Lower sulfurtransferase detoxification rates of cyanide in konzo-A tropical spastic paralysis linked to cassava cyanogenic poisoning.

Using a matched case-control design, we sought to determine whether the odds of konzo, a distinct spastic paraparesis associated with food (cassava) cyanogenic exposure in the tropics, were associated with lower cyanide detoxification rates (CDR) and malnutrition. Children with konzo (N=122, 5-17 years of age) were age- and sex-matched with presumably healthy controls (N=87) and assessed for motor and cognition performances, cyanogenic exposure, nutritional status, and cyanide detoxification rates (CDR). Cyanogenic exposure was ascertained by thiocyanate (SCN) concentrations in plasma (P-SCN) and urine (U-SCN). Children with a height-for-age z-score (HAZNCHS)<-2 were classified as nutritionally stunted. CDR was measured as time required to convert cyanide to SCN, and expressed as ms/µmol SCN/mg protein or as mmolSCN/ml plasma/min. Mean (SD) U-SCN in children with konzo was 521.9 (353.6) µmol/l and was, significantly higher than 384.6 (223.7) µmol/l in those without konzo. Conditional regression analysis of data for age- and sex- matched case-control pairs showed that konzo was associated with stunting (OR: 5.8; 95% CI: 2.7-12.8; p<0.01; N=83 paired groups) and higher U-SCN (OR: 1.1; 95% CI: 1.02-1.20 per 50-µmol increase in U-SCN; p=0.02; N=47 paired groups). After adjusting for stunting and U-SCN, the odds of developing konzo was reduced by 63% (95% CI: 11-85%, p=0.03; N=41 paired groups) for each 5mmol SCN/(ml plasma/min)-increase in CDR. Linear regression analysis indicated a significant association between BOT-2 or KABC-II scores and both the HAZNCHS z-score and the U-SCN concentration, but not the CDR. Our findings provide evidence in support of interventions to remove cyanogenic compounds from cassava prior to human consumption or, peharps, enhance the detoxification of cyanide in those relying on the cassava as the main source of food.

Methadone-related fatalities: review in the Ghent district between 1978-2008.

Previous research demonstrated that Methadone Maintenance Programs (MMP) and Methadone Maintenance Treatment/Therapy (MMT) could significantly reduce the mortality risk. However, in current forensic practice, methadone ingestion can still directly or indirectly be involved in fatalities. The objectives of this study were twofold. Firstly, referring to the wide range of blood levels reported in methadone-related fatalities, we aimed to provide insight into the interpretation of a quantitative post-mortem blood concentration. Secondly, to examine and discuss possible causes, mechanisms and manners of death. During a 30-year-period, all medico-legal files at the Department of Forensic Medicine (Ghent University) were searched through, to investigate whether methadone was involved in the fatal outcome. A significant increase in the methadone-related fatalities was found since 1995, which has also been noticed in other studies. In our study (n=48), the most frequent cause of death was intoxication: only one was due to a pure methadone intoxication, whereas in all other fatal intoxications, a poly-drug intoxication was found. In this study, cardiopulmonary failure, induced by depression of the vital centres in the brainstem, was--as expected--the most important mechanism of death. When we considered the post-mortem blood levels in our study group, we observed a wide range, namely between 0.10 and 4.13 microg/ml (median: 0.54 microg/ml, mean: 0.81 microg/ml, SD: 0.14). This was in line with previous reports, although the extreme values differed. We conclude that the interpretation of post-mortem methadone blood levels is still hazardous due to e.g. difficulties to assess the individual tolerance level, the variety of surviving periods after ingestion, interfering post-mortem redistribution and the combined ingestion of methadone with other drugs. Therefore, a close collaboration between the forensic pathologist and toxicologist is recommended in order to provide a well-grounded conclusion.

Fatality due to combined use of the designer drugs MDMA and PMA: a distribution study.

We present a fatal case involving the combined ingestion of amphetamine, 3,4-methylenedioxymethylamphetamine, 3,4-methylenedioxyamphetamine, and paramethoxyamphetamine. Various postmortem specimens (e.g., several blood samples, urine, and tissue samples) were analyzed to study the distribution of the compounds and their metabolites in the human body. Quantitation took place using liquid chromatography-sonic spray ionization-mass spectrometry after pretreatment with a liquid-liquid extraction. The medico-legal findings were compatible with a disseminated intravascular coagulation induced by hyperthermia caused by the simultaneous intake of the amphetamine analogues.

