleterious effect on the lifetime of the graphite tube and alter the absorption signal. To circumvent this problem we opted for a solubilization procedure in a 100 g/L TMAH solution wherein the element could either be measured directly or after a limited dilution in TMAH-EDTA. Since dissolution was performed in graduated polystyrene tubes, quantitative volume adjustment could be performed in the same recipient by which an additional transfer step to volumetric flasks was omitted. Here the addition of EDTA resulted in a significant ([+ or -] 30%) increase in sensitivity. Perhaps this is due to the ability of EDTA to form a chelate with Sr, which results in a more adequate atomization of the element by preventing it from forming either stable or volatile compounds with cations present in the sample matrix. The use of TMAH has previously been used successfully in our laboratory for the determination of Pb and Cd in soft tissue samples also (data not published). Stevens [24] used the latter reagent for the determination of Al in soft tissues. However, with TMAH, bone samples cannot be solubilized, making the latter method unsuitable for these specimens.

CALIBRATION

The linearity of the calibration curves was tested by calculating the correlation coefficient, which should be >0.99 in combination with the determination of the y-residuals (i.e., difference between the experimental y-values and the fitted y-values) representing the random experimental errors over the concentration range (see inset, Fig. 4). Calibration curves were linear up to a Sr concentration of 200.0 [micro]g/L, provided a slit width of 0.2 nm was used. Here a 200 [micro]g/L Sr concentration corresponded with 2.5 [A.sub.i] units. This corresponds with serum and urine Sr concentrations of 6.2-800 [micro]g/L and 31.2-4000 [micro]g/L, respectively. Calibration in the lower concentration range is depicted in the inset.

Calibration curves prepared for the biological matrices under study and comparison with aqueous calibrators are presented in Fig. 5. Confidence interval analysis on the difference between slopes of linear regression curves [25] indicated a significant difference between slopes of aqueous and urine-matched standard-addition curves. No significant differences were noted between aqueous calibration curves and standard-addition curves prepared for the other matrices under study. For each of the matrices under study, the slopes of the standard-addition curves prepared from samples of different subjects did not differ significantly from each other. Hence, the cumbersome standard-addition technique was not required for each of the matrices under study. Direct standardization with aqueous calibration curves could be used for the determination of Sr in bone and soft tissues. Because for serum a better analytical performance was obtained with matrix-matched calibration curves than with aqueous calibration curves, as for urine, we used the former calibration technique for serum also.

ANALYTICAL PERFORMANCE

Data on the analytical performance of the Sr determination in the various biological matrices as evaluated in terms of within- and between-run CVs, recovery of added analyte, recovery of digestion, sensitivity (determined by the characteristic mass, i.e., the amount of Sr yielding 0.0044 [A.sub.i] units), limit of detection (mean blank + 3 SD), and limit of quantification (mean blank + 10 SD) are presented in Tables 2 and 3.

For each of the matrices under study the within-run precision (repeatability) was assessed by determination of the Sr concentration in three samples containing Sr from the lower up to the higher concentration range (see Tables 2 and 3). Five dilutions were prepared per specimen, which were analyzed in two replicates within one run. For between-run precision (reproducibility), we determined weekly over a 21-day period in two replicates the Sr content in two dilutions of three samples of each of the biological matrices under study, covering a low to high concentration range.

[FIGURE 5 OMITTED]

At present, no Certified Reference Material (CRM), external quality-control schemes, or reference quality methods are available for the determination of Sr in serum or urine. Accuracy of the bone and tissue analyses was checked by means of the Community Bureau of Reference (BCR) CRM no. 278 (mussel tissue) (Table 4). To check the accuracy of the proposed method for tissue analysis in the ultralow concentration range, we used the Standard Reference Material (SRM) no. 1577 (bovine liver tissue material) (Table 5). Data indicate the proposed method to be accurate.

