Assessment of nationally recommended antibiotics for treatment of UTI in U.S.-Mexico border emergency departments.

BACKGROUND: Urinary tract infections (UTIs) seen in the emergency department are commonly treated as an outpatient with oral antibiotics. Given that antibiotics are available for over-the-counter purchase in Mexico, there is speculation that potential misuse and overuse of antibiotics in United States-Mexico border areas could lead to antibiotic resistance patterns that would render some empiric treatments for UTIs less effective. The purpose of this study was to examine the effectiveness of Infectious Disease Society of America (IDSA) guideline-recommended antibiotics for treatment of outpatient UTI diagnosed in the emergency department. Data were collected from a county hospital on the U.S.-Mexico border with a metropolitan area of over 2 million people. Secondary analysis included frequency of urine culture isolated, resistance rates of urine pathogens, and prescriber habits. METHODS: This study was a retrospective chart review of adult patients diagnosed and treated for UTI from August 1, 2019, to February 29, 2020. Culture results of included patients were analyzed against in vitro-tested antibiotics. Bacterial isolate frequency, resistance rates, and prescribing habits were collected. RESULTS: A total of 985 patient charts were reviewed, of which 520 patients met inclusion criteria for analysis of prescribing habits. Of these, 329 positive bacterial culture growths were included in the analysis of antibiotic resistance rates. Oral antibiotics with comparatively lower resistance rates were amoxicillin/clavulanate, cefdinir, cefuroxime, and nitrofurantoin. Oral antibiotics with notably high resistance rates included trimethoprim-sulfamethoxazole (TMP-SMX), tetracycline, ciprofloxacin, levofloxacin, and cephalexin. Nitrofurantoin was prescribed most frequently for outpatient treatment of UTI/cystitis (41.6%) while cephalexin was the most commonly prescribed antibiotic for outpatient treatment of pyelonephritis (50%). CONCLUSION: Our findings suggest that, while part of standard IDSA guidelines, fluoroquinolones and TMP-SMX are not ideal empiric antibiotics for treatment of outpatient UTI in the U.S.-Mexico border region studied due to high resistance rates. Although not listed as first line agents per current IDSA recommendations, 2nd and 3rd generation cephalosporins, and amoxicillin/clavulanate would be acceptable options given resistance patterns demonstrated in accordance with IDSA allowance for tailoring selection to local resistance. Nitrofurantoin appears to be consistent with recommendations and demonstrates a favorable resistance profile for treatment of outpatient UTI within this region.

Probability of target attainment of oral antimicrobials for Escherichia coli and Klebsiella pneumoniae based on Monte Carlo simulations.

Although early transition from intravenous to oral antimicrobials can reduce hospitalization duration, susceptibility breakpoints have not been established for many oral antimicrobials against Escherichia coli and Klebsiella pneumoniae bacteremia. Thus, we used population pharmacokinetic models, pharmacokinetic/pharmacodynamic indices, and Monte Carlo simulations to evaluate the probability of target attainment (PTA) for common oral antimicrobial dosages against E. coli and K. pneumoniae. The oral antimicrobial agents evaluated included cephalexin, cefaclor, cefditoren, amoxicillin/clavulanic acid, faropenem, and levofloxacin. For E. coli, the percentage of isolates with minimum inhibitory concentrations for which a PTA >90% was achieved was 53% and less than 20% for levofloxacin and the ß-lactams, respectively. For K. pneumoniae, the percentages of isolates for which a PTA >90% was achieved were comparatively higher (cephalexin, 73%; amoxicillin/clavulanic acid, 83%; levofloxacin, 96%). Our results suggest clinicians should check if pharmacokinetic/pharmacodynamic indices are achieved in individual patients before transitioning to oral antimicrobial therapy.

Photocatalyst production from wasted sediment and quality improvement with titanium dioxide to remove cephalexin in the presence of hydrogen peroxide and ultrasonic waves: A cost-effective technique.

