Ortho-phthalaldehyde Research Articles

Study on the Bactericidal Effect of Ortho-Phthalaldehyde Combination Disinfectant

This study used combination of ortho-phthalaldehyde(OPA) and quaternary ammonium surfactants (QAS) with different concentration to kill Escherichia coli, Staphylococcus aureus, Candida albicans and Aspergillus niger conidia. The concentration of the combination disinfectant to kill Escherichia coli, Staphylococcus aureus and Candida albicans was 75mg/L OPA and 30mg/L QAS within 10 min, respectively. The average killing log value was above 4 when Aspergillus niger conidia was exposed in 3.75g/L OPA and 1.50g/L QAS for one hour. Then the effects of time, temperature and organic concentration on the bactericidal effect were measured. And the metal corrosion and stability of the combination disinfectant were determined. In addition, these results showed that the combination disinfectant had a high efficiency of bacteria killing and the concentration of organic affected its bactericidal effect. The average killing log value gradually increased along with the temperature within 10°C. to 40°C. And the combination disinfectant also has the advantages of low corrosive of metal, good stability and so on.

High-performance liquid chromatography evaluation of the enantiomeric purity of amino acids by means of automated precolumn derivatization with ortho-phthalaldehyde and chiral thiols.

The use of ortho-phthalaldehyde (OPA) for the derivatization of amino acids (AA) is well known. It enables the separation of the derivatives on common reversed phase columns and improves the sensitivity with fluorescence detection. With the use of a chiral thiol an indirect enantioseparation of chiral amines and AAs is feasible. The major drawback of the OPA-derivatization is the poor stability of the products. Here, a method with an in-needle derivatization procedure is optimized to facilitate a quantitative conversion of the AA with OPA and the chiral thiols N-acetyl-L-cysteine or N-isobutyryl-L-cysteine, followed by a subsequent analysis, eluding the stability issue. Both enantiomers of a single AA were separated as OPA-derivatives with a pentafluorophenyl column and a gradient program consisting of 50mM sodium acetate buffer pH=5.0 and acetonitrile. Fluorescence detection is commonly used to achieve sufficient sensitivity. In this study, the enantiomeric impurity of an AA can be detected indirectly with common UV spectrophotometric detection with a limit of quantitation of 0.04%. Seventeen different L-AAs were tested and the amount of D-AA for each individual AA was calculated by means of area normalization, which ranged from not detectable up to 4.29%. The recovery of the minor enantiomer of L- and D-AA was demonstrated for three AAs at a 0.04% level and ranged between 92.3 and 113.3%, with the relative standard deviation between 1.7 and 8.2%.

Ortho-phthalaldehyde(OPA)による呼吸器内視鏡洗浄消毒法の有効性の検討(第27回日本呼吸器内視鏡学会総会 : シンポジウム3-5)

Ortho-phthalaldehyde(OPA)による呼吸器内視鏡洗浄消毒法の有効性の検討(第27回日本呼吸器内視鏡学会総会 : シンポジウム3-5)

Protein bioaccessibility of soymilk and soymilk curd prepared with two Lactobacillus plantarum strains as assessed by in vitro gastrointestinal digestion

Abstract The current study was conducted to compare the protein bioaccessibility of soymilk and two soymilk curds generated by fermentation with Lactobacillus plantarum B1-6 and Lactobacillus plantarum 70810. Soluble protein content, degree of hydrolysis, and electrophoretic patterns were monitored during in vitro gastrointestinal digestion of the soymilk and curds. Soluble protein content of digested soymilk was 2.0–2.6 times higher than that of digested curds as measured by the Bradford assay. The degree of hydrolysis was 1.6 times higher in digested soymilk than in digested soymilk curds as determined by the ortho -phthalaldehyde (OPA) method. Electrophoretic data showed that soymilk curds had slower protein digestion rates during simulated gastric and intestinal digestion. The soy protein 7S α′, 7S α and 11S acidic subunits and a peptide of 28 kDa were observed even after simulated intestinal digestion. The different degradation profiles suggest that the soymilk curd hinders enzymatic hydrolysis during simulated gastrointestinal digestion. Industrial relevance Soymilk curd is a popular food in East and Southeast Asian countries. Lactic acid fermentation is an emerging technology to form soymilk curds that creates a product with improved functional and probiotic properties. The purpose of the present study was to compare protein degradation profiles of soymilk curds produced by fermentation with Lactobacillus plantarum B1-6 and 70810, and of soymilk using an in vitro gastrointestinal digestion system. Results of the study will expand the knowledge base of the effects of this novel coagulation technology on the nutritional characteristics, especially protein bioaccessibility of soymilk curds and thus would be expected to provide fundamental information in the future application of the emerging coagulation technology using lactic acid bacteria.