CLINICAL RELEVANCE OF DETERMINING SR IN BLOOD AND TISSUES

Mean Sr concentrations assessed in serum of 25 subjects with normal renal function were 30.9 [+ or -] 11.7 [micro]g/L. These correspond well with the lower concentration range of the data reported in the literature [26]. Preliminary data of our group obtained in an ongoing worldwide multicenter study indicate that, as previously noted for other elements, e.g., silicon [27], serum Sr concentrations in dialysis patients may differ from country to country and center to center, varying between values noted in individuals with normal renal function up to concentrations 10-20 times higher. This is in agreement with the scarce data in the literature on Sr concentrations in dialysis patients, which also show a wide variation in the serum Sr concentrations [6, 28,29]. Canavese et al. [6] reported values in patients on renal dialysis treatment of 56.1 [+ or -] 17.4 [micro]g/L, whereas Wilhelm et al. X28] found predialysis serum Sr concentrations of 185.3 [micro]g/L [+ or -] 34.8 [micro]g/L, which increased up to 330.0 [+ or -] 91.6 [micro]g/L at the end of dialysis. These latter figures indicate that the Sr content of the dialysis fluid plays an important role in the accumulation of the element in the dialysis population. As already established for other elements [27], further investigations are required to determine, besides contamination of the dialysis fluid, to what extent renal failure itself and (or) diet also determine serum Sr concentrations in dialysis patients.

Our recent observation of increased Sr concentrations in bone of dialysis patients with osteomalacia is intriguing [3]. At present it is not yet clear whether Sr has played a causative role in the development of this type of bone disease, or if accumulation of the element occurred secondary to the presence of osteomalacia. In this context it is worth mentioning that in preliminary studies in a chronic renal failure rat model, osteomalacia could be induced after Sr loading [30]. Others have also in both experimental and human studies demonstrated the element to interfere with bone metabolism [1, 2, 8]. On the other hand, Sr has a potential therapeutic value in the prevention and treatment of osteopenic disorders [7].

At present it is not yet clear to what extent serum Sr concentrations reflect the element's body burden, i.e., bone Sr concentrations or potential toxicity. In view of the above observations, validation of a serum Sr determination in the monitoring and diagnosis of Sr overload/ deficiency and treatment follow-up might become important.

The accurate determination of Sr gains in interest in view of the element's potential to mimic Ca absorption. Indeed, measurement of stable Sr instead of using the [sup.45]Ca isotope has recently been found to be a valuable safe alternative for Ca absorption tests [31, 32]. The latter issue might become of particular interest in dialysis patients, who are known to be prone to a disturbed Ca absorption mechanism.

The determination of Sr in urine will be a useful tool in both experimental and human studies to (a) investigate the interactions of the element with, e.g., Ca; (b) get a better insight into the renal clearance; and (c) define a possible role for impaired renal function in the accumulation of the element in chronic renal failure patients.

The methodology for the determination of Sr in tissues will be helpful to get a better insight into the element's metabolism and tissue/cellular uptake and distribution. Also, the determination of Sr in organs and tissues might become of interest in view of the potential interaction of the element with, e.g., Ca, which in turn might influence parathyroid gland function and vitamin D synthesis, two biochemical processes that are altered in renal failure [4,5].

The proposed methods for Sr determination in serum, urine, bone, and tissues are simple, sensitive, accurate, and precise. They allow routine determination as well as research investigations on the element at the lowest concentrations encountered in biological materials without the need for matrix modification. The STPF concept recommended for several trace metal analyses cannot be used for Sr because the element requires atomization from the wall of pyrolytically coated graphite tubes. We found that the use of TMAH offers a valuable alternative for the acid digestion of soft tissue samples, particularly when sample dilution of the digestion liquid is not possible because of the very low concentration at which the analyte occurs in the solid matrix under study. The described methods are useful for routine applications and are currently used successfully in both experimental and epidemiological studies on the potential role of Sr in the development of osteomalacia in chronic renal failure patients.

We are indebted to E. Snelders for expert desk editing and to D. De Weerdt for his excellent drawings. Thanks are also due to M. Elseviers for her advice on statistics.

Received January 2, 1996; revised July 22, 1996; accepted July 23, 1996.

References

[1.] Marie PJ, Garha MT, Hott M, Miravet L. Effect of low doses of stable strontium on bone metabolism in rats. Miner Electrolyte Metab 1985;11:5-13.

[2.] Grynpas M, Marie JP. Effects of low doses of strontium on bone quality and quantity in rats. Bone 1990;11:313-9.

[3.] D'Haese PC, Couttenye M-M, Lamberts LV, Goodman WG, De Broe ME. Increased strontium levels in bone of dialysis patients with osteomalacia [Abstract]. J Am Soc Nephrol 1995;6:935.