In this study, wasted sediment (sludge waste from shipping docks) was coupled with titanium isopropoxide by the thermal and sol-gel method as a new photocatalyst. The sediment-titanate catalyst alongside ultrasonic and UV was activated hydrogen peroxide to produce OH radicals and decompose cephalexin (CEP). The photocatalyst was crystalline with 52.29 m2/g BET area. The best destruction rate of 87.01% based on COD test was achieved at optimal conditions (pH: 8, cephalexin concentration: 100 mg/L, H2O2: 1.63 mg/L, UV: 15 W/m2, ultrasonication time: 100 min at 40 kHz, photocatalyst quantity: 1.5 g/L). The trend of anions effect was NO3- ≤ SO42- ≤ Cl-. Decomposition of cephalexin in water solution followed the first-order kinetics (k > 0.01 min-1, R2 > 0.9). The percentage of cephalexin removal from urban water (76%) and hospital wastewater (63%) has decreased compared to the distilled water solution (87%), which is probably due to the presence of radical inhibitors. The consumed electrical energy of the studied system was calculated by 0.031 kW/h. The developed system is a promising and economical method to remove cephalexin.

Degradation of micropollutant cephalexin by ultraviolet (UV) and assessment of residual antimicrobial activity of transformation products.

Cephalexin (CEX) is an antibiotic commonly used to treat bacterial infections in humans and animals. However, it is also a micropollutant. Thus, this study evaluated the degradation of CEX using ultraviolet irradiation (UV-C) and analyzed the by-products as well as their residual antimicrobial activity. A reactor with a mercury vapor lamp was used for the degradation. Irradiated CEX solutions were collected over a period of 4 hours and analyzed using high-performance liquid chromatography coupled with mass spectrometry. For the residual antimicrobial activity the susceptibility test was performed using Staphylococcus aureus and Escherichia coli microorganisms by broth microdilution. It was found that CEX, after treatment, generated a metabolite with a mass of 150 m/z in 15 min. A four- and eightfold increase in the minimum inhibitory concentration of the drug against S. aureus and E. coli could be observed, respectively, after 20 min. Therefore, this treatment proved to be effective in the degradation of CEX, being able to degrade 81% of the initial molecule of the drug in 20 min. Furthermore, the antimicrobial activity of the CEX solution decreased as the irradiation time increased, indicating loss of antimicrobial function of the initial CEX molecule and the resulting by-products.

Comparación de antibióticoterapia en la apendicitis complicada en pediatría / Comparison of antibiotic therapy in complicated appendicitis in pediatrics

INTRODUCCIÓN. La apendicitis aguda es una patología pediátrica quirúrgica que en su fase complicada requiere uso de antibióticos en el postoperatorio; encontrar la mejor opción como experiencia local, fue necesario. OBJETIVO. Comparar un esquema de antibioticoterapia triple vs un doble respecto a variables: demográficas, clínicas y de costos en pacientes pediátricos con apendicitis complicada. MATERIALES y MÉTODOS. Estudio analítico transversal. Población de 133 pacientes y una muestra de 93, operados por apendicitis complicada; 58 recibieron ampicilina + metronidazol + gentamicina y 35 ceftriaxona + metronidazol. Se comparó las variables: estadía hospitalaria, complicaciones y costo monetario de cada esquema. Se realizó en la Unidad de Cirugía Pediátrica del Hospital de Especialidades Carlos Andrade Marín, en el periodo enero de 2017 a octubre de 2018. Los datos fueron analizados con R-Studio 1.8.366 para Windows. RESULTADOS. No hubo diferencia estadística con respecto a: estadía hospitalaria (p=0,261); complicaciones como infección del sitio quirúrgico (p=0,196), re-intervención quirúrgica (p=0,653) y costo (p=0,059). CONCLUSIÓN. Se comparó el esquema de antibioticoterapia triple vs un doble, utilizados en apendicitis complicada en pediatría. No se encontró diferencias estadísticamente significativas en este reporte preliminar, con la diferencia de que con el esquema doble la frecuencia de administración fue menor y se evitó la exposición a los efectos colaterales de los aminoglicósidos.