Analysis of flecainide and two metabolites in biological specimens by HPLC: application to a fatal intoxication.

A few days after her admittance to a hospital for a suicide attempt with benzodiazepines, a 15-year-old girl was found dead in bed. At autopsy, no specific anatomo-pathologic cause of death was identified. Systematic toxicological analysis (HPLC-DAD, GC-NPD, and GC-MS) of postmortem blood and urine revealed the presence of high concentrations of flecainide and its two major metabolites. Flecainide is a class IC anti-arrhythmic drug causing a decreased intracardiac conduction velocity in all parts of the heart. To identify and quantitate flecainide together with its metabolites in blood, urine, and other toxicologically relevant matrices, a new method was developed using high-performance liquid chromatography with diode-array detection. All compounds were separated on a Hypersil BDS phenyl column using water, methanol, and 1.5M ammonium acetate in a gradient system. Chromatographic analysis was preceded by an optimized solid-phase extraction procedure on RP-C18 extraction columns. The flecainide concentrations in blood and urine were 18.73 and 28.3 mg/L, respectively, and the metabolites were detected only in urine at the following concentrations: 9.4 mg/L for meta-O-dealkylated flecainide and 8.59 mg/L for meta-O-dealkylated flecainide lactam. Based on these results, it was concluded that the suicide was consistent with an overdose of this anti-arrhythmic drug.

Simultaneous determination of in total 17 opium alkaloids and opioids in blood and urine by fast liquid chromatography-diode-array detection-fluorescence detection, after solid-phase extraction.

A fast liquid chromatographic method with tandem diode array-fluorescence detection for the simultaneous determination of in total 17 opium alkaloids and opioids is presented. Blank blood and urine samples (1 ml) were spiked with different concentrations of a standard mixture, as well as with the internal standard, butorphanol (2000 ng/ml). After solid-phase extraction, based on weak cation exchange (Bond Elut CBA SPE columns), the extracts were examined by HPLC-DAD-FL. By using a "high-speed" phenyl column (53 x 7.0 mm I.D., particle size 3 microm) eluted with a gradient system (A: water-methanol (90:10, v/v), B: methanol, both containing 25 mM triethylammoniumformate (pH(A) = 4.5)) all compounds could be baseline separated within 12 min. The method was validated and its applicability was demonstrated by the analysis of real-time forensic cases.

Heroin impurity profiling: trends throughout a decade of experimenting.

Heroin is still one of the most frequently abused drugs of today. All over the world, law enforcement agencies try to eradicate the illicit production and trafficking of this potent and highly addictive narcotic. To this aim, important information is provided by physical and chemical toxicological analysis of confiscated samples, with special attention for the identification and the quantification of minor components, such as the impurities related to the origin and manufacturing. By combining these data complex characterisations, i.e. impurity profiles, chemical signatures or fingerprints, can be obtained and used for comparative analysis. This review focuses on heroin impurity profiling during the 1990s, proclaimed by the United Nations as the 'Decade for Eradicating Drug Abuse'. Special attention will be given to the new trends in analytical techniques as well as in data handling strategies, so called chemometrics, to produce these profiles. The latter can be used in comparative analysis of seized heroin samples for tactical (batch-to-batch comparison) and strategic (origin determination) intelligence purposes.

Symptoms of Gulf War veterans possibly exposed to organophosphate chemical warfare agents at Khamisiyah, Iraq.

During the 1991 Gulf War, some Allied troops were potentially exposed to sarin/cyclosarin as the result of the destruction of Iraqi munitions at Khamisiyah. To evaluate the prevalence of past and current symptoms known to be associated with exposure to these chemical warfare agents, the authors conducted a computer-assisted telephone survey of 2,918 U.S. Gulf War veterans. Veterans who had participated in or witnessed the demolition in 1991 were more likely to report historical or extant symptoms than were veterans from other military units. These results should be viewed cautiously because they are based on symptoms recalled nine years after the event without precise characterization of exposure. Nonetheless, the findings suggest that symptoms consistent with low-level sarin exposure may have initially occurred, and health effects may have persisted in the veterans who were nearest to the demolition activity. Further research is warranted.

