This work was aimed to implement a fast and simple method to quantify cholesterol (CHOL) and 7-dehydrocholesterol (7-DHC) in dried blood spot (DBS) to diagnose the Smith-Lemli-Opitz syndrome (SLOS), an inborn error of CHOL biosynthesis. We developed and validated a GC-FID method for separation and quantification of underivatized CHOL and 7-DHC using a DBS disc of 6 mm with a run time of 9. min. Correlation coefficients (r) of calibration curves ranged from 0.998 to 0.999 for CHOL and from 0.997 to 0.998 for 7-DHC. Within-day and between-day imprecision (CV%), accuracy (%), carry-over, and extraction efficacy (%) were also evaluated for validation. CHOL and 7-DHC were analyzed in DBS and plasma samples from 8 SLOS patients and 30 unaffected subjects. In SLOS patients, 7-DHC/CHOL ratios in DBS and plasma samples ranged from 0.035 to 1.448 and from 0.012 to 0.926, respectively. Results from calibration curves, quality controls and patient samples reveal that the method is suitable to analyze DBS to screen patients affected by SLOS.

Ascending aortic aneurysm (AsAA) is a consequence of medial degeneration (MD), deriving from apoptotic loss of smooth muscle cells (SMC) and fragmentation of elastin and collagen fibers. Alterations of extracellular matrix structure and protein composition, typical of medial degeneration, can modulate intracellular pathways. In this study we examined the relevance of superoxide dismutase (SOD3) and Akt in AsAA pathogenesis, evaluating their tissue distribution and protein levels in ascending aortic tissues from controls (n=6), patients affected by AsAA associated to tricuspid aortic valve (TAV, n=9) or bicuspid aortic valve (BAV, n=9). The results showed a significant reduction of SOD3, phospho-Akt and Akt protein levels in AsAA tissues from patients with BAV, compared to controls, whereas the differences observed between controls and patients with TAV were not significant. The decreased levels of SOD3 and Akt in BAV aortic tissues are associated with decreased Erk1/Erk2 phosphorylation and MMP-9 levels increase. The authors suggest a role of decreased SOD3 protein levels in the progression of AsAA with BAV and a link between ECM modifications of aortic media layer and impaired Erk1/Erk2 and Akt signaling in the late stages of the aortopathy associated with BAV.

Hepatocellular carcinoma (HCC) is a very aggressive neoplasia requiring early and accurate diagnosis to improve patient outcomes with timely treatment. The liver is also very frequently colonized by metastases, and the most frequent differential diagnosis is HCC against intrahepatic cholangiocarcinoma or metastatic adenocarcinoma. Metabolomics is a powerful tool for identification of altered biomarkers in cancer, and to evaluate the efficacy of drug treatments. Here we analyzed by HILIC-MS/MS methylated arginines, basic amino acids (Arg, Cit, Orn), and their ratios in the extracts of primary HCC tissues, liver metastases from colorectal carcinoma (MET), cirrhotic related hepatitis-C-virus (CIR), and non-cirrhotic normal liver (NT) adjacent tissues. We found high levels of Arg (p < 0.0001) and Arg/Orn (p < 0.01) in MET compared to other tissues. In MET, compared to NT and CIR, Arg concentration was fivefold higher, while in HCC it was twofold higher. ADMA increased twofold compared to NT and CIR, while in HCC it was 50 % higher. Arg/Cit and ADMA/SDMA ratios were significantly higher in MET compared to NT and CIR (p < 0.005). Arg/Orn, Arg/Cit, and ADMA/SDMA ratios increased progressively from NT, CIR, HCC, to MET tissues. Arg/Cit correlated significantly with Arg/Orn ratios (r = 0.77; p < 0.0001), and discriminates tumor from non-tumor samples. In addition, the discriminant lactate/glucose ratio we previously found by NMR, also correlated significantly with the Arg levels (r = 0.64; p < 0.0001), and discriminated MET from all other tissues. The results indicated that Arg in MET is higher than other tissue classes, suggesting that, together with the lactate/glucose ratio, it can be considered a further biomarker for HCC-metastases differentiation.