Differential proteomic analysis of platelets suggested target-related proteins in rabbit platelets treated with Rhizoma Corydalis

Context: Corydalis yanhusuo W.T. Wang (Papaveraceae) (Rhizoma Corydalis) showed inhibitory effects on rabbit platelet aggregation induced by ADP, thrombin (THR) or arachidonic acid (AA). Objective: This study separates and identifies the possible target-related platelet proteins and suggests possible signal cascades of RC antiplatelet aggregation. Materials and methods: Based on comparative proteomics, the differentially expressed platelet proteins treated before and after with 50 mg/mL RC 90% ethanol extract (for 15 min at 37 °C) were analyzed and identified by two dimensional gel electrophoresis (2-DE) and MALDI-TOF-MS/MS. To further verify the possible signalling pathways of RC antiplatelet aggregation function, the concentration of calcium (Ca2+) was measured by Fura-2/AM fluorescence (Ex 340/380 nm, Em 500 nm) (RC final concentrations of 0.0156–0.1563 mg/mL), the levels of P-selectin and cyclic guanosine monophosphate (cGMP) were quantified by ELISA (OD. 450 nm) (RC final concentrations of 0.0156–1.5625 mg/mL), and the 5-hydroxytryptamine (5-HT) level was measured using ortho-phthalaldehyde (OPT) fluorescence (Ex 340 nm, Em 470 nm) (RC final concentrations of 0.3125–1.5625 mg/mL). Results: The expression of 52 proteins were altered in rabbit platelets after the treatment and the MALDI-TOF-MS analysis indicated that those proteins include 12 cytoskeleton proteins, 7 cell signalling proteins, 3 molecular chaperone proteins, 6 proteins related to platelet function, 16 enzymes and 7 other related proteins. Furthermore, RC extract could decrease the levels of 5-HT [inhibition rate of 96.80% (p < 0.05, vs. THR-activated group) treated with 0.7813 mg/mL of RC], Ca2+ [172.73 ± 5.07 to 113.56 ± 5.46 nM (p < 0.001, vs. THR-activated group) treated with 0.0313 mg/mL of RC] and P-selectin [13.48 ± 0.96 ng/3 × 108 to 11.64 ± 0.17 ng/3 × 108 (p < 0.05, vs. THR-activated group) treated with 0.0156 mg/mL of RC], and increase in cGMP level [38.93 ± 0.57 to 50.26 ± 4.05 ng/3 × 108 (p < 0.05, vs. THR-activated group) treated with 1.5165 mg/mL of RC] in ADP (10 μmol/L), THR (0.25 u/mL) or AA-(0.205 mmol/L) activated rabbit platelets. Discussion and conclusion: The present study indicated that P2Y12 receptor might be one of the direct target proteins of RC in platelets. The signal cascades network of RC after binding with P2Y12 receptor is mediating Gαi proteins to activate downstream signalling pathways (AC and/or PI3K signalling pathways) for the inhibition of platelet aggregation.