A fatal case of serotonin syndrome after combined moclobemide-citalopram intoxication.

We present a case involving a fatality due to the combined ingestion of two different types of antidepressants. A 41-year-old Caucasian male, with a history of depression and suicide attempts, was found deceased at home. Multiple containers of medication, the MAO-inhibitor moclobemide (Aurorix), the SSRI citalopram (Cipramil), and the benzodiazepine lormetazepam (Noctamid) as active substance, as well as a bottle of whiskey were present at the scene. The autopsy findings were unremarkable, but systematic toxicological analysis (EMIT, radioimmunoassay, high-performance liquid chromatography-diode-array detection [HPLC-DAD], gas chromatography-nitrogen-phosphorus detection, and gas chromatography-mass spectrometry) revealed the following: ethanol (0.23 g/L blood, 0.67 g/L urine), lormetazepam (1.65 microg/mL urine), cotinine (0.63 microg/mL blood, 5.08 microg/mL urine), caffeine (1.20 microg/mL urine), moclobemide (and metabolites), and citalopram (and metabolite). There upon, we developed a new liquid chromatographic separation with optimized DAD, preceded by an automated solid-phase extraction, for the quantitation of the previously mentioned antidepressive drugs. The results obtained for blood and urine, respectively, were as follows: Ro 12-5637 (moclobemide N'-oxide) not detected and 424 microg/mL; Ro 12-8095 (3-keto-moclobemide) 2.26 microg/mL and 49.7 microg/mL; moclobemide 5.62 microg/mL and 204 microg/mL; desmethylcitalopram 0.42 microg/mL and 1.22 microg/mL; and citalopram 4.47 microg/mL and 19.7 microg/mL. The cause of death was attributed to the synergistic toxicity of moclobemide and citalopram, both antidepressants, which, by intentional or accidental combined ingestion, can produce a potentially lethal hyperserotoninergic state. Based on the history of the case and pharmacology of the drugs involved, the forensic pathologists ruled that the cause of death was multiple drug intoxication, resulting in a fatal "serotonin syndrome," and that the manner of death was suicide.

Determination of the designer drugs 3, 4-methylenedioxymethamphetamine, 3,4-methylenedioxyethylamphetamine, and 3,4-methylenedioxyamphetamine with HPLC and fluorescence detection in whole blood, serum, vitreous humor, and urine.

BACKGROUND: The popular designer drugs 3, 4-methylenedioxymethamphetamine (MDMA) and 3, 4-methylenedioxyethylamphetamine (MDEA) can be determined in serum, whole blood, and urine, but also in vitreous humor. The latter matrix is interesting when dealing with decomposed bodies in a toxicological setting. METHODS: After extraction, chromatographic separation was achieved on a narrow-bore C(18) column by gradient elution with fluorometric detection; results were confirmed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). RESULTS: The method was linear over the range of 2-1000 microg/L for whole blood, serum, and vitreous humor, and 0.1-5 mg/L for urine. Extraction recoveries were >70%, imprecision (CV) was 2.5-19%, and analytical recoveries were 95.5-104.4%. The limit of detection (LOD) and the limit of quantification (LOQ) were 0.8 and 2 microg/L, respectively, for whole blood, serum, and vitreous humor, and 2.5 microg/L and 0.1 mg/L, respectively, for urine. Excellent correlations between the quantitative LC-fluorescence and LC-MS/MS results were obtained. We found the following concentrations in a thanatochemical distribution study in rabbits: in serum, 5.3-685 microg/L for MDMA and from the LOQ to 14.5 microg/L for 3, 4-methylenedioxyamphetamine (MDA); in whole blood, 19.7-710 microg/L for MDMA and from the LOQ to 17.8 microg/L for MDA; in vitreous humor, 12.1-97.8 microg/L for MDMA and from the LOQ to 3.86 microg/L for MDA. In routine toxicological urine samples, concentrations ranged from LOQ to 14.62 mg/L for MDA, from LOQ to 157 mg/L for MDMA, and from LOQ to 32.54 mg/L for MDEA. CONCLUSIONS: The HPLC method described is sensitive, specific, and suitable for the determination of MDMA, MDEA, and MDA in whole blood, serum, vitreous humor, and urine.

Comparison of phenyl-type columns in the development of a fast liquid chromatographic system for eighteen opiates commonly found in forensic toxicology.