Spectroscopic and MS techniques were used to characterize the pigments and the composition of polar and nonpolar binders of a stray wall painting fragment from Liternum (Italy) archaeological excavation. X-ray fluorescence and diffraction analysis of the decorations indicated mainly the presence of calcite, quartz, hematite, cinnabar, and cuprorivaite. Infrared spectroscopy, GC coupled to flame-ionization detector, and MS analysis of the polar and nonpolar components extracted from paint layers from three different color regions revealed the presence of free amino acids, sugars, and fatty acids. Interestingly, LC-MS shotgun analysis of the red painting region showed the presence of S1-casein of buffalo origin. Compared to our previous results from Pompeii’s wall paintings, even though the Liternum painting mixture contained also binders of animal origin, the data strongly suggest that in both cases a tempera painting technique was utilized.

To achieve clinical validation of cutoff values for newborn screening by tandem mass spectrometry through a worldwide collaborative effort. Methods: Cumulative percentiles of amino acids and acylcarnitines in dried blood spots of approximately 25–30 million normal newborns and 10,742 deidentified true positive cases are compared to assign clinical significance, which is achieved when the median of a disorder range is, and usually markedly outside, either the 99th or the 1st percentile of the normal population. The cutoff target ranges of analytes and ratios are then defined as the interval between selected percentiles of the two populations. When overlaps occur, adjustments are made to maximize sensitivity and specificity taking all available factors into consideration. Results: As of December 1, 2010, 130 sites in 45 countries have uploaded a total of 25,114 percentile data points, 565,232 analyte results of true positive cases with 64 conditions, and 5,341 cutoff values. The average rate of submission of true positive cases between December 1, 2008, and December 1, 2010, was 5.1 cases/day. This cumulative evidence generated 91 high and 23 low cutoff target ranges. The overall proportion of cutoff values within the respective target range was 42% (2,269/5,341). Conclusion: An unprecedented level of cooperation and collaboration has allowed the objective definition of cutoff target ranges for 114 markers to be applied to newborn screening of rare metabolic disorders.

BACKGROUND: Although the pigment composition of Pompeian wall paintings has been the object of several studies, a comprehensive characterization of paint binder components is still lacking. This work aimed investigated at a molecular level the binder composition differences among wall paintings belonging to different periods of Pompeii's history. Analytical investigations were performed on representative samples of the first, second, third, and fourth painting styles excavated from the house of Marcus Fabius Rufus (Insula Occidentalis). The application of sensitive experimental methodologies was complemented by historical knowledge to gain insight in painting techniques and materials used by Pompeian artists. RESULTS: Fourier transform infrared spectroscopy and Raman spectroscopy were used to investigate the organic components and pigments present in powders obtained from samples of the four painting styles. No proteinaceous components were detected in the samples with liquid chromatography-electrospray ionization-hybrid quadrupole/time-of-flight mass spectrometry. Liquid chromatography, gas chromatography with flame-ionization detection, and gas chromatography-mass spectrometry of polar and non-polar components extracted from powders were used to evaluate and compare the free amino acids, sugars, and fatty acids profiles. CONCLUSIONS: Pigments and natural products (lipids, gums and wheat flours) were the main components of all samples. This supports the hypothesis that artists likely used water tempera for Pompeian wall paintings. Graphical AbstractScheme of the multi-analytical approach followed to compare Pompeian paint binders composition.Scheme of the multi-analytical approach followed to compare Pompeian paint binders composition.

Background: High serum cholesterol represents a risk factor for cardiovascular disease. This study aims to quantify total cholesterol in dried blood spot (DBS) by direct enzymatic method. Methods: Three hundred seventeen blood samples with serum cholesterol level ranging from 81 to 337 mg/dl were collected. DBS were manually prepared, cholesterol was extracted using methanol and analyzed by a manual enzymatic method. DBS cholesterol method was validated for imprecision and extraction efficacy. DBS cholesterol values were correlated (training test) with serum values measured by automated enzymatic method (reference method). The obtained correlation was used for predicting serum cholesterol from DBS analysis of a new sample group (validation test, n = 58). Results: Within-day and between-day coefficient of variation (CV%) were lower than 7.69 and 6.32, respectively. Residual cholesterol in DBS after extraction was 16%. DBS cholesterol and serum cholesterol showed a linear correlation (slope = 0.5217; r = 0.9139) and a bias of -28%. Furthermore, DBS cholesterol values of validation test (n = 58), converted using the training test correlation, were not statistically different compared to the corresponding plasma values (P = 0.9487), and the comparison by Passing and Bablok showed a linear regression with a slope of 1.068 (r = 0.611) and a bias of -0.22%. Conclusions: The results show that this enzymatic method is suitable to analyze cholesterol in DBS and it could be automated and used for population screening of total blood cholesterol.

