Analysis of Urinary Calculi of Mixed and Unusual Composition: Raman Spectroscopic Investigation VENKATA RAMANA KODATI, ANTHONY T. TU,* RAVINDRA NATH, and JACOB L. TURUMIN Department o[ Biochemistry, Colorado State University, Fort Collins, Colorado 80523, U.S.A. (V.R.K., A.T.T.); Department of Biochemistry, Postgraduate Institute o[ Medical Education and Research, Chandigarh, India (R.N.); and Institute o[ Surgery, East-Siberian Department, Siberian Division of Medical Science, Irkutsk, Russia (J.L.T.) Raman spectroscopic analysis of kidney stones of mixed composition and of unusual stones is presented. Raman spectroscopy was used to analyze the chemical composition of kidney stones without extraction. A kidney stone sample was irradiated by laser light of 514.5 nm, and the scattered light was analyzed by a Raman spectrometer. The first stone was determined to be a mixture of calcium oxalate dihydrate and hydroxyapatite and the second one to be a mixture of calcium oxalate monohydrate, uric acid, and hydroxyapatite. The third and fourth stones were not mixed-composition stones, but rather stones with unusual com- position. One was concluded to be carboxyapatite, and the other was mucoprotein-type stone. The present investigation indicates that Raman spectroscopy is a useful tool for direct analysis of kidney stone without going through the usual procedure of crushing, extracting, and carrying out tedious wet chemical analysis. Index Headings: Kidney stones; Urinary calculi; Raman spectroscopy. INTRODUCTION The chemical composition of urinary calculi is impor- tant in understanding their pathogenesis, which in turn provides the direction for preventive medical treatment. There are several analytical techniques available for the analysis of kidney stones, such as infrared, ~Raman spec- troscopy, 2 x-ray diffraction,3 scanning electron micros- copy,4 polarization microscopy,5 and wet chemical pro- cedures, s Among these methods, Raman spectroscopy is a simple, nondestructive analytical technique that can identify the composition of urinary calculi by analyzing scattered light after irradiation of a stone with laser light. Some urinary calculi are composed of only one type of compound and some are of mixed composition. Pure stone formation generally requires specific conditions. Accord- ing to Hada et al., uric acid stones form in neutral or slightly basic solutions, while calcium oxalate and cal- cium phosphate stones require a more acidic condition.7 Struvite stones are invariably the result of renal infec- tion, while ammonium urate stones reflect undernutri- tion. We have systematically investigated different types of single-component stones by Raman spectroscopy. Our investigation indicated that one can readily identify stones of cystine, uric acid, calcium oxalate, and calcium phosphate types, s-H After analysis of all types of single- component kidney stones, stones of mixed composition were investigated by Raman spectroscopy. Received 26 October 1992. * Author to whomcorrespondenceshouldbe sent. EXPERIMENTAL Uric acid and hydroxyapatite were purchased from Sigma Chemical Company. Calcium oxalate monohy- drate was from Aldrich Chemical Company. Calcium ox- alate dihydrate was synthesized as reported. 1° Raman spectra of the standard compounds in powder form were recorded with a Spex Ramalog 5 Raman spec- trometer equipped with a Spex SCAMP data acquisition processor. The 514.5-nm excitation line from an argon- ion laser (Spectra-Physics, Model 164) at 200 mW power with a green interference filter was used to excite the samples. The scattered radiation was collected at 90° to the incident beam. A spectral bandwidth of approxi- mately 6 cm -1 and a scan rate of 1 cm-~/s were adopted. The signal was detected by an RCA C31034 photomul- tiplier tube and traced on paper by a Hewlett-Packard 7041A X-Y recorder. Each spectrum reported represents an average of 5 to 10 scans. The kidney stones were provided by Jacob L. Turumin, Irkutsk, Russia, and Ravindra Nath, Chandigarh, India. The small kidney stones were mounted intact, while the larger stones were cut in half with a jeweler's saw, and the cut surface was exposed to the laser beam at different locations. The kidney stones tended to give high fluo- rescence background; however, this problem was mini- mized by irradiating the samples with laser light at high power for several hours before recording the spectra. The spectra were smoothed and baseline corrected to resolve Raman bands from the high fluorescence background. RESULTS Altogether, four samples were analyzed by laser Ra- man spectroscopy, and these samples are shown in Fig. 1. The sites of laser irradiation on the stones are indicated by arrows. The spectrum of sample I together with standard com- pounds of calcium oxalate and hydroxyapatite are shown in Fig. 2. Sample 1, obtained from J. L. Turumin, gave three distinct Raman bands at 907, 962, and 1476 cm -1. The band at 962 cm -1 exactly coincided with the 962- cm -1 line of hydroxyapatite, and the 907- and 1476-cm -~ lines are very similar to the 912- and 1477-cm -1 lines of calcium oxalate. Therefore, it is reasonable to conclude that sample 1 has a mixed composition of calcium oxalate and hydroxyapatite. Sample 2 is a kidney stone obtained from northern India, and its composition had already been determined by chemical analysis. It was identified as a mixed composition of calcium oxalate, hydroxyapa- 334 Volume 47, Number 3, 1993 ooo3-7028/93/47o3-o33452.0o/0 APPLIED SPECTROSCOPY © 1993 Society for Applied Spectroscopy