INTRODUCTION. Acute appendicitis is a pediatric surgical pathology that in its complicated phase requires the use of antibiotics during the postoperative period; finding the best option as local experience was a must. OBJECTIVE. Compare a triple vs a double antibiotic therapy scheme respect demographic, clinical and cost variables in pediatric patients whit complicated apendicitis. MATERIALS AND METHODS. Cross-sectional analytical study. Population of 133 patients and sample of 93, with complicated appendicitis; 58 received ampicillin + metronidazole + gentamicin and 35 ceftriaxone + metronidazole. The following variables were compared; hospital stay, complications and monetary cost of each scheme. It was performed in the Pediatric Surgery Unit of the Hospital of Specialties Carlos Andrade Marin, during january 2017, through october 2018. The data were analyzed with R-Studio 1.8.366 for Windows. RESULTS. There was no statistical difference regarding hospital stay (p=0.261); complications such as surgical site infection (p=0.196), re-surgical intervention (p=0.653); nor cost (p=0.059). CONCLUSION. Triple vs. Double antibiotic therapy scheme used in complicated appendicitis in pediatrics was compared. No statistically significant differences were found in this preliminary report, how ever with the double scheme the frequency of administration was lower and exposure to side effects of aminoglycosides was avoided.

Emergency department medication dispensing reduces return visits and admissions.

OBJECTIVES: Return visits to the emergency department (ED) and subsequent readmissions are common for patients who are unable to fill their prescriptions. We sought to determine if dispensing medications to patients in an ED was a cost-effective way to decrease return ED visits and hospital admissions for skin and soft tissue infections (SSTIs). METHODS: A retrospective review of ED visits for SSTIs, during the 24 weeks before and after the implementation of a medication dispensing program, was conducted. Charts were analyzed for both ED return visits and hospital admissions within 7 days and 30 days of the initial ED visit. Return visits were further reviewed to determine if the clinical conditions on subsequent visits were related to the initial ED presentation. A cost analysis comparing the cost of treatment to cost savings for return visits was also performed. RESULTS: Before the implementation of the medication dispensing program, the return rate in 7 days for the same condition was 9.1% and the rate of admission was 2.8%. The return rate for the same condition in 8-30 days was 2.1% and the rate of admission was 1.0%. After the implementation of the medication dispensing program, the return rate for the same condition in 7 days was 8.0%, and the admission rate was 1.7%. The return rate for the same condition in 8-30 days was 0.8%, and the admission rate was 0%. The total cost of dispensed medications was $4050, while total cost savings were estimated to be $95,477. CONCLUSION: A medication dispensing program in the ED led to a reduction in return visits and admissions for SSTIs at both 7 days and 30 days. For a cost of only $4050, an estimated total of $95,477 was saved. A medication dispensing program is a cost-effective way to reduce return visits to the ED and subsequent admissions for certain conditions.

Collaboration with an infection control team promoted appropriate antibiotic use for third molar extraction at a Japanese hospital.

Oral antibiotic therapy is routinely administered when a third molar (M3) is extracted to prevent infectious complications after surgery. Oral third-generation cephalosporins are frequently used after M3 extraction in Japan but at the expense of an increased risk of antimicrobial resistance. Therefore, the infection control team (ICT) at our institution recommended a reduction in use of these agents after M3 extraction. In this study, we compared the types of antibiotic agents prescribed for patients undergoing M3 extraction before and after this recommendation. We investigated the relationship between type of antibiotic used and the likelihood of infectious complications as well as cost savings in patients who underwent M3 extraction in the 6 months before and after the ICT recommendation in July 2018. There was a marked reduction in use of oral third-generation cephalosporins after M3 extraction (P < 0.0001) and increased use of oral penicillins and first-generation cephalosporins after the ICT recommendation. Moreover, surgical site infection (SSIs) were significantly less common after the ICT recommendation (P = 0.0099); however, the SSI rate was higher in patients who received a third-generation cephalosporin than in those who received penicillin (8.8% vs 0.5%). There was also a significant saving in per-patient antibiotic costs after the ICT recommendation (269.5 ± 282.0 JPY vs 454.7 ± 376.6 JPY; P < 0.0001). These findings suggest that collaboration with an ICT promotes appropriate antibiotic use, decreases the risk of an SSI, and improves the cost-benefit ratio in patients undergoing M3 extraction.