Element profiling is an interesting approach for understanding neurodegenerative processes, considering that compelling evidences show that element toxicity might play a crucial role in the onset and progression of Alzheimer's disease (AD). Aim of this study was to profile 22 serum elements in subjects with or at risk of AD. Thirtyfour patients with probable AD, 20 with mild cognitive impairment (MCI), 24 with subjective memory complaint (SMC) and 40 healthy subjects (HS) were included in the study. Manganese, iron, copper, zinc, selenium, thallium, antimony, mercury, vanadium and molybdenum changed significantly among the 4 groups. Several essential elements, such as manganese, selenium, zinc and iron tended to increase in SMC and then progressively to decrease in MCI and AD. Toxic elements show a variable behavior, since some elements tended to increase, while others tended to decrease in AD. A multivariate model, built using a panel of six essential elements (manganese, iron, copper, zinc, selenium and calcium) and their ratios, discriminated AD patients from HS with over 90% accuracy. These findings suggest that essential and toxic elements contribute to generate a distinctive signature during the progression of AD, and their monitoring in elderly might help to detect preclinical stages of AD.

Ultravioletradiationisthemaincauseofskincancers,andmelanomaisthemostseriousform of tumor.Thereisnotherapyforadvanced-stagemelanomaanditsmetastasisbecauseoftheirhigh resistance tovariousanticancertherapies.Humanskinisanimportantmetabolicorganinwhichoccurs photoinducedsynthesisofvitaminD3from7-dehydrocholesterol(7-DHC).7-DHC,theprecursorof cholesterolbiosynthesis,ishighlyreactiveandeasilymodifiable toproduce7-DHC-derivedcompounds. The intracellularlevelsof7-DHCoritsderivativescanhavedeleteriouseffectsoncellularfunctionality and viability.Inthisstudyweevaluatedtheeffectsonmelanomacelllinesof7-DHCassuchandforthis aim weusedmuchcaretominimize7-DHCmodifications. Wefoundthatfrom12to72hoftreatment 82–86% of7-DHCenteredthecells,andthelevelsof7-DHC-derivedcompoundswerenotsignificant. Simultaneously,reactiveoxygenspeciesproductionwassignificantly increasedalreadyafter2h.After 24 handupto72h,7-DHC-treatedmelanomacellsshowedareductionincellgrowthandviability.The cytotoxiceffectof7-DHCwasassociatedwithanincreaseinBaxlevels,decreaseinBcl-2/Baxratio, reduction ofmitochondrialmembranepotential,increaseinapoptosis-inducingfactorlevels,unchanged caspase-3 activity,andabsenceofcleavageofPARP-1.These findings couldexplainthemechanism through which7-DHCexertsitscytotoxiceffects.Thisisthe first reportinwhichthebiologicaleffects found inmelanomacellsaremainlyattributableto7-DHCassuch.

INTRODUZIONE: Il rene trapiantato è esposto agli effetti dell’ischemia-riperfusione responsabili di ritardata ripresa funzionale dell’organo (delayed graft function; DGF). DGF può incidere negativamente sull’evoluzione del rene trapiantato.Nel presente studio è stato valutato il ruolo della lipocalina urinaria quale predittore di DGF. MATERIALI E METODI: Sono stati valutati pazienti sottoposti a trapianto di rene da donatore cadavere. Prelievi ematici erano effettuati immediatamente prima del trapianto. Le urine erano raccolte per le 24 ore successive al trapianto.DGF era definita dalla necessità di trattamento dialitico entro la prima settimana dal trapianto. RISULTATI: Sono stati valutati 20 pazienti. I pazienti che avevano una rapida ripresa funzionale del rene trapiantato (NO-DGF) erano 14 (70%). DGF era osservata in 6 pazienti. L’età media nei DGF era superiore (58±6 Vs 51±11, p=0.001). Nei pazienti DGF risultavano significativamente ridotta la diuresi (57±35 Vs 4150 ± 2230 ml/24h; p=0.001) e la escrezione urinaria di creatinina (191±184 Vs 683±660 mg/24h; p=0.001), misurate nel primo giorno successivo al trapianto. Non erano osservate significative differenze tra pazienti DGF e NO-DGF per la escrezione urinaria di lipocalina (1,20±2,20 Vs 2,44±4,0 mg/24h; p<0.20). In univariata, DGF risultava associata negativamente alla diuresi (r2=-0.795, p=0.001) ed alla escrezione urinaria di creatinina (r2=-0.480, p=0.037) e positivamente all’età (r2=0.446, p=0.049). In multivariata diuresi (p=0.014) ed escrezione urinaria di creatinina (p=0.039) erano associati a DFG. CONCLUSIONI: Lipocalina urinaria, misurata nel giorno successivo al trapianto renale, non è biomarcatore predittivo di DGF. I risultati del presente studio possono essere stati influenzati dal campione limitato di pazienti e dalla bassa incidenza di DGF.

