Point prevalence survey of antibiotic use and healthcare-associated infections in acute care hospitals: a comprehensive report from the Marche Region of Italy.

BACKGROUND: Healthcare-associated infections (HAIs) and antimicrobial resistance (AMR) are serious health challenges. Point prevalence surveys (PPSs) are valuable tools for monitoring HAIs and AMR. AIM: To describe results of the ECDC PPS 2022 dealing with the prevalence of HAIs, antimicrobial consumption, and associated factors, in acute care hospitals. METHODS: The survey was performed in November 2022 in 14 hospitals according to the protocol proposed by the European Centre for Disease Prevention and Control. Multilevel logistic regression was performed using geographical area/hospital type as cluster variable to evaluate the factors independently associated with HAIs and antibiotics. FINDINGS: The point prevalence of HAIs was 7.43%. Patients hospitalized for longer periods were more likely to have an HAI as well as those aged 15-44 years, with a rapidly fatal disease, intubated, and with one or two devices. Antibiotics prevalence was 47.30%. Males, unknown McCabe scores, minimally invasive/non-National Healthcare Safety Network (NHSN) surgery, patients with HAIs, hospitals with a higher alcohol hand-rub consumption, hospitals with a greater number of IPC personnel, geriatric wards, and hospitals with 300-600 beds were more likely to be under antimicrobial therapy. CONCLUSION: This PPS provided valuable information on the prevalence of HAIs and antimicrobial consumption and variables associated. The high prevalence of HAIs highlights the need for improved infection control measures.

Checking rounds for isolation precautions in the control of multidrug-resistant organisms: reduction achieved.

The objective of this investigation was to analyze the effectiveness of a quality improvement initiative in limiting the spread of multidrug-resistant organisms (MDROs) in the hospital setting. During the period 2011-2013, a multimodal intervention was activated at a tertiary care center in Italy. The intervention included: laboratory-based surveillance, interdisciplinary training sessions, monitoring the adoption of isolation precautions and daily supervision provided by infection control nurses, and a monthly feedback. Time series analysis was used to evaluate the trends and correlations between the MDROs rate, intensity of checking rounds, and hospital-wide data (i.e., transfer of patients, patients' days, site of isolation, etc.). A total of 149,251 patients were included in the study. The proportion of patients undergoing transmission-based isolation precautions within 24 h from a positive laboratory finding increased from 83% in 2011 to 99% in 2013 (p < 0.05). The wards appropriately adopting the correct isolation precaution increased from 83% in 2011 to 97.6% in 2013 (p < 0.05). The frequency of controls was significantly reduced after the observation of compliance in the appropriate wards (p < 0.05). After three years, the incidence rate changed from 5.8/1000 days of stay [95% confidence interval (CI) 5.6-6.1] in 2011 to 4.7 (95% CI 4.4-4.9) in 2013 (p < 0.0001). Moreover, microorganisms isolated from different types of specimens showed variable potential for transmission (i.e., skin as the most potential and urine the least). The results demonstrate the efficacy of the multimodal intervention, with sustained reduction of MDROs rate, besides check reduction, and highlight the long-term efficacy of checking rounds in changing professionals' behaviors.

Liquid chromatographic analysis of guanidino compounds using furoin as a new fluorogenic reagent.

Furoin, a benzoin analogue, was examined as novel fluorogenic reagent for the selective and sensitive LC determination of various guanidines after pre-column derivatization. The derivatization reaction was carried out at 100 degrees C for 5 min to give adducts that were separated on a Phenomenex Synergi MAX-RP column and detected at lambda(em)=410 nm with lambda(ex)=325 nm. The reagent showed to be useful both for determining together arginine (Arg) and creatine (CT) in dietary supplements under elution isocratic conditions and for the simultaneous analysis of a variety of guanidines in biological samples (human plasma and urine) under elution gradient conditions. The detection limits ranged from 7 to 25 fmol. Recovery studies showed good results for all determined guanidino compounds (85.6-106.2%; R.S.D.=1.1-6.2%).

