Development and validation of a new HPLC-MS method for meglumine impurity profiling.

We report a new High Performance Liquid Chromatography-Mass Spectrometry (HPLC-MS) method to rapidly detect and quantify meglumine by-products (specifically reducing sugar(s) and nitrogen impurities) that could be present in the meglumine samples. Meglumine is a secondary amine obtained from glucose and it is an excipient used as counter-ion in several pharmaceutical formulations, especially when the concentration of the active pharmaceutical ingredient (API) is so high that the sodium is not a suitable option. Moreover, the increased use of meglumine is related to its ability to improve solubility in aqueous solutions due to the presence of a large number of hydroxyl groups. Thus, even if meglumine is widely used as excipient in pharmaceutical formulations, its impurity profile has never been fully evaluated. Here, we propose the use of a commercial agent that specifically reacts with carbonyl compounds, 1-(4-aminobenzyl)-1,2,4-triazole, with the aim of improving the detection of reducing sugars, such as glucose, after an easy derivatization procedure. Finally, we describe the method validation and the analysis of the impurity profile of meglumine samples from different manufacturers.

In Vivo MR Imaging of Fibrin in a Neuroblastoma Tumor Model by Means of a Targeting Gd-Containing Peptide.

PURPOSE: A magnetic resonance imaging contrast agent based on a tetrameric Gd-DTPA-like system linked to a fibrin-targeting peptide (Gd-F) has been designed for in vivo tumor characterization. PROCEDURES: Gd-F was synthesized following Fmoc-SPPS strategy. Binding was measured using soluble fibrin DD(E) fragment and a dried fibrin assay. Contrast efficiency was tested on human and mouse clots and in vivo on Neuro2A tumor model. An anti-thrombotic drug was used to evaluate Gd-F sensitivity for changes in fibrin availability at the tumor site. RESULTS: The high relaxivity of Gd-F (42 mM(-1) s(-1), per molecule) yielded a strong signal enhancement in human and murine clots. High contrast was also measured in vivo in Neuro2A tumors, with a persistent enhancement in tumor rim and stroma. Upon treatment with an anti-thrombotic drug, the contrast uptake was significantly reduced in the tumor area confirming the specificity of the probe. CONCLUSIONS: Gd-F resulted to be an efficient probe for tumor delineation and for monitoring fibrin deposits during tumor progression and anti-thrombotic therapy.

Mitochondrial DNA sequence variations in some Italian wild boar populations.

In order to investigate the relationships between Italian wild boar and major pig breeds, we studied the genetic variability of four wild boar populations in Italy (Arezzo, Pisa, Parma, Bergamo) using a 533-bp fragment of the mitochondrial control region. Sixty-nine wild boar samples were analysed, allowing the identification of 10 distinct haplotypes, which involve a total of 15 single nucleotide polymorphisms. Phylogenetic and network analyses were performed also considering several sequences of wild and domesticated forms available in the databases. The Bayesian phylogenetic tree and the Median-Joining network analyses show three main groups: the Italian (IT), European (EU) and Asian (AS) clades. The IT clade corresponds to the Maremma endemic wild boar population and also includes Sardinian individuals, while the EU and AS groups include wild boars as well as domestic pig breeds. Only two individuals from Pisa cluster in the IT group, whereas two haplotypes from Bergamo cluster in the AS group and all other samples cluster in the EU clade. These findings suggest that in Italy wild boar populations have a mixed origin, both EU and AS, and that an interbreeding between wild and domesticated strains has probably occurred. Eight of the 10 wild boars coming from the Migliarino-San Rossore-Massaciuccoli Regional Park (Pisa) belong to H2 and H3 haplotypes, and cluster into the EU clade, suggesting that this regional park is not anymore exclusive of the endemic Maremma wild boar.

[Evaluation of the appropriateness of public hospital use using AEP in the Abruzzo Region of Italy: results by hospital and ward, main predictors and potential implications]. / Valutazione dell'appropriatezza dei ricoveri ospedalieri pubblici nella regione abruzzo tramite PRuo: risultati per presidio e disciplina, principali predittori e potenziali implicazioni.