We report the outcome of liver transplantation (LT) in the only surviving patient with lathosterolosis, a defect of cholesterol biosynthesis characterized by high lathosterol levels associated with progressive cholestasis, multiple congenital anomalies and mental retardation. From her diagnosis at age 2 she had shown autistic behavior, was unable to walk unaided and her sight was impaired by cataracts. By age 7 she developed end-stage liver disease. After a soul-searching discussion within the transplantation team, she was treated with LT as this represented her only lifesaving option. At 1-year follow-up, her lathosterol levels had returned to normal (0.61mg/dL from 13.042.65) and her nutrition improved. She began exploring her environment and walking by holding onto an adult’s hand and then independently. Her brain magnetic resonance imaging (MRI) had shown a normal picture at age 1, whereas a volume reduction of white matter with ex vacuo ventricular dilatation and defective myelinization were observed before transplant. At 5-year follow-up, a complete biochemical recovery, an arrest of mental deterioration and a stable MRI picture were achieved, with a return to her every day life albeit with limitations. Timely liver transplant in defects of cholesterol biosynthesis might arrest the progression of neurological damage.

Exhaled breath condensate (EBC) collection is a noninvasive method to investigate lung diseases. EBC is usually collected with commercial/custom-made condensers, but the optimal condensing temperature is often unknown. As such, the physical and chemical properties of exhaled metabolites should be considered when setting the temperature, therefore requiring validation and standardization of the collecting procedure. EBC is frequently used in nuclear magnetic resonance (NMR)-based metabolomics, which unambiguously recognizes different pulmonary pathological states. Here we applied NMR-based metabolomics to asthmatic and healthy EBC samples collected with two commercial condensers operating at -27.3 and -4.8 °C. Thirty-five mild asthmatic patients and 35 healthy subjects were included in the study, while blind validation was obtained from 20 asthmatic and 20 healthy different subjects not included in the primary analysis. We initially analyzed the samples separately and assessed the within-day, between-day, and technical repeatabilities. Next, samples were interchanged, and, finally, all samples were analyzed together, disregarding the condensing temperature. Partial least-squares discriminant analysis of NMR spectra correctly classified samples, without any influence from the temperature. Input variables were either integral bucket areas (spectral bucketing) or metabolite concentrations (targeted profiling). We always obtained strong regression models (95%), with high average-quality parameters for spectral profiling (R(2) = 0.84 and Q(2) = 0.78) and targeted profiling (R(2) = 0.91 and Q(2) = 0.87). In particular, although targeted profiling clustering is better than spectral profiling, all models reproduced the relative metabolite variations responsible for class differentiation. This warrants that cross comparisons are reliable and that NMR-based metabolomics could attenuate some specific problems linked to standardization of EBC collection.

Pompeii's archaeological site is of utmost importance for the study of ancient remains of great historical and scientifi c relevance [Saiz-Jimenez C., 2003]. However, very few studies have been performed on the constituent materials of ancient paintings whose characterization is fundamental for the understanding of the artists painting techniques. Moreover, these analyses should provide useful suggestions for the selection of suitable conservation treatments to assure their preservation and transfer to future generations. Given the complexity of their molecular components, the analysis of ancient painting samples is a very challenging task and requires the integration of several scientifi c and humanistic disciplines. In this contest, the study of wall painting samples from a relevant archaeological site might furnish such opportunity. In particular, the comparison of binders composition employed by artists over the time might be helpful to understand changes in the painting technique. Here, we have applied an experimental strategy for the analysis of pigments and binders (polar and non-polar) of Pompeian mural painting unearthed in the archaeological area of Marcus Fabius Rufus house, Insula Occidentalis, and classifi ed as representative of the fi rst, second, third and fourth painting styles.

The use of Fourier transform infrared spectromicroscopy and mass spectrometry (MS) allowed us to characterize the composition of polar and non-polar binders present in sporadic wall paint fragments taken from Pompeii’s archaeological excavation. The analyses of the polar and non-polar binder components extracted from paint powder layer showed the presence of amino acids, sugars, and fatty acids but the absence of proteinaceous material. These results are consistent with a water tempera painting mixture composed of pigments, flours, gums, and oils and are in agreement with those obtained from a simulated wall paint sample made for mimicking an ancient “a secco” technique. Notably, for the first time, we report the capability to discriminate by tandem MS the presence of free amino acids in the paint layer.