Development and validation of a liquid chromatographic method for the determination of ascorbic acid, dehydroascorbic acid and acetaminophen in pharmaceuticals.

A reversed-phase ion pair liquid chromatographic method (RP-LC) for the determination of dehydroascorbic acid (DHA) and ascorbic acid (AA) and also acetaminophen, which is combined in pharmaceuticals, is proposed and validated. AA and acetaminophen were analyzed directly, while DHA was determined after pre-column derivatization with 4,5-dimethyl-1,2-phenylenediamine (DMPD). The derivatization reaction was carried out under mild conditions (10min at ambient temperature) in the dark in sodium acetate buffer (80mM; pH 3.7) solution containing EDTA as metal scavenger. The chromatographic separations were performed on a Phenomenex Synergi 4u hydro-RP (150mmx4.6mm) under isocratic elution conditions, using cetyltrimethylammonium bromide (CTAB) as ion-pairing reagent in the mobile phase. Linear responses were observed for each compound. The intra-day precision (R.S.D.) was < or =1.40% and there was no significant difference between intra- and inter-day data. Recovery studies showed good results for all compounds (99.7-101.8%) with R.S.D. ranging from 0.56 to 1.82%. The limits of quantitation were about 40, 50 and 140pmol for acetaminophen, AA and DHA, respectively. The DHA impurity values found in dosage forms were < or =0.2% of AA.

Development and validation of a liquid chromatographic method for the determination of branched-chain amino acids in new dosage forms.

A reversed-phase liquid chromatographic method (RP-LC) is proposed and validated for the analysis of branched-chain amino acids (l-leucine, l-isoleucine and l-valine) in new pharmaceutical formulations. The pre-column derivatization reaction of these amino acids with 2,4-dinitrofluorobenzene (DNFB) has been investigated considering the matrix effect. The compound reacts at 60 degrees C for 10 min at pH 9 with the amino function, in presence of cetyltrimethylammonium bromide (CTAB), to give adducts that have been separated on a RP amide C16 column and detected at lambda=360 nm. Linear responses were observed for each derivative. The intra-day precision (R.S.D.) was

Anisoin: a useful pre-chromatographic derivatization fluorogenic reagent for LC analysis of guanidino compounds.

The use of anisoin as pre-chromatographic reagent for LC analysis of guanidino compounds is proposed. The reagent reacts (5 min at 100 degrees C) with guanidino function and the resulting adducts can be chromatographed under reversed-phase conditions. A fluorescence detector (lambda(ex)=325 nm; lambda(em)=435 nm) was used to detect guanidino adducts. The derivatization and chromatographic conditions were optimised by a series of experiments. Application to the determination of arginine and creatine in pharmaceuticals and arginine, guanidine, methylguanidine, guanidinosuccinic acid, beta-guanidinopropionic acid, gamma-guanidinobutyric acid, guanidinoacetic acid and homoarginine in human urine is described. Quantitation limits ranged from 6 to 30 fmol, except for creatine (510 fmol).

2,7-dimethyl-3,8-dinitrodipyrazolo[1,5-a:1',5'-d]pyrazine-4,9-dione: a new labelling reagent for liquid chromatographic analysis of amino acids.

The use of 2,7-dimethyl-3,8-dinitrodipyrazolo[1,5-a:1',5'-d]pyrazine-4,9-dione as pre-column reagent for LC analysis of amino acids is proposed. The compound reacts (30 min at 68 degrees C in presence of 0.04 M triethylamine in a dimethylsulfoxide-water mixture) with primary and secondary amino function and the stable resulting adducts can be chromatographed under reversed-phase conditions and detected at lambda=280 nm. The derivatization conditions were optimized by a series of experiments. The effect of temperature, triethylamine concentration and reagent on the reaction was investigated. The yield of the glycine derivative was found to be quantitative at a reagent amino acid molar ratio of about 6 by comparison with an authentic specimen of synthesized glycine adduct. Application of the method to quality control of commercially available oral polyaminoacid formulations is described.