Traceless and Chemoselective Amine Bioconjugation via Phthalimidine Formation in Native Protein Modification

ortho-Phthalaldehyde (OPA) and its derivatives are found to react chemoselectively with amino groups on peptides and proteins rapidly and tracelessly under the physiological condition via formation of phthalimidines, which provides a novel and promising approach when performing bioconjugation on native proteins. The notable advantages of this method over the existing native protein lysine-labeling approaches include a traceless process, a self-reacting, specific and fast reaction, ease of operation, and the ability to use nonhydrolyzable reagents. Its applications have been effectively demonstrated including conjugation of peptides and proteins, and generation of an active PEGlyated l-asparaginase.

Electrochemical Study of Reactivity of Orthophthalaldehyde with Primary Amines

Orthophthalaldehyde (OPA) is an aromatic dialdehyde with aldehydic groups in positions 1 and 2. The two formyl groups exhibit inductive, resonance and steric effects. Due to the inductive and resonance effects, one aldehydic group is activated by the other one and, as a result, the molecule is strongly reactive towards nucleophiles. In addition to this, the mutual ortho-position of the carbonyls enables intramolecular ring formation. The reactivity of OPA with amines is widely used since many years in the determination of amino acids [1] and in application of OPA as a biocide [2]. In the literature related to these applications the mechanism of reaction of OPA with amines is not completely understood. The present study was made in order to describe the detailed mechanism of OPA with primary amines using DC- and DP-polarography and UV/Vis spectrophotometry. The experiments were performed in aqueous buffered solutions of different pH, in non-aqueous acetonitrile and in mixed acetonitrile/H2O media with aqueous or non-aqueous stock solution of OPA. It has been observed that at pH between 7.5 and 10.5, OPA is reduced in three waves of reduction instead of two as mostly mentioned in the literature [3,4]. Its reduction is strongly influenced by pH-dependent hydration. It was found that there is a significant difference in reaction pattern, when using hydrated or non-hydrated form of OPA, respectively. Although the dialdehydic form was always considered as the species reacting with amines [5,6], the experiments showed that the reactivity of OPA with amines is directly related to the hydrated form of OPA, while the non-hydrated OPA undergoes hydration reaction prior the reaction with amines. It was also found that the reaction rate of OPA with primary amines is strongly influenced by the number of H-atoms at the α–carbon atom which may reflect the steric hindrance. Acknowledgement: The material support from the grant GACR No. 13-21704S and the institutional support RVO 61388955 are acknowledged. References (1) Norton, D. R.; Howell, N. Analytical Chemistry 1954, 26, 1116-1119. (2) Simoes, M.; Simoes, L. Journal of Basic Microbiology 2007, 47, 230-242. (3) Norton, D. R.; Mann, C. K. Analytical Chemistry 1954, 26, 1111-1115. (4) Zuman, P.; Salem, N.; Kulla, E. Electroanalysis 2009, 21, 645-649. (5) Kulla, E.; Zuman, P. Organic & Biomolecular Chemistry 2008, 6, 3771-3780.

Comparison of the efficacy of disinfectants in automated endoscope reprocessors for colonoscopes: tertiary amine compound (Sencron2®) versus ortho-phthalaldehyde (Cidex®OPA).

Background/AimsTo prevent the transmission of pathogens by endoscopes, following established reprocessing guidelines is critical. An ideal reprocessing step is simple, fast, and inexpensive. Here, we evaluated and compared the efficacy and safety of two disinfectants, a tertiary amine compound (TAC) and ortho-phthalaldehyde (OPA).MethodsA total of 100 colonoscopes were randomly reprocessed using two same automated endoscope reprocessors, according to disinfectant. The exposure time was 10 minutes for 0.55% OPA (Cidex® OPA, Johnson & Johnson) and 5 minutes for 4% TAC (Sencron2®, Bab Gencel Pharma & Chemical Ind. Co.). Three culture samples were obtained from each colonoscope after reprocessing.ResultsA total of nine samples were positive among the 300 culture samples. The positive culture rate was not statistically different between the two groups (4% for OPA and 2% for TAC, P=0.501). There were no incidents related to safety during the study period.ConclusionsTAC was non-inferior in terms of reprocessing efficacy to OPA and was safe to use. Therefore, TAC seems to be a good alternative disinfectant with a relatively short exposure time and is also less expensive than OPA.

