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Elevated spermidine level in pleural fluid of patients with lung cancer
Clinica Chimica Acta, 188 (1990) 183-184 Elsevier
183
CCA 04703
Letter to the Editor
Elevated spermidine level in pleural fluid of patients with lung cancer (Received
13 July 1989; revision received 23 December
1989; accepted
5 January
1990)
Dear Editor, The differentiation between malignant and benign pleural effusion is often difficult using biopsy, cytologic or bacteriologic examination of pleural fluid. However, increased polyamines in urine and blood of lung cancer patients have been reported [l-5]. Therefore, we developed an assay of spermidine, one of polyamines, in pleural fluid by high-performance liquid chromatography (HPLC) and applied it to pleural effusion. One milliliter of pleural fluid with 0.1 ml of 50 mmol/l 1,6-diaminohexane as an internal standard was deproteined by adding 1 ml of 100 g/l trichloroacetic acid and centrifuged for 5 min at 10000 X g. All supemate was shaked with 5 ml of diethyl ether for 5 min, and centrifuged for 1 min at 2000 X g. The upper layer was removed and the low layer was evaporated to dryness in vacua. The residue was dissolved with 0.5 ml of 6 mol/l hydrochloric acid, hydrolyzed for 16 h at 110 o C, and filtrated with a 0.45~pm membrane filter. The sample (50 ~1) was injected into the HPLC. Spermidine (retention time 10.4 min) was separated from the other amines on an ISC-OS/SO504 cation-exchange column (Shimadzu Co., Kyoto, Japan) at 70” C with isocratic elution and derived to a fluorescent substance with a post-column reaction and then monitored at 430 nm after excitation at 345 nm. The
Spermidine Primary Lung Cancer
(nmol/dl)
Of.ti:;.
M&as&tic Lung Cancer
Benign
I
Fig. 1. Distributions of spermidine in pleural fluid of patients with primary lung cancer (n =17), metastatic lung cancer (n = 7) and benign pleural effusion (n = 12). The spermidine level (mean k SD) in pleural fluid was 84k 39 nmol/dl for primary lung cancer, 94k32 nmol/dl for metastatic lung cancer and 49 f 26 nmol/dl for benign pleural effusion, respectively.
0009-8981/90/%03.50
0 1990 Elsevier Science Publishers
B.V. (Biomedical
Division)
184
mobile phase consisted of 0.17 mol/l trisodium citrate and 2.1 mol/l sodium chloride and flowed at 0.7 ml/mm. The post-column reagent solution consisted of 3.7 mmol/l o-phthalaldehyde, 12.8 mmol/l 2-mercaptoethanol, 5 g/l Brij 35 and 0.15 mol/l sodium borate, pH 10.5 and flowed at 1 ml/mm to a post-column reaction tube, 0.5 mm i.d. X 3 m. Figure 1 shows the distributions of pleural fluid spermidine in three patient groups. The mean spermidine levels in the primary and metastatic lung cancer groups were elevated significantly compared to the benign pleural effusion ( p < 0.01 by Student’s t test), but there was no significant difference between the means in primary and metastatic lung cancer (p > 0.05). However, only the spermidine concentrations in 4/17 patients with primary lung cancer and 3/7 patients with metastatic lung cancer were above the controls. These findings suggest that elevated spermidine may provide useful information about pleural effusions but a single assay hardly suffices to rule-out malignant disease. Kenji Abe I, Tatsuya Saito 2 Satoshi Kitamura 2 and Ryusei Konaka ’
2 Department
’ Shionogi Institute for Medical Science, Shionogi & Co., Ltd., Set&u, Osaka and of Pulmonary Medicine, Jichi Medical School, Minamikawachi, Tochigi, Japan
References 1 Nakamoto Y. Clinical and pathological study on urinary polyamine in lung cancer patients by FID gas chromatography. Lung Cancer 1982;22:397-402. 2 Woo KB, Waalkes TP, Abeloff MD, Lenhard Jr RE, Gehrke CW. Kuo KC. Urinary polyamines for evaluating the course of disease for patients with small cell carcinoma of the lung. Cancer 1983;52:1684-1690. 3 Moulinoux J-P, Quemener V, Larzul J-J, Le Calve M, Roth A-M, Toujas L, Quash G. Red blood cell polyamines in mice bearing the lewis lung carcinoma and in patients with bronchopulmonary cancers. Int J Cancer 1984;34:277-281. 4 Abe K, Hori T. Determination of polyamines in human urine by high-performance liquid chromatography with fluorescence detection. Rinsho Kagaku 1985;14:315-320. 5 Umeki S, Umeki Y, Okumoto T, Matsumori T. Urinary polyamines in malignancies. Med Lab Sci 1988;45:250-254.