[4.] Ross EA, Nissenson AR. Acid-base and electrolyte disturbances. In: Daugirdas JT, Todd SI, eds. Handbook of dialysis, 2nd ed. Boston: Little Brown and Co., 1994:401-15.

[5.] Kokot F, Wiecek A. Endocrine changes in chronic dialysis patients. In: Maher F, ed. Replacement of renal function by dialysis. Boston: Kluwer Academic Publishers, 1989:953-71.

[6.] Canavese C, Pacitti A, Salomone M, Santoro MA, Stratta P, Mangiarotti G, et al. Strontium overload in uremic patients on regular dialytic treatment. Trans Am Soc Artif Intern Organs 1986;32:120-2.

[7.] Marie PJ. Effects of strontium on bone and bone cells. J Trace Elements Med Biol 1995;9:227.

[8.] Omdahl JL, DeLuca HF. Strontium induced rickets: metabolic basis. Science 1971;174:949-51.

[9.] Matsumuto A. Effect of strontium on the epiphyseal cartilage of rat tibiae- histological and radiographic studies. Jpn J Pharmacol 1976;26:675-81.

[10.] Grime JK, Vickers TJ. Determination of lithium in microsamples of blood using flame atomic emission spectrometry with a filament vaporizer. Anal Chem 1975;47:432-3.

[11.] Piette M, Desmet B, Dams R. Determination of strontium in whole blood by ICP-AES. Sci Total Environ 1994;141:269-73.

[12.] Berndt H, MesserschmidtJ. Simultaneous determination of Na, K, Ca, Li, Fe, Cu, and Zn in human serum using direct current plasma. Fresenius Z Anal Chem 1981;301:104-5.

[13.] Probst T, Zeh P, Kim J. Multielement determinations in ground water ultrafiltrates using inductively coupled plasma mass spectrometry and monostandard neutron activation analysis. Fresenius J Anal Chem 1995;351:745-51.

[14.] Valkovic V. Analysis of biological material for trace elements using x-ray spectroscopy. Boca Raton, FL: CRC Press, 1980:242-5.

[15.] Leeuwenkamp OR, van der Vijgh WJF, Husken BCP, Lips P, Netelenbos JC. Identification of strontium in plasma and urine with flameless atomic absorption spectrometry. Clin Chem 1989; 35:1911-4.

[16.] Barto R, Sips ADAM, van der Vijgh WJF, Netelenbos JC. Sensitive method for analysis of strontium in human and animal plasma by graphite furnace atomic absorption spectrophotometry. Clin Chem 1995;41:1159-63.

[17.] D'Haese PC, Van de Vyver FL, de Wolff FA, De Broe ME. Measurement of aluminum in serum, blood, urine, and tissues of chronic hemodialyzed patients by use of electrothermal atomic absorption spectrometry. Clin Chem 1985;31:24-9.

[18.] D'Haese PC, Couttenye MM, Goodman WG, Lemoniatou E, Digenis P, Sotornik I, et al. Use of the low-dose desferrioxamine test to diagnose and differentiate between patients with aluminum-related bone disease, increased risk for toxicity, or aluminum overload. Nephrol Dial Transplant 1995;10:1874-84.

[19.] Couttenye MM, D'Haese PC, Van Hoof V0, Lemoniatou E, Goodman WG, Verpooten GA, De Broe ME. Low levels of serum alkaline phosphatase of bone origin: a good marker of adynamic bone disease in hemodialysis patients. Nephrol Dial Transplant 1996; 11:1065-72.

[20.] Slavin W, Carnrick GR, Manning DC, Pruszkowska E. Recent experiences with the stabilized temperature platform furnace and Zeeman background correction. At Spectros 1983;4:69-86.

[21.] Ohta K. Electrothermal atomic absorption spectrometry with improved tungsten tube atomizer. Anal Chem 1987;59:539-40.

[22.] Liang L, D'Haese PC, Lamberts LV, De Broe ME. Direct calibration for determining aluminum in bone and soft tissues by graphite furnace atomic absorption spectrometry. Clin Chem 1991;37:461-6.

[23.] Liang L. Fast furnace determinations of aluminum and iron in bone and soft tissues. Spetrochim Acta 1992;47B:239-44.

[24.] Stevens BJ. Electrothermal atomic absorption determination of aluminum in tissues dissolved in tetramethyl ammonium hydroxide. Clin Chem 1984;30:745-7.