In-Use Evaluation of Peracetic Acid for High-Level Disinfection of Endoscopes.

Many high-level disinfectants have been used for disinfection of endoscopes such as 2% glutaraldehyde (GA), 0.55% ortho-phthalaldehyde (OPA), and peracetic acid (PAA). Both GA and OPA are widely used in disinfection of endoscopes and have been previously discussed, but there is little research on the practical use of PAA as an endoscope disinfectant. An experimental model of a flexible gastrointestinal endoscope being contaminated with 9 strains of microorganism was designed. After the cleaning and disinfecting procedure was completed, we evaluated the biocidal activity (850 ppm PAA, 2% GA, and 0.55% OPA) on our flexible gastrointestinal endoscope model. We also evaluated sterilization effectiveness of PAA on other bacteria, including some antibiotic-resistant bacteria (methicillin-resistant Staphylococcus aureus, vancomycin-resistant Enterococcus, and Clostridium difficile). The residual bacterial colony count number of the PAA-disinfected endoscope was significantly lower than that of the GA- and OPA-disinfected endoscopes. The biocidal effect and efficiency of the endoscope disinfection by PAA appeared to be better than either the GA- or OPA-disinfected endoscope. PAA has demonstrated a good sterilization effect on other bacterial species; of particular note are common antibiotic-resistant bacteria such as methicillin-resistant Staphylococcus aureus, vancomycin-resistant Enterococcus, and Clostridium difficile. The results of this study demonstrate that PAA is a fast and effective high-level disinfectant for use in the reprocessing of flexible endoscopes.

Electrochemical and Quantum Chemical Study of Reactivity of Orthophthalaldehyde with Aliphatic Primary Amines

The reactivity of orthophthalaldehyde (OPA) with primary amines is a base for broad application of OPA: in the determination of amino acids, in medicine as a disinfection agent and in organic synthesis, therefore the detailed knowledge of the mechanism is of interest. The electrochemical reducibility of OPA was utilized for the investigation of its reactivity with eight aliphatic primary amines which could be separated into four types. Based on the electrochemical monitoring, the apparent reaction rates were evaluated. It was found that the reaction rate essentially depends on the substitution at the α-carbon atom of the amine. The quantum chemical computations revealed that the reaction rate well corresponds with the Hirshfeld and NBO atomic charges on the α-carbon atom. Because of the strong hydration of OPA in aqueous solutions, the reaction was performed both in aqueous solutions (pH 9–10) as well as in diethyl ether. For the identification of intermediates and products, NMR, MS and X-ray crystal structure analyses have been used. Besides the expected isoindolin-1-one main products, the phthalimide derivative as a side product was observed and a new type of the conjugate product has been found and identified. The plausible reaction pathway was proposed.

