Stanislav Pavel

Identification of three indolic compounds in a pigmented-melanoma cell-culture supernatant by gas chromatography-mass spectrometry

SummaryIn the supernatant of melanoma cell culture SK mel 25 three indolic compounds-5-hydroxy-6-methoxyindole, 5-hydroxy-6-methoxyindolyl-2-carboxylic, and 6-hydroxy-5-methoxyindolyl-2-carboxylic acids-have been identified. The supernatants were extracted with ethyl acetate, derivatized, and analyzed by gas chromatography-mass spectrometry. The significance of this finding is discussed.

Identification of two Thormählen-positive compounds from melanotic urine by gas chromatography—mass spectrometry

The isolation of two Thormählen-positive compounds from the urine of a patient with malignant melanoma and the elucidation of their structure by gas chromatography-mass spectometry is described. The compounds were isolated using a poly-N-vinylpyrrolidone column and separated by preparative thin-layer chromatography. After elution they were analyzed by gas chromatography and gas chromatography-mass spectrometry as their trimethylsilyl derivatives and after hydrolysis also as their tert.-butyldimethylsilyl derivatives. The results showed the main Thormählen-positive compound A to be the glucuronide of 5-hydroxy-6-methoxyindole, whereas the minor compound AX appeared to be the glucuronide of its isomer 6-hydroxy-5-methoxyindole.

A QUALITATIVE GAS-CHROMATOGRAPHIC ANALYSIS OF SUBSTITUTED 5,6-DIHYDROXYINDOLES FROM THE URINE OF PATIENTS WITH MELANOMA

A qualitative gas chromatographic analysis of trimethylsilylated ethyl acetate extracts of melanotic urine revealed 5 indolic compounds, which have been identified as substituted 5,6-dihydroxyindoles. Ethyl acetate extracts of melanotic urines at pH 2.0 contained isomeric 5-hydroxy-6-methoxy and 6-hydroxy-5-methoxy-indolyl-2-carboxylic acids which were not separable under the conditions used. A careful hydrolysis of melanotic urine with a Helix pomatia preparation followed by extraction at pH 6.5 in a nitrogen atmosphere released 3 additional indolic compounds from their conjugated form. Using gas chromatographic-mass spectrometric analysis they were identified as 5-hydroxy-6-methoxy, 6-hydroxy-5-methoxyindole and 5,6-dihydroxyindole.

IDENTIFICATION OF THORMAHLEN-POSITIVE COMPOUND-B IN URINE OF PATIENTS WITH MALIGNANT-MELANOMA

SummaryThe identification of the entire structure of the Thormählen-positive compound B from melanotic urine is described. The compound was separated from other Thormählen-positive compounds using DEAE-cellulose column chromatography. On the basis of differential enzymic hydrolysis followed by gas chromatography-mass spectrometry analysis and comparison with synthetically prepared compounds it is pissible to conclude that the Thormählen-positive compound B is a mixture of O-sulphate 5-hydroxy-6-methoxyindole and 6-hydroxy-5-methoxyindole with predominance of the latter.

The excretion of Thormählen positive melanogens in melanoma patients and its clinical significance

Abstract Thormahlen positive melanogens (TPM) were determined in the urine of 585 primary and/or metastatic cutaneous melanoma patients, in 105 primary and/or metastatic ocular melanoma patients, in 100 patients suffering from other diseases with the exception of melanoma and in 72 healthly persons. On the basis of our results we can recommend the determination of the sum of TPM as sufficient for routine monitoring of patients in the course of malignant melanoma. The determination of TPM urinary excretion cannot substitute histological and clinical examinations in an early diagnosis of malignant melanoma, but it is very important and useful for a differential diagnosis and prognosis of malignant melanoma. TPM urinary excretion can be followed in melanoma patients also for the indication of further treatment and the evaluation of the response on chemotherapy or immunotherapy.

Incorporation of L-3,4-dihydroxy-[2-14C]phenylalanine into hamster melanoma melanosomes

Melanosomes are specific particles of pigment’cells [ 11. They have been studied by classical isolation methods [2,3]. The isolation procedures aim at obtaining homogenous samples of melanosomes but neglect quantitative aspects [2]; consequently we lack information both on the amount of melanosomes in pigment tissue and on the recovery by the separation methods. As melanocytes possess a unique metabolic pathway towards melanins with L3,4dihydroxyphenylalanine (Dopa) as the specific precursor [ 1,4,5] and, as on the subcellular level, melanin is produced in melanosomes [ 1,6], experiments with labelled Dopa could yield both quantitative data, and give more information on the origin of Dopa, repeatedly found in melanosome hydrolysates [7-91. Here, we report on melanosome concentration in melanoma tissue and show that part of the [2-*4C]Dopa incorporated into melanosomes can be released by hydrolysis.

Melanogenuria in hamsters with transplantable melanoma.

Abstract Little has been known about the excretion of melanogens (melanogenuria) in animals with experimental melanoma. In this report the first information about melanogenuria in hamsters with transplantable melanotic and amelanotic melanoma is given. In hamsters with melanotic melanoma apparently increased concentration and excretion of so called Thormahlen positive melanogens in urine was found in comparison with hamsters with amelanotic melanoma or hamsters without tumours. Similar results were obtained when the excretion of all diazo positive compounds in urine was measured. Thin-layer chromatography of ethyl acetate extracts of hamster urine proved the presence of at least 15 phenolic or indolic acids. Some of them remained unidentified. However, such compounds as p-hydroxyphenylacetic acid, homovanillic acid, vanilmandelic acid or 5-hydroxyindole-acetic acid were identified. Only in the urine of hamsters with melanotic melanoma were we able to detect the isomeric 5-hydroxy-6-methoxyindolyl-2-carboxylic and 6-hydroxy-5-methoxyindolyl-2-carboxylic acids.