[25.] Altman DG, Gardner MJ. Calculating confidence intervals for regression and correlation. In: Gardner MJ, Altman DG, eds. Statistics with confidence-confidence intervals and statistical guidelines. London: British Medical Journal, 1989:34-49.

[26.] Sips ADAM, van der Vijgh WJF. Strontium. In: Seiler HG, Sigel A, Sigel H, eds. Handbook on metals in clinical and analytical chemistry. New York: Marcel Dekker, 1994:577-85.

[27.] D'Haese PC, Shaheen FA, Huraib SO, Djukanovic L, Polenakovic MH, Spasovski G, et al. Increased silicon levels in dialysis patients due to high silicon content of the drinking water, inadequate water treatment procedures and concentrate contamination: a multicentre study. Nephrol Dial Transplant 1995;10:1838-44.

[28.] Wilhelm M, Hanewinckel B, Blaker F. Influence of hemodialysis and renal transplantation on trace element concentrations in children with chronic renal failure. EurJ Pediatr 1986;145:372-6.

[29.] Mauras Y, Ang KS, Simon P, Tessier B, Cartier F, Allain P. Increase in blood plasma level of boron and strontium in hemodialysed patients. Clin Chim Acta 1986;156:315-20.

[30.] Cabrera W, D'Haese PC, Nouwen EJ, Schrooten I, Goodman WG, De Broe ME. Strontium-induced osteomalacia. A new disease entity in the dialysis population? Experimental evidence for a causative role [Abstract]. J Am Soc Nephrol 1996;7:1788.

[31.] Sips ADAM, Netelenbos JC, Barto R, Lips P, van der Vijgh WJF. One-hour test for estimating intestinal absorption of calcium by using stable strontium as a marker. Clin Chem 1994;40:257-9.

[32.] Blumsohn A, Morris B, Eastell R. Stable strontium absorption as a measure of intestinal calcium absorption: comparison with the double-radiotracer calcium absorption test. Clin Sci 1994;87: 363-8.

(2) Nonstandard abbreviations: FAAS, flame atomic absorption spectrometry; ETAAS, electrothermal atomic absorption spectrometry; TMAH, tetramethylammonium hydroxide; [A.sub.i], integrated absorbance; BCR, Community Bureau of Reference; SRM, Standard Reference Material; and CRM, Certified Reference Material.

PATRICK C. DHAESE, GLEN F. VAN LANDEGHEM, LUDWIG V. LAMBERTS, VERA A. BEKAERT, (1) IRIS SCHROOTEN, and MARC E. DE BROE *

Department of Nephrology-Hypertension, University of Antwerp, Antwerp, Belgium.

(1) Department of Industrial Sciences and Technology, Catholic Polytechnic Institute Antwerp, Antwerp, Belgium.

* Address correspondence to this author at: University of Antwerp, Department of Nephrology-Hypertension, p/a University Hospital Antwerp, Wilrijkstr. 10, B-2650 Edegem/Antwerpen, Belgium. Fax +32/3/829-0100; e-mail debroe@uia.ua.ac.be.
Table 1. ETAAS determination of Sr in serum, urine, bone,
and tissues.

 Time-temperature settings

 Temp, Ramp, s Hold, s Gas, mL/min
 [degrees]C

Step
 Dry 1 100 20 20 (40) (a) 300
 Dry 2 110 20 5 300
 Ash 1500 (1200) (b) 10(1) (a) 20 (10) (a) 300
 Atomize 2600 0 3 (5) (a) 50(10) (a)
 Clean 2750 1 5 300
Conditions
 Purge gas Argon
 Mode Peak area
 Injection 10 [micro]L (20 [micro]L) (a)
 volume
 Wavelength 460.7 nm
 Slit width 0.2 nm
 Furnace Pyrolytically coated
 graphite tube

(a) Soft tissues.

(b) Bone.

Table 2. ETAAS determination of Sr in serum and Urine
analytical performance.

 Serum Urine

Within-run CV, % 4.5 [+ or -] 2.1 3.3 [+ or -] 1.8
 (concentration range, (10-55) (27-178)
 [micro]g/L)
Between-run CV, % 3.2 [+ or -] 1.7 4.5 [+ or -] 1.1
 (concentration range, (10-55) (27-178)
 [micro]g/L)
Recovery added analyte, (a) % 101 [+ or -] 5.3 102 [+ or -] 3.5
Sensitivity, (b) pg 2.4 2.4
Limit of quantification, (c) 9.0 1.3
 [micro]g/L
Limit of detection, (d) 1.2 0.3
 [micro]g/L

(a) Aqueous calibration curve.