Assessment of the comparability of CLSI, EUCAST and Stokes antimicrobial susceptibility profiles for Escherichia coli uropathogenic isolates.

BACKGROUND: As many clinical laboratories convert between Stokes, Clinical and Laboratory Standards Institute (CLSI) and European Committee for Antimicrobial Susceptibility Testing (EUCAST) methods, the problem of comparing differently derived sets of antimicrobial susceptibility testing (AST) data with each other arises, owing to a scarcity of knowledge of inter-method comparability. The purpose of the current study was to determine the comparability of CLSI, EUCAST and Stokes AST methods for determining susceptibility of uropathogenic Escherichia coli to ampicillin, amoxicillin-clavulanate, trimethoprim, cephradine/cephalexin, ciprofloxacin and nitrofurantoin. METHODS: A total of 100 E. coli isolates were obtained from boric acid urine samples from patients attending GP surgeries. For EUCAST and CLSI, the Kirby-Bauer disc diffusion method was used and results interpreted using the respective breakpoint guidelines. For the Stokes method, direct susceptibility testing was performed on the urine samples. RESULTS: The lowest levels of agreement were for amoxicillin-clavulanate (60%) and ciprofloxacin (89%) between the three AST methods, when using 2017 interpretive guidelines for CLSI and EUCAST. A comparison of EUCAST and CLSI without Stokes showed 82% agreement for amoxicillin-clavulanate and 94% agreement for ciprofloxacin. Discrepancies were compounded by varying breakpoint susceptibility guidelines issued during the period 2011-2017, and through the inclusion of a definition of intermediate susceptibility in some cases. CONCLUSIONS: Our data indicate that the discrepancies generated through using different AST methods and different interpretive guidelines may result in confusion and inaccuracy when prescribing treatment for urinary tract infection.

Prevalence and correlates of antibiotic sharing in the Philippines: antibiotic misconceptions and community-level access to non-medical sources of antibiotics.

OBJECTIVE: To identify sociodemographic, knowledge and attitudinal correlates to antibiotic sharing among a community-based sample of adults (age 18 and older) in a low-income setting of the Philippines and to explore community-level data on informal antibiotic distribution in roadside stands (i.e., sari-sari stands). METHODS: Participants (n = 307) completed self-administered surveys. Correlates to antibiotic sharing were assessed using logistic regression with Firth's bias-adjusted estimates. Study staff also visited 106 roadside stands and collected data on availability and characteristics of antibiotics in the stands. RESULTS: 78% had shared antibiotics in their lifetime, most often with family members. In multivariable analysis, agreement with the belief that it is safe to prematurely stop an antibiotic course (OR: 2.8, CI: 1.3-5.8) and concerns about antibiotic side effects (OR: 2.1, CI: 1.1-4.4) were significantly associated with increased odds of reported antibiotic sharing. Antibiotic sharing was not associated with sociodemographic characteristics or antibiotic knowledge. Antibiotics were widely available in 60% of sampled sari-sari stands, in which 59% of antibiotics were missing expiration dates. Amoxicillin and cephalexin were the most commonly available antibiotics for sale at the stands (60% and 21%, respectively). CONCLUSIONS: Antibiotic sharing was common and was associated with misconceptions about proper antibiotic use. Antibiotics were widely available in sari-sari stands, and usually without expiration information. This study suggests that multipronged and locally tailored approaches to curbing informal antibiotic access are needed in the Philippines and similar Southeast-Asian countries.

A model assessment of the importance of direct photolysis in the photo-fate of cephalosporins in surface waters: Possible formation of toxic intermediates.