In the context of a wide healthcare system reorganization, the Abruzzo Region of Italy used the Appropriateness Evaluation Protocol (AEP) to carry out a systematic evaluation of the appropriateness of the admissions performed during 2006 in all public hospitals. After specific training courses, a sample representative of all ordinary admissions was assessed twice: first by regional investigators (external assessment) then by the local personnel (internal assessment). Random-effect logistic regression was used to evaluate potential inappropriateness predictors. On a total of 13081 hospital days (2393 hospitalizations), 39.7% (95% Confidence interval: 38.9%-40.6%) were inappropriate at the regional assessment; 39.5% at the internal assessment, with high correlation between the two controls (K = 0.73). Another 10.4% of admissions, excluded by the evaluation, was assigned to DRGs at high risk of inappropriateness and should be considered. In single hospitals, the inappropriateness ranged between 17.9% and 57.9%, with large variation across wards. Additional significant predictors of inappropriateness were the day and hour of admission and hospital size, with lower inappropriateness in bigger ones. In 2006, there was a large degree of hospital misuse in public hospitals in the Abruzzo Region. The approach used in the survey may have contributed to the drastic reduction of the number of ordinary admissions observed in the Region between 2006 and 2007.

ENA screen chemiluminescence immunoassay: a random access method in autoimmunology.

Recently, autoimmunity, due to an increase in examination requests, has become an independent area of laboratory research, which needs management optimization in terms of quality, time, and flexibility. Therefore, we have evaluated the screening of extractable nuclear antigens (ENA) antibodies both with a chemiluminescence immunoassay and the enzyme-linked immunosorbent assay (ELISA) method, which was used in our laboratory, as a reference kit. The most important difference between these two methods is the possibility of processing serum samples with a random access system, which is different from batch methods.

Effects of 2 patterns of prehospital care on the outcome of patients with severe head injury.

HYPOTHESIS: A pattern of prehospital care combining advanced life support, physician staffing, and helicopter transport improves the outcome of patients with severe brain injuries, compared with combined expanded basic life support, nurse staffing, and ground transport. DESIGN: Inception cohort from the data set of a population-based, prospective study on major trauma. SETTING: Prehospital and hospital trauma systems of an Italian region. PATIENTS: All patients with major trauma (Injury Severity Score, >or=16) and severe head injury (Abbreviated Injury Scale score for the head, >or=4) rescued alive from March 1, 1998, to February 28, 1999, who received either form of care. Patients with self-inflicted injuries were excluded. The 184 patients who met the entry criteria were divided equally between care groups. INTERVENTIONS: None. MAIN OUTCOME MEASURES: Mortality at 30 days and Glasgow Outcome Scale score of survivors. RESULTS: After verifying the comparability of the cohorts, no survival or disability benefit could be demonstrated (95% confidence interval [CI] of the odds ratio for mortality [helicopter/ambulance] [95% CI 1], 0.72 to 2.67; 95% CI of the difference in Glasgow Outcome Scale score medians between helicopter and ambulance groups [95% CI 2], 0.0 to 0.0). Similar results were derived from analyses restricted to the subgroups identified by low (

DOTA tris(phenylmethyl) ester: a new useful synthon for the synthesis of DOTA monoamides containing acid-labile bonds.

The synthesis of DOTA tris(phenylmethyl) ester 2, a new monoreactive derivative of DOTA, is described. This versatile synthon can be easily coupled to compounds bearing an amino group and then deprotected to DOTA monoamide under mild and neutral conditions by catalytic hydrogenolysis. Accordingly, compound 2 has been used in the synthesis of a DOTA monoamide gadolinium complex containing two palmitic esters, which is a component of mixed micelles as MRI contrast agents.

A population-based study on pneumothorax in severely traumatized patients.