아임계수를 이용한 분리대두단백질의 가수분해

최근 아임계수 가수분해는 전통적인 단백질 가수분해법의 대체방법으로서 관심을 받고 있으며, 고단백질원으로부터 아미노산을 회수하는데 효과적인 공정이 될 수 있을 것으로 기대되고 있다. 본 연구에서는 대표적인 식물성 단백질원인 분리대두단백질을 선택하여 대두단백질의 아임계수 가수분해에서 가장 중요한 인자인 대두단백질의 초기농도, 반응온도, 반응시간의 영향을 연구하여 대두단백질로부터 아미노산을 생산할 수 있는 조건을 최적화하고자 하였다. 대두단백질 수열분해액의 실온에서 pH는 <TEX>$200^{\circ}C$</TEX>에서 20분간 처리했을 때 pH는 7.3으로 중성이었으나 온도가 증가할수록 pH가 증가하여 <TEX>$270^{\circ}C$</TEX>에서 10.3으로 알카리성을 나타내었다. 반응온도 <TEX>$220^{\circ}C$</TEX>까지는 수열분해액은 분산상태를 이루었으나 <TEX>$230^{\circ}C$</TEX> 이상에서는 분산상태가 파괴되고 단백질이 분리되어 하부에 침강층을 이루었으며 <TEX>$240^{\circ}C$</TEX> 이상에서는 이 단백질층이 현저히 감소하였다. 반응온도는 <TEX>$250^{\circ}C$</TEX>, 반응시간 20분으로 고정한 조건에서 대두현탁액의 초기농도가 수열분해에 미치는 영향을 살펴본 결과 초기농도 10% (w/v)일 때 가수분해도 및 아미노산 수율이 각각 16.2% 및 22.3%로 가장 우수하였다. 또한 반응온도 <TEX>$200-220^{\circ}C$</TEX> 범위에서는 가수분해도와 아미노산 수율은 서서히 증가하였으나 <TEX>$230-250^{\circ}C$</TEX> 영역에서는 급격히 증가하였으며 <TEX>$250^{\circ}C$</TEX> 이상에서는 다시 완만히 증가하는 경향을 보여 <TEX>$250^{\circ}C$</TEX> 이상에서 아미노산 분해속도는 단백질 분해속도를 초과하였다. 표면반응분석법으로 예측한 결과 <TEX>$268^{\circ}C$</TEX>, 처리시간 35분에서 최적 아미노산 수율 43.5%를 얻을 수 있었다. Hydrolysis of isolate soybean protein (ISP) using subcritical water (SCW) was conducted to study the feasibility for producing protein hydrolyzate. SCW hydrolysis of SPI suspension (5-15%) was conducted in an electrically heated batch reactor (2 L). The effects of temperature (230 to <TEX>$270^{\circ}C$</TEX>) and holding time (10 to 50 min) on the degree of hydrolysis (DH) and the production of amino acids were studied by surface response method. The DH was determined by derivatizing the hydrolyzates with ortho-phthalaldehyde (OPA) solution. It was confirmed that reaction temperature and holding time affected the hydrothermolysis of soybean protein. However, the holding time was less effective on amino acid yield when the temperature was higher than <TEX>$230^{\circ}C$</TEX>. In order to achieve optimal yields of amino acids exceeding 43%, the temperature should be within the range between 256 and <TEX>$268^{\circ}C$</TEX> with holding time from 29 to 41 min, respectively. A maximum estimated amino acid yield of 43.5% was obtained at <TEX>$268^{\circ}C$</TEX> for 35 min.

Electrochemical and Spectrophotometric Study of the Hydration of Orthophthalaldehyde and Its Reaction with Amines and Aminoacids

Reactions of orthophthalaldehyde (OPA) with aliphatic α-amino acids and with aliphatic primary amines were studied in buffered aqueous solution and in non-aqueous acetonitrile. For comparison lysine and glycine ethyl ester were used. The role of the substitution on the α-carbon was discovered and the effect of OPA hydration on the course of the reaction was also investigated. In contrary to the published information, the dialdehydic unhydrated form is not the primary reactive species and for the successful reaction of OPA with amines, the presence of water is necessary.

Durability comparison and cost-effectiveness analysis of three disinfectants

Objective To compare the durability and cost-effectiveness of slightly acidic electrolyzed oxidizing water(SAEOW), ortho-phthalaldehyde(OPA) and glutaraldehyde(GA) in two conditions, i.e., continuous machine cleaning and continuous manual cleaning. Methods The maximal numbers of endoscopes which were disinfected by the three disinfectants at its minimal effective concentration, as well as the corresponding costs were compared. The cost-effectiveness of three disinfectants were evaluated with revenue-incremental cost analysis. Results The number of endoscopes disinfected by ortho-phthalaldehyde was larger than that by two other disinfectants, 140±2 by machine wash and 226±1 by hand washing; followed by glutaral disinfectant, 88±2 and 108±2 by machine wash and hand wash, respectively; slightly acidic electrolyzed oxidizing water disinfected the least endoscopes, 34±2 and 39±1 by machine wash and hand wash. There were significant differences among these three disinfectants (all P<0.05). Compared with GA, one more Yuan invested to OPA yielded 2.24 yuan more under the condition of machine wash and 3.14 yuan more under the condition of hand wash. Compared with GA, one Yuan invested in SAEOW produced 47.14 yuan by machine wash and 45.6 Yuan by hand wash. Conclusion Under full workload, ortho-phthalaldehyde shows the best durability among the three disinfectants, while slightly acidic electrolyzed oxidizing water shows the highest economic benefit. Key words: Slightly acidic electrolyzed oxidizing water; Ortho-phthalaldehyde; Glutaral; Disinfection; Economics