We report a precise and reliable method for the detection of 18 of the most commonly found opiates on the Belgian legal and illicit market, by ion-exchange, reversed-phase high-performance liquid chromatography, using a conventional phenyl-type analytical column (150x4.6 mm I.D., particle size 5 microm) and diode-array detection. We also describe a performance (efficiency and sensitivity) comparison of this column to a recently developed "high-speed" column (53x7.0 mm I.D., particle size 3 microm) packed with the same stationary phase, and used under slightly adjusted flow and gradient conditions. The final method, using the "high-speed" column, showed a significant reduction (55%) in analysis time without loss of resolution and sensitivity.

Ethnicity in America and feeling "American".

Ethnic group acculturation remains a concern in the United States today. In the present study, the authors explored the extent to which members of three ethnic groups (White American women, African American women, and Cuban American women) perceived themselves to be "American," how much each group felt that its members were perceived as being American by White Americans, and how these perceptions related to beliefs about their own group's economic and social status. The results showed that African Americans felt American but felt that they were not perceived as such by White Americans. African Americans also reported feeling economically and socially excluded. In contrast, Cuban Americans reported neither feeling they were American nor believing they were perceived as such by White Americans, but feelings of inclusion increased with length of residence. Implications of these results for the common ingroup identity model are discussed.

Medico-legal implications of hidden thyroid dysfunction: a study of two cases.

In this paper we present two cases in which thyroid disorders were unexpectedly brought to view. The question we ponder is whether hidden thyroid dysfunction may be important in the cause, mechanism and manner of death, or just an incidental discovery during the post-mortem examination. A short literature review has been conducted in order to evaluate previously reported cases of thyroid pathology and sudden death. The significance of post-mortem evaluation of thyroid hormones (T3 and T4) will be considered briefly.

Evaluation of automated single mass spectrometry to tandem mass spectrometry function switching for comprehensive drug profiling analysis using a quadrupole time-of-flight mass spectrometer.

A liquid chromatographic mass spectrometric strategy for systematic toxicological analysis (STA) is presented using the automatic 'on-the-fly' single mass spectrometry mode to tandem mass spectrometry mode (MS to MS/MS) switching abilities of a quadrupole time-of-flight (Q-TOF) instrument. During the chromatographic run, the quadrupole is initially set to transmit all masses until (an) ion(s) reaches a certain set threshold. Thereupon, the quadrupole automatically switches to the MS/MS mode, selecting the ion(s), which are subsequently fragmented in the high-efficiency hexapole collision cell, thus generating product ions that are further mass analyzed by the TOF. By limiting the TOF spectral accumulation time in the MS/MS mode to a statistically acceptable minimum, the quadrupole almost instantly switches back to the MS mode. Qualitative information, comprising the complementary MS ([M + H](+) ion mass) and MS/MS (informative product ion profile) data, as well as quantitative information obtained by integration of the MS extracted ion chromatogram(s), can be obtained in one single acquisition. Optimization of the automatic switching parameters, such as threshold, TOF spectral accumulation time, detection window and collision energy, was carried out by injection of a mix of 17 common drugs which were not necessarily baseline separated in the chromatographic system used. Indeed, the complete separation of the drugs is not deemed necessary since up to 8 different ions can 'simultaneously' be selected for MS/MS if they reach the preset criteria. In addition, the quantitative performance of the method was defined. In a second phase, the developed method was field-tested. To that end, the resulting data from extracts of urine samples were compared with and found to be in close concordance with those obtained by a standard toxicological analysis. This innovative approach clearly holds the potential for a substantial advance in the introduction of LC/MS in STA.

Liquid chromatography-mass spectrometry in forensic toxicology.