BACKGROUND: Pneumothorax (PNX) is a recognized cause of preventable deaths in trauma patients. Our objective was to determine the incidence of traumatic PNX, the characteristics of its victims, and the treatment they receive. METHODS: The study consisted of data set of a population-based study on major trauma. RESULTS: The incidence of PNX was 81 per 1 million population per year, mostly caused by transport accidents. PNX victims generally had multiple injuries, and they showed on-scene clinical parameters worse than victims of other chest injuries of comparable severity. Fifty-three percent of PNXs were drained during the prehospital and early (< 2 hours) hospital course. There was no uniformity of treatment among different types of rescue facilities, some of them never performing decompression despite clinical need. The z statistic for mortality was -0.63. CONCLUSION: PNX can be expected in one in five major trauma victims found alive. PNX is associated with a peculiar on-scene instability. Early decompression is often required. The effects of wider access to prehospital decompression and the reasons for its uneven availability in our setting need elucidation. Nevertheless, the present mortality follows the international standards.

Mixed micelles containing lipophilic gadolinium complexes as MRA contrast agents.

Mixed micelles for MRA are multicomponent systems containing a phospholipid, a biocompatible non-ionic surfactant (e.g. Synperonic(R) F-108) and a lipophilic gadolinium complex. A variety of lipophilic gadolinium complexes were designed taking into account features such as: (i) nature of ligand (cyclic versus acyclic); (ii) lipophilic moiety; (iii) global charge of the complex; and (iv) nature of bond connecting the complex to the lipophilic moiety. All the lipophilic gadolinium complexes after formulation as mixed micelles show high relaxivities in water and in blood (rat). Mixed micelles containing gadolinium complexes bearing only one aliphatic chain cannot be used as MRA contrast agents because they have a high haemolytic effect. Furthermore, in rats they are quickly eliminated from the blood stream. These drawbacks are completely circumvented using gadolinium complexes bearing two aliphatic chains. Mixed micelles containing such complexes show high relaxivities, no haemolytic effect and long blood permanence. This makes them promising candidates as MRA contrast agents. However, elimination, which occurs exclusively through the liver, is not complete, even after 7 days. Complexes containing labile (e.g. ester) bonds between the lipophilic moieties and the chelate subunit are eliminated through both the liver and the kidneys. However, elimination is still not complete after 7 days.

Antibiotic prescription in italian children: a population-based study in Friuli Venezia Giulia, north-east Italy.

UNLABELLED: Comprehensive information on prescription patterns of antibiotics in Italy is scarce. This study describes the use of systemic antibiotics in children according to age and sex in Friuli Venezia Giulia, north-east Italy. A pharmacological prescription database was used to identify individual prescriptions provided to all 0-15-y-old resident children (n = 140,630) during 1998. Overall, 124,383 prescriptions were identified. The prescription rate was highest in the 3-6 y olds, with 1491 antibiotic prescriptions per 1,000 children per year. Antibiotics were prescribed for 52% of infants, 57.2% of toddlers and 62% of preschool children. Twenty-nine percent of the prescriptions were for cephalosporins, 27% for macrolides and 24% for broad-spectrum penicillins. Prescription rates were much higher than in other countries such as Denmark, with more antibiotic courses prescribed for more children at all ages. Prescriptions from general practitioners and family paediatricians often included second-line antibiotics (e.g. cephalosporins and macrolides) or antibiotics that have not been approved for community-acquired paediatric infections (e.g. quinolones). CONCLUSION: The development of regional guidelines for antibiotic use in children should be urgently recommended.

Epidemiological study on high grade trauma. Friuli VG Major Trauma Study Group.

A better understanding of trauma epidemiology may allow to enhance the organisation of trauma systems with a potentially relevant impact on the level of trauma care. A one year epidemiology study (1st March 1998-28th February 1999) was planned in Friuli Venezia Giulia with the aim to collect all prehospital, hospital and outcome data of patients who sustained a major trauma (ISS > 15) within the regional border. In 12 months 15,429 traumatized patients (14,108 residents) were admitted to any one of the Regional hospitals. Over 1% of the whole population sustained injuries severe enough to cause hospital admission. 630 people (77.3% male, 27.7% female average age 42 ys) had a major trauma. The incidence of major trauma is 525 per million people per year. RTA was by far the most important cause of major injuries (78.6%) followed by work accidents (6.8%), domestic (5.9%) and sport accidents (1.9%). Only 1.2% of all the major injuries was the consequence of interpersonal violence. One hundred-sixty-six trauma victims died on the spot (149) or before hospital arrival (17). 464 patients with major injuries reached the hospital alive. More than two third of the patients with ISS > 15, suffered from a multiple trauma. 70% had a severe injury to the head (AIS > or = 3). Head trauma occurred as an isolated injury in only 35.3%. Hospital mortality within 30 days from admission (trauma death) was 25.1%. The results of the follow-up at 6 months are still incomplete. However the preliminary data clearly show that a high percentage of the patients who were discharged alive from the ICU had a good neurologic recovery.