Electrochemical and Spectrophotometric Study of the Hydration of Orthophthalaldehyde and Its Reaction with Amines and Aminoacids

Reactions of orthophthalaldehyde (OPA) with nucleophiles are used in analytical and biologically significant reactions for many years [1, 2]. Besides the analytical method for determination of amino acids, the solution of OPA is broadly applied in hospitals as a disinfecting agent for surgery instruments made from plastics. In the literature, there are several different protocols concerning the composition of the active solutions with different timing of the use. This situation is a consequence of the fact, that the mechanisms are not understood [3] and therefore the processes could not be optimized. OPA as a dialdehyde is a molecule with two communicating redox centers. The ortho-position of the two carbonyls enable intramolecular cyclization of intermediates, hence its reactivity with nucleophiles offers a number of structural and stoichiometric possibilities. In addition to this, hydration of the original structure plays a crucial role, since H2O is, in fact, the most abundant nucleophile in the system. Nevertheless, for the above mentioned use, the reaction of OPA with amines and particularly with amino acids is the most important. Due to the reducibility of carbonyl groups and their nitrogen analogues, the electrochemical approach is favorable for this mechanistic investigation. Besides the hydration(dehydration studies, the first series of experiments were focused on the reaction of OPA with aliphatic amino acids: eight simple amino acids with only one α-amino group (glycine, alanine, leucine, isoleucine, valine, norvaline, aminobutyric and aminoisobutyric acid), one ester preventing zwitterionic equilibrium (glycine ethyl ester) and lysine with two amino groups. Polarography and spectrometry were the main used techniques. Measurements were done in two phosphate buffers (pH 7,90 and 11,20) with pH above and under the pKa of amino groups in used amino acids. It was found out that the hydration of orthophthalaldehyde plays a very important role. If stock solution of OPA is prepared in acetonitrile, then in aqueous phosphate buffer the hydration of OPA competes with the reaction with amino acids more than it was expected in published articles [4]. If stock solution of OPA is prepared in water, the reaction with amino acids is faster than with non-aqueous stock solution. The main result is that according to our measurements the reactive species is not OPA as dialdehydic form but its (mono)hydrated form. At present the investigation continues with various aliphatic amino compounds, the relationship with hydration of OPA and the reaction products are presented and discussed. Acknowledgments The research was supported by GA CR (project No. 13-21704S) and by institutional support (RVO: 61388955).

Synergistic sporicidal effect of ethanol on a combination of orthophthalaldehyde and didecyldimethylammonium chloride.

The objective of this study was to evaluate the potential synergistic effect of ethanol on a combination of orthophthalaldehyde (OPA) and didecyldimethylammonium chloride (DDAC) against the spores of Bacillus subtilis var. Niger. The quantitative carrier test for sporicidal testing of high-level disinfectants according to the guideline of China (Technical Standard for Disinfection 2002) was used as method. Considerable synergistic effect was observed after a 30-min treatment at 20°C. There was an augment in mean log reduction as the concentration of DDAC was increased ranging from 0·2 to 3gl(-1) in combination with 6gl(-1) OPA. Ten and 20% ethanol in combination with 6gl(-1) OPA and 2gl(-1) DDAC caused more than a 3-log reduction while either 6gl(-1) OPA, 2gl(-1) DDAC and 20% ethanol alone or a combination of two of the three agents produced less than a 1-log reduction. Further, 40-min exposure time of combination of OPA, DDAC and 20% ethanol led to greater than a 5-log reduction in spores, and no spore growth was observed following 60- and 90-min exposures. Orthophthalaldehyde (OPA) is very effective at concentrations far lower than its recommended in-use concentration of 0·5% (w/v) and is equally effective against both the gram-negative and gram-positive bacteria. However, it shows lower activity against spores. The synergistic sporicidal effect exhibited by ethanol on a combination of OPA and DDAC can be considered to enhance sporicidal activity for using in situations of sterilization, to reduce in-use concentration of OPA used alone, which may minimize its side effect. OPA may be a more satisfactory and the first-choice agent to replace glutaraldehyde (GTA) as a high-level disinfectant for medical devices.