The direct and indirect photodegradation of six cephalosporins was predicted using a photochemical model, on the basis of literature values of photochemical reactivity. Environmental photodegradation would be important in surface water bodies with depth ⩽ 2-3m, and/or in deeper waters with low values of the dissolved organic carbon (DOC ⩽ 1 mg C L(-1)). The half-life times would range from a few days to a couple of weeks in summertime. In deeper and higher-DOC waters and/or in different seasons, hydrolysis could prevail over photodegradation. The direct photolysis of cephalosporins is environmentally concerning because it is known to produce toxic intermediates. It would be a major pathway for cefazolin, an important one for amoxicillin and cefotaxime and, at pH<6.5, for cefapirin as well. In contrast, direct photolysis would be negligible for cefradine and cefalexin. The DOC values would influence the fraction of photodegradation accounted for by direct photolysis in shallow water, to a different extent depending on the role of sensitisation by the triplet states of chromophoric dissolved organic matter.

Outcome assessment in cellulitis clinical trials: is telephone follow up sufficient?

The US Food and Drug Administration has scrutinized clinical trial methodology in cellulitis, partly because the definition and timing of cure are debatable. We analysed the validity of telephone self-report as a proxy for in-person follow up in a cellulitis treatment trial comparing cephalexin alone with cephalexin-plus-trimethoprim/sulfamethoxazole. Our results demonstrate poor agreement between these two methods of outcome determination and have implications for future cellulitis clinical trial design and clinical management.

Quantitative analysis of cefalexin based on artificial neural networks combined with modified genetic algorithm using short near-infrared spectroscopy.

In this paper, a novel chemometric method was developed for rapid, accurate, and quantitative analysis of cefalexin in samples. The experiments were carried out by using the short near-infrared spectroscopy coupled with artificial neural networks. In order to enhancing the predictive ability of artificial neural networks model, a modified genetic algorithm was used to select fixed number of wavelength.

Distribution, fate and risk assessment of antibiotics in sewage treatment plants in Hong Kong, South China.

Occurrence, removal, consumption and environmental risks of sixteen antibiotics were investigated in several sewage treatment plants (STPs) featuring different treatment levels in Hong Kong, China. Cefalexin, ofloxacin and erythromycin-H(2)O were predominant with concentrations of 1020-5640, 142-7900 and 243-4740 ng/L in influent, respectively; their mass loads were comparable to levels reported in urban regions in China and were at the high end of the range reported for western countries. The target antibiotics behaved differently depending on the treatment level employed at the STPs and relatively higher removal efficiencies (>70%) were observed for cefalexin, cefotaxime, amoxicillin, sulfamethoxazole and chloramphenicol during secondary treatment. ß-lactams were especially susceptible to removal via the activated sludge process while macrolides were recalcitrant (<20%) in the dissolved phase. Two fluoroquinolones, ofloxacin (4%) and norfloxacin (52%), differed greatly in their removal efficiencies, probably because of disparities in their pK(a) values which resulted in different sorption behaviour in sludge. Overall antibiotic consumption in Hong Kong was back-calculated based on influent mass flows and compared with available prescription and usage data. This model was verified by a good approximation of 82% and 141% to the predicted consumption of total ofloxacin, but a less accurate estimate was obtained for erythromycin usage. Risk assessment indicated that algae are susceptible to the environmental concentrations of amoxicillin as well as the mixture of the nine detected antibiotics in receiving surface waters.

Effect of loading rate and HRT on the removal of cephalosporin and their intermediates during the operation of a membrane bioreactor treating pharmaceutical wastewater.