L-Glutamic acid and L-lysine as useful building blocks for the preparation of bifunctional DTPA-like ligands.

Bisalkylation of suitably protected L-glutamic acid and L-lysine derivatives with tert-butyl N-(2-bromoethyl)iminodiacetate 2, followed by deprotection of the omega functional group affords N, N-bis[2-[bis[2-(1, 1-dimethylethoxy)-2-oxoethyl]amino]ethyl]-L-glutamic acid 1-(1, 1-dimethylethyl) ester 4 and N2,N2-bis[2-[bis[2-(1, 1-dimethylethoxy)-2-oxoethyl]amino]ethyl]-L-lysine 1,1-dimethylethyl ester 7. Such compounds feature a carboxylic or an amino group, respectively, which are available for conjugation with a suitable partner via formation of an amide bond. The conjugates, which can be prepared in this way, contain a chelating subunit in which all five acetic residues of DTPA are available for the complexation of metal ions. Direct bisalkylation of glycine with 2 promptly gives N, N-bis[2-[bis[2-(1,1-dimethylethoxy)-2-oxoethyl]amino]ethyl]glycine 11. The latter allows to achieve conjugates in which the central acetic group of DTPA is selectively converted into an acetamide.

Hepatocyte-directed MR contrast agents. Can we take advantage of bile acids?

A series of gadolinium complexes conjugated to bile acids was prepared and investigated as possible hepatospecific MR imaging contrast agents. In the design of such compounds, features such as the nature of the bile acid, the site of conjugation on the bile acid skeleton, and the global charge of the conjugate were taken into account. Relaxivity measurements carried out in human serum indicate interaction of the conjugates with human serum proteins; even small structural variations significantly affect relaxivity in human serum. Pharmacokinetic data (biliary elimination in the range of 18.4-45.6%) show that bile acids can be used as address moieties to transport gadolinium complexes through hepatocytes. For a homogeneous series of compounds, differing only in the bile acid residue conjugated, it was unexpectedly found that cholic acid is twice as efficient an address moiety as cholylglycine or cholyltaurine. Preliminary results show that none of the conjugates is transported through the basolateral membrane of hepatocytes by the Na+/taurocholate carrier.

[Dental caries in a group of Carnia children. Epidemiological study]. / Carie dentarie in un campione di bambini della Carnia. Studio epidemiologico.

The purpose of the research is to analyse the prevalence of dental caries of children living in Carnia divided into 2 groups, one of which was treated with fluoride tablets since the first months of life. In the considered area there is a reduction of dental caries even if the problem remains a major one. The prevention programme, based on collaboration with relatives from the first months of their babies, can be considered a valid operative support.

[Changes in the circadian rhythm of urinary cyclic adenosine monophosphate during the alcoholic withdrawal syndrome and related neurobiological correlations]. / Alterazioni del ritmo circadiano dell'adenosinmonofosfato ciclico urinario in corso di sindrome da astinenza alcolica e relative correlazioni neurobiologiche.

Urinary cyclic adenosine monophosphate (AMP-c) was measured morning and evening in 35 patients with alcohol withdrawal syndrome (AWS). 65% of the patients revealed a higher night time than day time concentration of AMP-c in the urine, reflecting increased sympathetic adrenergic activity. The circadian rhythm was lost in 88.55% of the 35 patients. The pathogenic factors and mechanisms involved in AWS are discussed and the contribution of sympathetic adrenergic hyperactivity to the onset of the withdrawal syndrome with its concomitant depression of the cholinergic and GABAergic systems is emphasised. Finally it is suggested that insomnia and the loss of REM sleep may also contribute to the onset of the condition.