Free carnitine and acylcarnitines play an important role in the metabolism of fatty acids. Sterols are structural lipids found in themembranes ofmany eukaryotic cells, and they also have functional roles such as the regulation of membrane permeability and fluidity, activity of membrane-bound enzymes and signals transduction. Abnormal profiles of these compounds in biological fluids may be useful markers of metabolic changes. In this review,we describe the subset of the lipidome represented by acylcarnitines and sterols, andwe summarize how these compounds have been analyzed in the past. Over the last 50 years, lipid mass spectrometry (MS) has evolved to become one of the most useful techniques for metabolic analysis. Today, the introduction of new ambient ionization techniques coupled to MS (AMS), which are characterized by the direct desorbing/ionizing of molecules from solid samples, is generating new possibilities for in situ analysis. Recently, we developed an AMS approach called APTDCI to desorb/ionize using a heated gas flow and an electrical discharge to directly analyze sterols and indirectly investigate acylcarnitines in dried blood or plasma spot samples. Here, we also describe the APTDCI method and some of its clinical applications, and we underline the common complications and issues that remain to be resolved. This article is part of a Special Issue entitled: Lipodomics and ImagingMass Spectrometry.

BACKGROUND: Low cholesterol is typically observed in the plasma of patients with cystic fibrosis (CF) contrasting with the subcellular accumulation of cholesterol demonstrated in CF cells and in mice models. However, the homeostasis of cholesterol has not been well investigated in patients with CF. METHODS: We studied the plasma of 26 patients with CF and 33 unaffected controls campesterol and β-sitosterol as markers of intestinal absorption and lathosterol as a marker of de novo cholesterol biosynthesis by gas chromatography (GC-FID and GC-MS). RESULTS: Plasma campesterol and β-sitosterol results were significantly (p=0.01) lower while plasma lathosterol was significantly higher (p=0.001) in patients with CF as compared to control subjects. Plasma cholesterol results were significantly lower (p=0.01) in CF patients. CONCLUSIONS: Our data suggest that the impaired intestinal absorption of exogenous sterols in patients with CF stimulates the endogenous synthesis of cholesterol, but the levels of total cholesterol in plasma remain lower. This may be due to the CFTR dysfunction that reduces cholesterol blood excretion causing the accumulation of cholesterol in liver cells and in other tissues contributing to trigger CF chronic inflammation

Orotic acid (OA), a marker of hereditary orotic aciduria, is usually used for the differential diagnosis of some hyperammonemic inherited defects of urea cycle and of basic amino acid transporters. This study was aimed to establish age related reference intervals of OA in urine, and for the first time in plasma, and dried blood spot (DBS) from 229 apparently healthy subjects aged from three days to 40 years. The quantification of OA was performed by a previously implemented method, using a stable isotope dilution with 1,3-[15N2]-orotic acid and hydrophilic interaction liquid chromatography–tandem mass spectrometry (HILIC–MS/MS). The method has proved to be sensitive and accurate for a quantitative analysis of OA also in DBS and plasma. According to previous studies, urinary OA levels (mmol/mol of creatinine) decrease significantly with age. The upper limits (as 99th %ile) were of 3.44 and 1.30 in groups aged from three days to 1 year (group 1) and from 1 year to 12 years (group 2), respectively; in teenagers (from 13 to 19 years; group 3) and adults (from 20 to 40 years; group 4) urinary levels became more stable and the upper limits were of 0.64 and 1.21, respectively. Furthermore, OA levels in DBS (M) also resulted significantly higher in subjects of group 1 (upper limit of 0.89) than in subjects of groups 2, 3 and 4 (upper limits of 0.24, 0.21, and 0.29, respectively). OA levels in plasma (M) were significantly lower in subjects of group 3 (upper limit of 0.30) than in subjects of groups 1, 2, and 4 (upper limits of 0.59, 0.48, and 0.77, respectively). This method was also employed for OA quantification in plasma and DBS of 17 newborns affected by urea cycle defects, resulting sensitive and specific enough to screen these disorders.