Liquid chromatography-mass spectrometry has evolved from a topic of mainly research interest into a routinely usable tool in various application fields. With the advent of new ionization approaches, especially atmospheric pressure, the technique has established itself firmly in many areas of research. Although many applications prove that LC-MS is a valuable complementary analytical tool to GC-MS and has the potential to largely extend the application field of mass spectrometry to hitherto "MS-phobic" molecules, we must recognize that the use of LC-MS in forensic toxicology remains relatively rare. This rarity is all the more surprising because forensic toxicologists find themselves often confronted with the daunting task of actually searching for evidence materials on a scientific basis without any indication of the direction in which to search. Through the years, mass spectrometry, mainly in the GC-MS form, has gained a leading role in the way such quandaries are tackled. The advent of robust, bioanalytically compatible combinations of liquid chromatographic separation with mass spectrometric detection really opens new perspectives in terms of mass spectrometric identification of difficult molecules (e.g., polar metabolites) or biopolymers with toxicological relevance, high throughput, and versatility. Of course, analytical toxicologists are generally mass spectrometry users rather than mass spectrometrists, and this difference certainly explains the slow start of LC-MS in this field. Nevertheless, some valuable applications have been published, and it seems that the introduction of the more universal atmospheric pressure ionization interfaces really has boosted interests. This review presents an overview of what has been realized in forensic toxicological LC-MS. After a short introduction into LC-MS interfacing operational characteristics (or limitations), it covers applications that range from illicit drugs to often abused prescription medicines and some natural poisons. As such, we hope it can act as an appetizer to those involved in forensic toxicology but still hesitating to invest in LC-MS.

Segmental analysis for cocaine and metabolites by HPLC in hair of suspected drug overdose cases.

Hair samples of eight postmortem cases were analyzed in segments of 1 to 3 cm for cocaine, benzoylecgonine and cocaethylene. Samples were prepared for analysis by digestion in 0.1 M HCl and subsequent extraction with mixed-mode solid-phase extraction columns. Measurement was made by reversed-phase, narrow-bore HPLC and fluorescence detection using two laboratory-made internal standards. The concentrations were in the region of 0.29-316 ng/mg of hair for cocaine, 0.43-141 ng/mg of hair for benzoylecgonine and 0.93-1.83 ng/mg of hair for cocaethylene. All eight investigated cases had cocaine-positive segments. In six of the cases, all segments were positive, suggesting regular cocaine use and two showed in-between negative segments indicating an interruption or a change of the abuse intensity. The results showed a second, remarkable observation, i.e. enormous concentration differences (factor >150) for both cocaine and benzoylecgonine between the different subjects. Furthermore, interindividual cocaine/benzoylecgonine ratios ranged from 0.02 to 8.43. We believe these observations could in part be attributed to both some of the still existing limitations in the analytical approach(es), especially the mandatory hair washing steps, and in our still too limited knowledge of the hair incorporation processes. Nevertheless, in some cases, segmental analysis proved to be an important tool to distinguish, together with postmortem examination, deadly chronic abuse from single acute drug overdosage.

A parental history of asthma is a risk factor for wheezing and nonwheezing respiratory illnesses in infants younger than 18 months of age.

The relationship between respiratory infection and allergy as risk factors for the development of wheezing illnesses in infants has been in dispute. We hypothesized that a parental history of allergic diseases would be associated with an increased rate of respiratory infections as well as an increased rate of wheezing during infectious episodes. We prospectively evaluated 1,193 infants from birth to 18 mo of age, using bi-weekly telephone surveillance to document all respiratory events. The overall rate of respiratory illness (all RI) increased to a maximum of 10.6 illnesses/infant/year in the 7- to 9-mo age group and then leveled off in the older infants. Multivariable models adjusting for demographic variables, breast feeding, month of illness, number of siblings, and attendance at day care showed an increase in the rate of all RI in infants older than 7 mo of age who had a parental history of asthma (OR = 1.24, CI = 1.09 to 1.41) or a parental history of atopy (OR = 1.14, CI = 1.03 to 1.26). The rate of lower respiratory illnesses accompanied by wheezing was related only to a parental history of asthma (OR = 2.06, CI = 1.36 to 3.11). We conclude that all RI, most of which represent viral infections, are increased in otherwise normal infants with a parental history of asthma or atopy, whereas wheezing is related only to a parental history of asthma.

Is vitreous humour useful for the interpretation of 3,4-methylenedioxymethamphetamine (MDMA) blood levels? Experimental approach with rabbits.