(b) Slope of calibration curve, i.e., amount
of Sr producing a 0.0044 [A.sub.j] change in signal.

(c) Mean blank + 10 SD.

(d) Mean blank + 3 SD.

Table 3. ETAAS determination of Sr in bone and soft
tissues--analytical performance.

 Bone

Within-run CV, % 5.7 [+ or -] 2.1
 (concentration range, [micro]g/L (a)) (4.7 [+ or -] 43.3)
Between-run CV, % 6.0 [+ or -] 3.0
(concentration range, [micro]g/L (a)) (4.6 [+ or -] 47.2)
Recovery digestion, (b) % 103 [+ or -] 2.1
Recovery added analyte, (b) % 99 [+ or -] 3.1
Sensitivity, (c) pg 3.9
Limit of quantification (d) 0.9 [micro]g/g
Limit of detection (e) 0.4 [micro]g/g

 Tissues

Within-run CV, % 4.3 [+ or -] 1.4
 (concentration range, [micro]g/L (a)) (5.6-12.0)
Between-run CV, % 9.0 [+ or -] 1.8
(concentration range, [micro]g/L (a)) (5.6-10.5)
Recovery digestion, (b) % 99 [+ or -] 4.2
Recovery added analyte, (b) % 100 [+ or -] 2.4
Sensitivity, (c) pg 2.6
Limit of quantification (d) 8.3 ng/g
Limit of detection (e) 2.2 ng/g

(a) Concentration of Sr in digestion liquid.

(b) Aqueous calibration curve.

(c) Slope of calibration curve, i.e., amount of
Sr producing a 0.0044 [A.sub.j] change in signal.

(d) Mean blank + 10 SD; calculated for a 500-mg bone
or tissue sample.

(e) Mean blank + 3 SD; calculated for a 500-mg bone
or tissue sample.

Table 4. Accuracy of methods for determination of
Sr in bone and soft tissues.

 Standard reference material: BCR
 CRM no. 278 (mussel tissue)

Technique Procedure Found, [micro]g/g Number of
 replicates

ICPES Pressurized digestion 13.8 [+ or -] 0.4 (a) 6
 with [HN0.sub.3]
EDXRF Pelletizing 14.8 [+ or -] 0.2 (a) 10
Zeeman-AAS Solubilization in 16.0 [+ or -] 0.7 (b) 6
 TMAH (60 [degrees]C)
Zeeman-AAS [HNO.sub.3] digestion 16.8 [+ or -] 0.5 (b) 6
 at atmospheric
 pressure
 (90 [degrees]C)

(a) Data provided by supplier of Reference Material.

(b) Determined in our laboratory.

ICPES, inductively coupled plasma emission spectroscopy;
EDXRF, energy dispersive x-ray fluorescence.

Table 5. Accuracy of method for determination of
ultralow concentrations of Sr in soft tissues.

 Standard reference material: SRM no. 1577
 (bovine liver tissue)

Technique Procedure Found, Number of
 [micro]/g replicates

Isotope dilution 0.14 (a)
mass spectrometry (indicative
 value)

Zeeman-AAS Solubilization in 0.11 [+ or -] 10
 TMAH (60 [degrees]C) 0.006 (b)

(a) Data provided by supplier of Reference Material. Indicative
value means that the value has not yet been certified, i.e., the
reported concentrations are not based on results of 6 to 12
determinations obtained by Reference Methods of known accuracy
and performed by two or more analysts or alternatively from
results obtained by two or more independent reliable analytical
methods (National Bureau of Standards information).

(b) Determined in our laboratory.
COPYRIGHT 1997 American Association for Clinical Chemistry, Inc.
No portion of this article can be reproduced without the express written permission from the copyright holder.
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Article Details
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Title Annotation:Automation and Analytical Techniques
Author:D'Haese, Patrick C.; Van Landeghem, Glen F.; Lamberts, Ludwig V.; Bekaert, Vera A.; Schrooten, Iris;
Publication:Clinical Chemistry
Article Type:Report
Date:Jan 1, 1997
Words:5680
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