LC determination of leuprolide component amino acids in injectable solution by phanquinone pre-column derivatization labelling procedure.

A sensitive LC method for the determination of leuprolide acetate component amino acids in injectable solution with fluorogenic pre-column derivatization has been developed. The derivatization reaction with phanquinone was optimised by a series of experiments. Histidine, arginine, serine, tryptophan, glutamic acid, tyrosine, methionine, isoleucine, leucine and phenylalanine were separated on a reversed-phase ODS column using as eluent a binary mixture of triethylammonium phosphate buffer-methanol, under gradient elution conditions. The derivatives were eluted in 30 min with good reproducibility. The hydrolysis reaction of the peptide was carried out at reflux with 12 N hydrochloric acid for 2 h 30 min. The intra-day accuracy of the entire procedure (hydrolysis, derivatization, LC separation) ranged from 80.5 to 109.5% of the nominal concentration of leuprolide acetate and the precision (%R.S.D.) was less than 5.8%; the inter-day accuracy was in the range 81.5-107.2% and corresponding R.S.D. values were less than 4.6%. The detection limits (signal-to-noise ratio=3) for the adducts are 30-800 fmol.

HPLC-fluorescence determination of amino acids in pharmaceuticals after pre-column derivatization with phanquinone.

4,7-Phenanthroline-5,6-dione (phanquinone) was used as a fluorogenic labeling reagent in pre-column derivatization for the quality control of amino acids in pharmaceuticals. The amino acid adducts were efficiently separated by C12 RP high-performance liquid chromatography (HPLC) using a ternary mixture of triethylamine (TEA) phosphate buffer (pH 2.5, 0.05 M)-methanol- tetrahydrofuran (THF) as mobile phase by varying composition gradient elution and detected fluorometrically. The results obtained by the proposed method were compared statistically, by means of the Student's t-test and the variance ratio F-test, with those obtained by a rapid reference method, which involved o-phthaldialdehyde (OPA) as pre-column reagent; no significant difference was found. The stronger derivatization conditions (60 degrees C, pH 8, 60 min) required for the method with phanquinone are compensated by the major stability of derivatives and by the absence of fluorescent degradation products.

Validation of a spectrophotometric method for the determination of iron (III) impurities in dosage forms.

A spectrophotometric method (lambda=535 nm) for the iron (III) impurities determination in iron protein-succinylate complex syrup using thioglycolic acid in basic ambient was proposed and validated. Assay samples were treated with 0.1 N hydrochloric acid and centrifuged to remove the interfering active drug. Linear response (r=0.9999) was observed over the range of 0.005-0.2% of the iron (III) with respect to the complex nominal concentration. The accuracy could be considered very satisfactory (recovery=97-99%). The intra-day precision (RSD) of impurity amongst six independent sample preparations, was 1.4%, and there was no significant difference between intra- and inter-day studies. Intermediate precision indicated that the assay possessed high degrees of ruggedness. The limit of quantitation was 0.005% of impurity with respect to the active drug. The results obtained for iron (III) were compared statistically with those obtained with the standard addition method by means of the Student's t-test and the variance ratio F-test; no significative difference was found.

Determination of retinoids in galenicals by column liquid chromatography with fluorescence and diode-array detection.

Simple and rapid reversed-phase gradient column liquid chromatography (LC) with fluorescence detection at different wavelengths was developed for the simultaneous analysis of all-trans, 13-cis, 9-cis retinoic acids, vitamin A palmitate and beta-carotene in galenicals. The assay results agreed with those obtained by an LC method with diode-array UV detection. A post-column on-line photochemical reactor (irradiation at 254 and 366 nm) was inserted between the LC column and the fluorescence detector to enhance the performance of the method. Two fluorescence spectra (photoreactor on and off) were obtained for each analyte which proved useful for the unambiguous identification of the various analytes.