Development, evaluation and comparison of two independent sampling and analytical methods for ortho-phthalaldehyde vapors and condensation aerosols in air.

Two independent sampling and analytical methods for ortho-phthalaldehyde (OPA) in air have been developed, evaluated and compared (1) a reagent-coated solid sorbent HPLC-UV method and (2) an impinger-fluorescence method. In the first method, air sampling is conducted at 1.0 L min-1 with a sampler containing 350 mg of silica gel coated with 1 mg of acidified 2,4-dinitrophenylhydrazine (DNPH). After sampling, excess DNPH in ethyl acetate is added to the sampler prior to storage for 68 hours. The OPA-DNPH derivative is eluted with 4.0 mL of dimethyl sulfoxide (DMSO) for measurement by HPLC with a UV detector set at 3S5 nm. The estimated detection limit is 0.016 µg per sample or 0.067 µg m-3 (0.012 ppb) for a 240 L air sample. Recoveries of vapor spikes at levels of 1.2 to 6.2 µg were 96 to 101%. Recoveries of spikes as mixtures of vapor and condensation aerosols were 97 to 100%. In the second method, air sampling is conducted at 1.0 L mm-1 with a midget impinger containing 10 mL of DMSO solution containing N-acetyl-l-cysteine and ethylenediamine. The fluorescence reading is taken 80 min after the completion of air sampling. Since the time of taking the fluorescence reading is critical, the reading is taken with a portable fluorometer. The estimated detection limit is 0.024 µg per sample or 0.1 µg m-3 (0.018 ppb) for a 240 L air sample. Recoveries of OPA vapor spikes at levels of 1.4 to 5.0 µg per sample were 97 to 105%. Recoveries of spikes as mixtures of vapors and condensation aerosols were 95 to 99%. The collection efficiency for a mixture of vapor and condensation aerosol was 99.4%. The two methods were compared side-by-side in a generation system constructed for producing controlled atmospheres of OPA vapor in air. Average air concentrations of OPA vapor found by both methods agreed within ±10%.

Action of a pancreatin and an Aspergillus oryzae protease on whey proteins: correlation among the methods of analysis of the enzymatic hydrolysates

The objectives of this study were to hydrolyze whey proteins using a pancreatin and an Aspergillus oryzae protease; to evaluate the degree of hydrolysis (DH) and the peptide profile; and to establish the correlations among the analytical methods. Ten hydrolysates were prepared at different reaction times and the highest DH was obtained by the protein content method. Good correlations (r > 0.87) between the methods of formaldehyde and orthophthalaldehyde (OPA), formaldehyde and osmometry as well as osmometry and OPA were observed using pancreatin. Similar results were obtained between OPA and soluble protein content for the A. oryzae protease. The action of pancreatin produced the highest contents of di- and tripeptides (9.07, 7.12 and 6.46%) and the lowest of large peptides (42.43, 41.33 and 41.13%), after 3, 4 and 5 h of hydrolysis, respectively. Using pancreatin, the DH measured by formol titration and OPA was positively correlated with medium peptide content and negatively correlated with large peptide content. For the A. oryzae protease, a strong negative correlation was observed between the large peptide content and the DH measured by the OPA method.