The viability of treating high-concentration antibiotic wastewater by an anaerobic membrane bioreactor was studied using submerged flat sheet membrane. The objective of the study was to determine the effect of organic loading rate and hydraulic retention time on the removal of cephalosporin derivative, viz. cephalexin, and the intermediates 7-amino-3-deacetoxycephalosporanic acid (7-ADCA) and acyl group (phenyl acetic acid) in an anaerobic membrane bioreactor with enhanced biodegradation using the bioaugmentation technique. The pharmaceutical industry is looking for alternatives to either direct disposal of 7-amino-3-deacetoxycephalosporanic acid and phenyl acetic acid, or further degradation and disposal, which will essentially require additional costs and maintenance. The present regulatory standard, implemented at a global level, does not allow for such disposal alternatives and hence the present study was aimed at the complete removal of the intermediates 7-ADCA and phenyl acetic acid prior to discharge.

Cephalexin residue detection in milk and beef by ELISA and colloidal gold based one-step strip assay.

An evaluation of a rapid enzyme-linked immunosorbent assay (ELISA) and colloidal gold based one-step strip assay for cephalexin (CEX) residue detection in milk and beef is described. A monoclonal antibody (mAb) against CEX was produced using cephalexin-bovine serum albumin (CEX-BSA) conjugate as the immunogen, which exhibited no cross-reactivity with applied chemicals in the studied concentration range. The detection limit of rapid ELISA was calculated as 0.39 microg/kg in PBS and 19.5 microg/kg in beef and milk, which was quite lower than the European Union Maximum Residue Limit (MRL) of 100 microg/kg in milk and 200 microg/kg in muscle. Spiked samples were detected with a mean recovery of 82.8-124% and coefficient of variation of 4.88-25%, which indicated a good agreement with the spiked concentration. Accuracy and reproducibility were determined using spiked samples with four different final concentrations of 1, 2, 5, and 10 microg/kg of CEX (n = 7). Mean intra-assay variation of 6.67% and inter-assay variation of 10.66% were obtained. In contrast, the strip test for CEX had a visual detection limit of 0.5 microg/kg, which could be evaluated within 3-10 min. However, positive samples should be further quantified by more sensitive and accurate competitive indirect ELISA method. In conclusion, the described strip test is rapid, simple, and cost-effective as well as sensitive and specific enough for reliable and accurate on-site screening.

Antibiotic selection for purulent skin and soft-tissue infections in ambulatory care: a decision-analytic approach.

OBJECTIVE: Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) has caused a nationwide epidemic of skin and soft-tissue infections in ambulatory pediatrics. Antibiotic treatment recommendations suggest incorporating local epidemiology for the prevalence of CA-MRSA. We sought to identify the antibiotic strategy with the highest probability of activity and to identify threshold values for epidemiologic variables including bacterial prevalence and antibiotic resistance. METHODS: We used decision analysis to evaluate 3 empiric antibiotic strategies: clindamycin, trimethoprim/sulfamethoxazole (T/S), and cephalexin. We calculated the probability of activity against the bacteria causing the infection (CA-MRSA, methicillin-sensitive S. aureus and group A Streptococcus [GAS]) by incorporating estimates of prevalence and antibiotic resistance to determine the optimal strategy. Sensitivity analysis was used to identify thresholds for prevalence and antibiotic resistance where 2 strategies were equal. RESULTS: Clindamycin (0.95) and T/S (0.89) had substantially higher probability of activity than cephalexin (0.28) using baseline estimates for bacterial prevalence and antibiotic resistance. Cephalexin was the optimal antibiotic only when CA-MRSA prevalence was <10%. The probability of activity for clindamycin and T/S was highly sensitive to changes in the values for bacterial prevalence (both CA-MRSA and GAS) and CA-MRSA resistance to clindamycin. CONCLUSIONS: Empiric treatment of skin and soft-tissue infections with either clindamycin or T/S maximizes the probability that the antibiotic will be active when CA-MRSA prevalence is >10%. Deciding between T/S and clindamycin requires consideration of antibiotic resistance and prevalence of GAS. This model can be customized to local communities and illustrates the importance of ongoing epidemiologic surveillance in primary care settings.

Management of vagal nerve stimulator infections: do they need to be removed?