183
CCA 04703
Letter to the Editor
Elevated spermidine level in pleural fluid of patients with lung cancer (Received
13 July 1989; revision received 23 December
1989; accepted
5 January
1990)
Dear Editor, The differentiation between malignant and benign pleural effusion is often difficult using biopsy, cytologic or bacteriologic examination of pleural fluid. However, increased polyamines in urine and blood of lung cancer patients have been reported [l-5]. Therefore, we developed an assay of spermidine, one of polyamines, in pleural fluid by high-performance liquid chromatography (HPLC) and applied it to pleural effusion. One milliliter of pleural fluid with 0.1 ml of 50 mmol/l 1,6-diaminohexane as an internal standard was deproteined by adding 1 ml of 100 g/l trichloroacetic acid and centrifuged for 5 min at 10000 X g. All supemate was shaked with 5 ml of diethyl ether for 5 min, and centrifuged for 1 min at 2000 X g. The upper layer was removed and the low layer was evaporated to dryness in vacua. The residue was dissolved with 0.5 ml of 6 mol/l hydrochloric acid, hydrolyzed for 16 h at 110 o C, and filtrated with a 0.45~pm membrane filter. The sample (50 ~1) was injected into the HPLC. Spermidine (retention time 10.4 min) was separated from the other amines on an ISC-OS/SO504 cation-exchange column (Shimadzu Co., Kyoto, Japan) at 70” C with isocratic elution and derived to a fluorescent substance with a post-column reaction and then monitored at 430 nm after excitation at 345 nm. The
Spermidine Primary Lung Cancer
(nmol/dl)
Of.ti:;.
M&as&tic Lung Cancer
Benign
I
Fig. 1. Distributions of spermidine in pleural fluid of patients with primary lung cancer (n =17), metastatic lung cancer (n = 7) and benign pleural effusion (n = 12). The spermidine level (mean k SD) in pleural fluid was 84k 39 nmol/dl for primary lung cancer, 94k32 nmol/dl for metastatic lung cancer and 49 f 26 nmol/dl for benign pleural effusion, respectively.
0009-8981/90/%03.50
0 1990 Elsevier Science Publishers
B.V. (Biomedical
Division)
184
mobile phase consisted of 0.17 mol/l trisodium citrate and 2.1 mol/l sodium chloride and flowed at 0.7 ml/mm. The post-column reagent solution consisted of 3.7 mmol/l o-phthalaldehyde, 12.8 mmol/l 2-mercaptoethanol, 5 g/l Brij 35 and 0.15 mol/l sodium borate, pH 10.5 and flowed at 1 ml/mm to a post-column reaction tube, 0.5 mm i.d. X 3 m. Figure 1 shows the distributions of pleural fluid spermidine in three patient groups. The mean spermidine levels in the primary and metastatic lung cancer groups were elevated significantly compared to the benign pleural effusion ( p < 0.01 by Student’s t test), but there was no significant difference between the means in primary and metastatic lung cancer (p > 0.05). However, only the spermidine concentrations in 4/17 patients with primary lung cancer and 3/7 patients with metastatic lung cancer were above the controls. These findings suggest that elevated spermidine may provide useful information about pleural effusions but a single assay hardly suffices to rule-out malignant disease. Kenji Abe I, Tatsuya Saito 2 Satoshi Kitamura 2 and Ryusei Konaka ’
2 Department
’ Shionogi Institute for Medical Science, Shionogi & Co., Ltd., Set&u, Osaka and of Pulmonary Medicine, Jichi Medical School, Minamikawachi, Tochigi, Japan
References 1 Nakamoto Y. Clinical and pathological study on urinary polyamine in lung cancer patients by FID gas chromatography. Lung Cancer 1982;22:397-402. 2 Woo KB, Waalkes TP, Abeloff MD, Lenhard Jr RE, Gehrke CW. Kuo KC. Urinary polyamines for evaluating the course of disease for patients with small cell carcinoma of the lung. Cancer 1983;52:1684-1690. 3 Moulinoux J-P, Quemener V, Larzul J-J, Le Calve M, Roth A-M, Toujas L, Quash G. Red blood cell polyamines in mice bearing the lewis lung carcinoma and in patients with bronchopulmonary cancers. Int J Cancer 1984;34:277-281. 4 Abe K, Hori T. Determination of polyamines in human urine by high-performance liquid chromatography with fluorescence detection. Rinsho Kagaku 1985;14:315-320. 5 Umeki S, Umeki Y, Okumoto T, Matsumori T. Urinary polyamines in malignancies. Med Lab Sci 1988;45:250-254.