Analysis and stability study of retinoids in pharmaceuticals by LC with fluorescence detection.

Liquid chromatographic (HPLC) methods with fluorescence detection at different wavelengths were developed for measurements of retinoic acids (13-cis and all-trans) in pharmaceutical dosage forms and components of 'retinoid solution' (all-trans retinoic acid, vitamin A palmitate and beta-carotene), a galenical of 'Di Bella therapy', using reversed phase columns under isocratic conditions. The stability of all-trans retinoic acid in cream and all-trans retinoic acid and vitamin A palmitate in 'retinoid solution' was investigated. Solid-phase extraction (SPE), using C18 sorbent was applied to the analysis of retinoic acids (9-cis, 13-cis and all-trans) in the 'retinoid solution' to obtain a practical and reliable sample clean-up. The results showed that these preparations (cream and solution) can be conveniently stored in the dark (t.a. or 2-8 degrees C): under these conditions about 86-87% of the all-trans retinoic acid initial concentration in both formulations and about 73-78% of vitamin A palmitate in the 'retinoid solution' remained after 90 days, while under sunlight exposure rapid degradation of the drugs was observed.

HPLC-fluorescence determination of equilin and equilenin in postmenopausal women's urine.

A high-performance liquid chromatographic (HPLC) method with fluorescence detection (lambda(ex) = 280 nm; lambda(em) = 410 and 312 nm) in combination with a post-column on-line photochemical derivatization is described for the determination of equilin and equilenin in urine from normal postmenopausal women after therapy with conjugated oestrogens. The column effluents were subjected on-line to UV irradiation (254 nm) and the photo-induced modifications were useful for the identification of the analytes. The conjugated (sulphate and glucuronide) forms were analysed after enzymatic or chemical hydrolysis and extracted with chloroform. Solid-phase extraction using strong anion-exchange sorbent was applied to the analysis of unconjugated oestrogen fraction to obtain a practical and reliable sample clean-up. The HPLC separations were achieved using ODS columns with a mobile phase consisting of 0.05 M triethylamine phosphate buffer (pH 4.0)-acetonitrile (64:36, v/v) at a flow rate of 1.0 mL/min. The method was accurate and reproducible; for the equilin and equilenin separation isocratic conditions were satisfactory, allowing a sensitive detection in urine samples with a detection limit of about 50 fmol for equilin (lambda(ex) = 280 nm; lambda(em) = 312 nm, after photoderivatization) and 10 fmol for equilenin (lambda(ex) = 280 nm; lambda(em) = 410 nm).

HPLC-fluorescence determination of unconjugated estrogens in pharmaceuticals.

A fluorimetric liquid chromatographic method (lambda(ex) = 280 nm; lambda(em) = 312 nm) was developed for measurements of unconjugated estrogens (estradiol and estriol) in pharmaceutical dosage forms using a reversed-phase column with water acetonitrile at different composition as mobile phase. The in vitro release profiles of three different estradiol transdermal therapeutic systems were determined through a medical-grade silicone rubber subdermal implant material membrane, using a modified Franz diffusion apparatus at 37 degrees C in presence of PEG 400. The HPLC method possesses advantages of rapidity, simplicity and accuracy.

HPLC analysis of pharmaceutical estrogens in raw materials and dosage forms.

The use of HPLC with fluorescence detection (lambda ex = 280 nm; lambda em = 410 or 312 nm) in combination with a postcolumn on line photochemical derivatization was investigated for the analysis of conjugated and unconjugated estrogens and their correlated impurities. The column effluents were subjected on-line to UV irradiation (254 nm) and the photo induced modifications were useful for the identification of the various estrogens. The proposed HPLC methods were successfully applied to the analysis of commercially available conjugated estrogens (raw materials and pharmaceuticals) and estrogen samples. The assays results relative to the pharmaceutical formulations were in agreement with those obtained by a reference HPLC method with UV detection (lambda = 280 nm).