This study aimed to determine the serum levels of free L-carnitine, acetyl-L-carnitine and 34 acyl-L-carnitine in healthy subjects and in patients with or at risk of Alzheimer's disease. Twenty-nine patients with probable Alzheimer's disease, 18 with mild cognitive impairment of the amnestic type, 24 with subjective memory complaint and 46 healthy subjects were enrolled in the study, and the levels of carnitine and acyl-carnitines were measured by tandem mass spectrometry. The concentrations of acetyl-L-carnitine progressively decreased passing from healthy subjects group (mean±SD, 5.6±1.3 μmol/L) to subjective memory complaint (4.3±0.9 μmol/L), mild cognitive impairment (4.0±0.53 μmol/L), up to Alzheimer's disease (3.5±0.6 μmol/L) group (p<0.001). The differences were significant for the comparisons: healthy subjects vs. subjective memory complaint, mild cognitive impairment or Alzheimer's disease group; and subjective memory complaint vs. Alzheimer's disease group. Other acyl-carnitines, such as malonyl-, 3-hydroxyisovaleryl-, hexenoyl-, decanoyl-, dodecanoyl-, dodecenoyl-, myristoyl-, tetradecenoyl-, hexadecenoyl-, stearoyl-, oleyl- and linoleyl-L-carnitine, showed a similar decreasing trend, passing from healthy subjects to patients at risk of or with Alzheimer's disease. These results suggest that serum acetyl-L-carnitine and other acyl-L-carnitine levels decrease along the continuum from healthy subjects to subjective memory complaint and mild cognitive impairment subjects, up to patients with Alzheimer's disease, and that the metabolism of some acyl-carnitines is finely connected among them. These findings also suggest that the serum levels of acetyl-L-carnitine and other acyl-L-carnitines could help to identify the patients before the phenotype conversion to Alzheimer's disease and the patients who would benefit from the treatment with acetyl-L-carnitine. However, further validation on a larger number of samples in a longitudinal study is needed before application to clinical practice.

Considering the clinical importance of implementing a screening method for cholesterol and dehydrocholesterols in dried blood spot, this study aims to standardize the preanalytical variability of DHC in DBS for detecting SLOS by using filter paper impregnated with BHT as an antioxidant.

This study reports our experience over the last six years in the diagnosis of Smith-Lemli-Opitz syndrome and other inborn errors of cholesterol biosynthesis. Methods: Gas chromatography/mass spectrometry was used to obtain sterol profiles in plasma and erythrocyte membranes of suspected patients. Results: Plasma sterol reference values calculated in unaffected subjects (ns276) were in agreement with those previously reported. Among patients investigated from 2005 to 2010, we report 16 patients affected by Smith-Lemli-Opitz syndrome, three of whom represent new cases and 13 of whom were follow-up patients. In this period we also identified a new case of chondrodysplasia punctata 2 X-linked. The estimated incidence obtained for Smith-Lemli-Opitz syndrome was 1:93 suspected patients (1.08%). We also studied the effect of storage on the dehydrocholesterols/ cholesterol ratio in plasma and erythrocyte membranes of patients affected by Smith-Lemli-Opitz syndrome stored at –208C for up to 22 and 20 months, respectively. A significant negative linear correlation between storage time and the dehydrocholesterols/cholesterol ratio was identified in both plasma and erythrocyte membranes. The decrease in the dehydrocholesterols/cholesterol ratio in erythrocyte membranes was at least two-fold higher than in plasma. Conclusions: The results of this study may be helpful for diagnosis and interpretation of data in patients with findings suggestive of a cholesterol biosynthesis defect.

Pseudoalteromonas haloplanktis, a psychrotrophilic marine bacterium of biotechnological interest, shows anti-biofilm properties and is particularly relevant for cold storage of vacuum packed seafood. We focused our interest on the activation of cholesterol metabolism in this bacterium as the presence in its genome of a putative 3-ketosteroid-Δ1-dehydrogenase. This study reports GC-MS and LC-MS/MS profiles of sterols/steroids and their derivatives found in cell extracts of P. haloplanktis grown in a medium with a low content of cholesterol. Here, for the first time, we suggest that P. haloplanktis produces some intermediates of cholesterol catabolism, putatively identified as 24-hydroxycholest-1,4-dien-3-one-26-oic acid, chol-1,4-dien-3-one-24-oic acid, 26-hydroxycholest-4-en-3-one, and pregn-4-en-3-one-20-carboxylic acid, a finding already reported in other microorganisms. The presence of these compounds, also considered steroid precursors, produced by P. haloplanktis in vacuum packed seafood could be of interest for healthy of consumers, as well as, for biotechnological applications in pharmaceutical industry.