OBJECT: Vagal nerve stimulators (VNSs) have been used successfully to treat medically refractory epilepsy. Although their efficacy is well established, appropriate management of infections is less clearly defined. In the authors' experience, patients who have gained a benefit from VNS implantation have been reluctant to have the device removed. The authors therefore sought conservative management options to salvage infected VNS systems. METHODS: The authors performed a retrospective review of 191 (93 female and 98 male) consecutive patients in whom VNS systems were placed between 2000 and 2007. RESULTS: They identified 10 infections (5.2%). In 9 of 10 patients the cultured organism was Staphylococcus aureus. Three (30%) of 10 patients underwent early removal (within 1 month) of the VNS as the initial treatment. The remaining 7 patients were initially treated with antibiotics. Two (28.6%) of these patients were successfully treated using antibiotics without VNS removal. Patients in whom conservative treatment failed were given cephalexin as first-line antibiotic treatment. All patients recovered completely regardless of treatment regimen. CONCLUSIONS: This study confirms the low rate of infection associated with VNS placement and suggests that, in the case of infection, treatment without removal is a viable option. However, the authors' data suggest that oral antibiotics are not the best first-line therapy.

Synthesis of cephalexin in aqueous medium with carrier-bound and carrier-free penicillin acylase biocatalysts.

The use of very high substrate concentrations favors the kinetically controlled synthesis of cephalexin with penicillin acylase (PA) not only by Michaelian considerations, but also because water activity is depressed, so reducing the rates of the competing reactions of product and acyl donor hydrolysis. Commercial PGA-450, glyoxyl agarose immobilized (PAIGA) and carrier-free cross-linked enzyme aggregates of penicillin acylase (PACLEA) were tested in aqueous media at concentrations close to the solubility of nucleophile and at previously determined enzyme to nucleophile and acid donor to nucleophile ratios. The best temperature and pH were determined for each biocatalyst based on an objective function considering conversion yield, productivity, and enzyme stability as evaluation parameters. Stability was higher with PAIGA and specific productivity higher with PACLEA, but best results based on such objective function were obtained with PGA-450. Yields were stoichiometric and productivities higher than those previously reported in organic medium, which implies significant savings in terms of costs and environmental protection. At the optimum conditions for the selected biocatalyst, operational stability was determined in sequential batch reactor operation. The experimental information gathered is being used for a technical and economic evaluation of an industrial process for enzymatic production of cephalexin in aqueous medium.

Conjugation of penicillin acylase with the reactive copolymer of N-isopropylacrylamide: a step toward a thermosensitive industrial biocatalyst.

Conjugation of penicillin acylase (PA) to poly-N-isopropylacrylamide (polyNIPAM) was studied as a way to prepare a thermosensitive biocatalyst for industrial applications to antibiotic synthesis. Condensation of PA with the copolymer of NIPAM containing active ester groups resulted in higher coupling yields of the enzyme (37%) compared to its chemical modification and copolymerization with the monomer (9% coupling yield) at the same NIPAM:enzyme weight ratio of ca. 35. A 10-fold increase of the enzyme loading on the copolymer resulted in 24% coupling yield and increased by 4-fold the specific PA activity of the conjugate. Two molecular forms of the conjugate were found by gel filtration on Sepharose CL 4B: the lower molecular weight fraction of ca. 10(6) and, presumably, cross-linked protein-polymer aggregates of MW > 10(7). Michaelis constant for 5-nitro-3-phenylacetamidobenzoic acid hydrolysis by the PA conjugate (20 microM) was found to be slightly higher than that of the free enzyme (12 microM), and evaluation of V(max) testifies to the high catalytic efficiency of the conjugated enzyme. PolyNIPAM-cross-linked PA retained its capacity to synthesize cephalexin from d-phenylglycin amide and 7-aminodeacetoxycephalosporanic acid. The synthesis-hydrolysis ratios of free and polyNIPAM-cross-linked enzyme in cephalexin synthesis were 7.46 and 7.49, respectively. Thus, diffusional limitation, which is a problem in the industrial production of beta-lactam antibiotics, can be successfully eliminated by cross-linking penicillin acylase to a smart polymer (i.e., polyNIPAM).