PIN2 - Alternative High Level Disinfectants to Processing Flexible Endoscopes

Endoscopy is fundamental in different medical specialties, with increasing use. Endoscopes are expensive equipment, complex, thermo sensitive, delicate materials and submitted to high-level disinfection (HLD). The most commonly used disinfectant is glutaraldehyde (GLU) due to its high compatibility with the materials and low cost. However, evidence of toxicity to professionals and the identification of mycobacteria tolerant to GLU caused changes in Brazilian legislation on the issue, pressing health services to search for alternative desinfectants. Search for evidence on the effectiveness, toxicity and potential damage to endoscopes by alternative disinfectants to GLU available in the Brazilian market. The study sample was semi-critical endoscopes flexible (digestive, respiratory and cystoscope), the intervention was HLD with peracetic acid (PA), Ortho-Phthalaldehyde (OP) and Electrolyzed Acid Water (EAW), compared to GLU, with outcomes HLD effectiveness, toxicity and damage to equipment. Were identified 822 publications (2008-2013) on 13 databases, 23 studies were selected considering the best quality of available evidence. As for effectiveness, the findings draw attention to the intrinsic resistance of subclass Coccidia spp to all evaluated disinfectants and acquired tolerance by GLU strain of Mycobacterium massilienses which caused an outbreak of infection with more than 2000 cases in Brazil. About toxicity, the most frequently reported adverse events are colitis (no definitive causal relationship with the germicide used) and anaphylactic reactions by OP in cystoscopy. There is lack of published data on damage caused in endoscopes by the disinfectants; the few studies on the theme indicate the importance of adequate handling of equipment for conservation of its functionality. The publications show superiority of the PA and OP for efficacy in HLD. Only the OP clearly had adverse event related to their use. There is insufficient evidence in literature to assert the inferiority of some disinfectant for damage to equipment.

Development and validation of a spectrofluorimetric method for the quantification of ceftriaxone in pharmaceutical formulations and plasma

We describe the development and validation of a new, simple, sensitive and cost-effective method for the determination of ceftriaxone in commercial formulations and spiked human plasma. The method proposes the conversion of ceftriaxone into a fluorescent product by reacting with ortho-phthalaldehyde (OPA) in the presence of sulfite at room temperature. The reaction medium is buffered to pH 10 using borate buffer. The derivatized reaction product is highly fluorescent and exhibits maximum fluorescence intensity at λ(em) = 386 nm after excitation at λ(ex) = 324 nm. The experimental parameters affecting progress of the derivatization reaction were carefully studied and optimized. Under optimum experimental conditions, the method has an excellent correlation coefficient of 0.9984 with a broad linear range of 0.4-20 µg/mL. The limit of detection (LOD) and limit of quantification (LOQ) were found to be 1.30 × 10(-3) and 3.90 × 10(-3) µg/mL, respectively. The interference effects of common excipients on the quantification of drug were investigated and no interference effect was observed. The proposed method has been successfully applied to the determination of ceftriaxone in pharmaceutical formulations and spiked human plasma samples. The method has been validated statistically through percent recovery studies using standard addition and by comparison with a reference HPLC method. The developed method exhibits excellent inter- and intraday precision.

Sensitive determination of hydrazine in water by gas chromatography–mass spectrometry after derivatization with ortho-phthalaldehyde

A gas chromatography–mass spectrometric (GC–MS) method has been established for the determination of hydrazine in drinking water and surface water. This method is based on the derivatization of hydrazine with ortho-phthalaldehyde (OPA) in water. The following optimum reaction conditions were established: reagent dosage, 40mgmL−1 of OPA; pH 2; reaction for 20min at 70°C. The organic derivative was extracted with methylene chloride and then measured by GC–MS. Under the established condition, the detection and the quantification limits were 0.002μgL−1 and 0.007μgL−1 by using 5.0-mL of surface water or drinking water, respectively. The calibration curve showed good linearity with r2=0.9991 (for working range of 0.05–100μgL−1) and the accuracy was in a range of 95–106%, and the precision of the assay was less than 13% in water. Hydrazine was detected in a concentration range of 0.05–0.14μgL−1 in 2 samples of 10 raw drinking water samples and in a concentration range of 0.09–0.55μgL−1 in 4 samples of 10 treated drinking water samples.