As drug instability and redistribution are factors known to affect the interpretation of post-mortem blood levels, we questioned whether post-mortem vitreous humour concentrations could be useful as predictors for the MDMA load at the time of death. In a first series of in vivo experiments using rabbits, 3,4-methylenedioxymethamphetamine (MDMA) concentrations in plasma, blood and vitreous humour were studied as a function of time after intravenous (i.v.) administration of MDMA. Equilibration between the vascular compartment and vitreous humour was attained about 1 h after i.v. MDMA administration. In a second series of experiments, the post-mortem stability of MDMA in vitreous humour in relation to ambient temperature was investigated. Post-mortem MDMA concentrations in vitreous humour were closer to the ante-mortem blood levels when compared to cardiac blood samples. These preliminary investigations in the rabbit model indicate that measurements of vitreous humour concentrations could also be of interest for predicting the blood concentration at the time of death in humans.

Identification and quantitation of despropionyl-bezitramide in postmortem samples by liquid chromatography coupled to electrospray ionization tandem mass spectrometry.

A sensitive and specific method for the determination and quantitation of (despropionyl) bezitramide in postmortem samples using liquid chromatography combined with electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) is presented. The method is the result from a simple methodological transfer of a liquid chromatographic method with fluorescence detection (LC-FL) previously developed in our laboratory. A liquid-liquid back-extraction procedure using n-hexane isoamyl alcohol (93:7, v/v) as the extraction solvent is performed for a basic sample cleanup. N-Methyldespropionyl bezitramide is used as the internal standard. Chromatographic separation of the analytes of interest is achieved on a Hypersil ODS 5-micron column, using a 80:20 (v/v) mixture of 1.0 mM ammonium acetate and methanol/acetonitrile (50:50, v/v) and 1.0 mM ammonium acetate as the mobile phase. To obtain as high a sensitivity and selectivity as possible, a selected reaction-monitoring mass spectrometric technique is applied. In addition, low-energy collisional-activated dissociation (CAD) product ion spectra are recorded for a few samples. Calibration graphs are prepared for blood and urine, and good linearity is achieved over a concentration range of 1-150 ng/mL. The intra- and interassay coefficients of variation (CV%) for the analysis of quality control samples at 10 and 50 ng/mL concentration levels do not exceed 10.2% and percent of targets are within 12.1%. Postmortem samples (blood, urine, stomach contents, bile, liver, and kidney) from three fatalities, all suspected victims of drug overdoses, are analyzed, and the results are reported. The results obtained with LC-ESI-MS/MS are in close agreement with those obtained using the LC-FL method. Moreover, the isolates' identity and structure are confirmed by the CAD product ion spectra, thus allowing to make unequivocal conclusions about the prior intake of bezitramide by the three subjects.

Narrow-bore HPLC in combination with fluorescence and electrospray mass spectrometric detection for the analysis of cocaine and metabolites in human hair.

A simple, but sensitive and specific, high-performance liquid chromatographic assay for cocaine, cocaethylene, and benzoylecgonine is described. Using direct fluorometric detection, the procedure is particularly interesting for the routine analysis of human hair samples. In the sample preparation part, the hair samples are cut and washed and two internal standards with close structural resemblance to benzoylecgonine and cocaine as well as to cocaethylene are added. Subsequently, the hair samples are homogenized, hydrolyzed overnight in a 0.1 M HCl solution at 56 degrees C, and extracted on IST Confirm HCX solid-phase extraction columns. Chromatographic separation is achieved on a narrow-bore Hypersil BDS C18 column (125 x 2.1 mm, 3 microns) by gradient elution with an ammonium acetate buffer-methanol/acetonitrile mixture. For the fluorometric detection, excitation and emission wavelengths of 242 and 315 nm, respectively, are used. This analysis protocol affords a method of high sensitivity and specificity which has been fully evaluated and validated. The data presented show good accuracy and linearity with excellent reproducibility and recovery. Because unequivocal identity confirmation is mandatory in forensic applications, an extension of the analysis protocol was accomplished toward mass spectrometric detection. We succeeded in a simple methodological transfer from LC/FL to LC/ESI-MS/MS, thus providing two complementary approaches after a single, common sample-processing step. Hair samples from 29 fatalities, all known drug users and suspected victims from a drug overdose, were analyzed in this way. Of the investigated samples, 12 were positive and the concentrations found range from 0.98 to 938 ng/mg of hair for cocaine and from 1.45 to 388 ng/mg of hair for benzoylecgonine. Traces of cocaethylene were also found in two of the hair samples. The results obtained with LC/ESI-MS/MS were in close agreement with those obtained with LC/FL, positively confirming the isolates' identity and structure by means of the resulting MS/MS spectra.