Laminar Air-flow Cabinet Research Articles

Ultrasonic atomization and subsequent polymer desolvation for peptide and protein microencapsulation into biodegradable polyesters

Peptide and protein microencapsulation into poly(lactide) (PLA) and poly(lactide-co-glycolide) (PLGA) microspheres continues to represent a technological challenge in terms of product sterility and up-scaling. The primary objective of this study was to examine the feasibility of a novel method for peptide and protein entrapment into PLA and PLGA microspheres, particularly suitable for up-scaling and aseptic processing. The method involves ultrasonic atomization of an organic polymer solution combined with subsequent organic solvent extraction by a hardening agent. The study evaluated the critical atomization conditions, the required molecular cohesion parameters of polymer solvents and hardening agent for particle preparation as well as the quality of entrapment and release as a function of polymer and peptide/protein type. Suitable polymer solvents and hardening agents were restricted to defined domains of fractional cohesion parameters: fp = 0.2–0.35 and fh = 0.2–0.4 for the polymer solvents, and fp = 0-0.1 and fh = 0-0.25 for the hardening agents. Microsphere size (0.1–100 μm) was largely controlled by the viscosity of the atomized solution. Microencapsulation of the freely water-soluble bovine serum albumin and tetrapeptide thymocartin yielded modest efficiencies of 12–35%, whereas the slightly water-soluble octapeptide vapreotide pamoate was entrapped with 63–93% efficiency. Drug release was mainly governed by the polymer type, lasting over 100 days for BSA entrapped in PLA microspheres and ∼ 20 days for vapreotide pamoate in PLGA 50:50 and for thymocartin in PLA. Very importantly, the novel method was readily accommodated within a laminar air-flow cabinet. Under aseptic conditions, sterile microspheres could be prepared. In conclusion, the novel method described may have potential in industrial environments.

High frequency plant regeneration of garlic (Allium sativum L.) calli immobilized in calcium alginate gel

Calli obtained from a shoot-tip of garlic,Allium sativum L., were encapsulated using a calcium alginate gel. Some of the encapsulated calli were cultured on a 1/2 MS medium supplemented with 3% sucrose, 10−5 M kinetin, and 5×10−6 M NAA, whereas the remainder was stored for 40 days at 4°C. All the naked calli regenerated on the solid medium, while 95% of the encapsulated calli regenerated, and 88% of the encapsulated calli regenerated after 40 days of storage at 4°C. The capsule matrix delayed the germination time of the encapsulated calli, yet activated the shoot formation of the artificial garlic seeds. The shoot length of the encapsulated garlic calli was much longer than that of the naked garlic calli. The encapsulated garlic calli were dried in a laminar airflow cabinet and the conversion frequency of the dried artificial garlic seeds on a 1/2 MS medium remained at 93% with a water loss of less than 50%.

In vitro propagation of Opuntia ellisiana Griff. and acclimatization to field conditions

The genus Opuntia is a valuable forage resource in arid and semiarid lands during periods of drought and shortage of herbaceous plants. However, absolute minimum temperatures in the plains of Mendoza represent a limiting factor to cultivate several species. Opuntia ellisiana is a cold hardy species, so the goals of this study were to massively propagate is using in vitro culture techniques, and then to acclimatize plantlets obtained to field conditions. Different sterilization protocols were tested. Areoles were isolated in laminar airflow cabinet, and cultured on Murashige-Skoog medium, supplemented with sucrose and different BAP and IBA combinations. Explants were grown at 27 +/- 2 degrees C, under a 16-h photoperiod. The shoots produced were used in the rooting assay using different auxin combinations. In the most efficient growth treatment, plantlets reached 100% shooting after 35 days of culture, and a mean length of 10.2 mm after 49 days of culture. A 100% rooted plantlets was obtained on a medium containing 5 mg L-1 IBA, after 12 days of culture. Acclimatization was achieved under greenhouse conditions, showing 100% plantlet survival. This study suggests that O. ellisiana can be successfully micropropagated by areoles, and easily acclimatizated to field conditions.

薬剤師による病棟での注射薬混合業務の定量的解析 （１） 骨髄移植を目的とした無菌病棟の場合

We analyzed the mixing services performed by pharmacists in the aseptic ward at Tokyo University Hospital as a model case, in order to establish a new mixing service system in clinical wards. The average number of prescriptions for injection was 5 per day during the 5 day investigation, and the number of injections prescribed and mixed were 44.6±4.4 and 33.2±4.6 per day, respectively. The most common drugs for the mixed injections were antibiotics (41.6%), antineoplastic agents and immunosuppressants agents (both together 21.6%). The ratios of the ampule/vial was 5: 5, and solution/lyophilized powder was 7: 3. When two pharmacists with 3 months and 3 years of mixing experience, respectively, mixed drugs, the required time for the three processes of preparation, mixing, and checking were approximately 40, 40, and 10 min, respectively, and the total time required was 90 min. The average required time per injection was 2.7 min (90min/33.3 injections), and the mixing time per injection in the laminar air flow cabinet was 1.2 min (40 min/33.3 injections). Regarding the mixing experiment, the average time required for 5 ampules (10 mL solution) was shorter than that for 5 vials (lyophilized powder) in all groups tested. In the 3 years-experience group, the average time was approximately 7 min/5 ampules and 13 min/5 vials, and these values were half of that for beginners. In the 3 years-experience group, the time required for ampules was almost the same as for the 3 months-experience group, and that of vials decreased to 8 min. We propose that the data from these time study can be utilized to estimate the effectiveness of the mixing services in the ward, in the hope that the fee for such important aseptic techniques in the mixing service provided by pharmacists, the same as for IVH admixturing, will be authorized by the regulatory agency in the future.

Two Tropical Plants as Plant Materials for Teaching Seed Germination

A local ground orchid, Spathoglottis plicata Blume, and coconut, Cocos nucifera L., were used in the classroom to teach seed germination. S. plicata, a common orchid on Guam, was utilized to demonstrate the aseptic culture of seeds under non-sterile conditions. The procedures were done in the classroom without a laminar air-flow cabinet. Nonsterile seeds were sown on growing media which were prepared without autoclaving, but by incorporating sodium hypochlorite into the media. Students had a high rate of success in germinating the orchid seeds without contamination by spraying sodium hypochlorite on the seeds. Different stages of coconut seed development were presented to students by simply cutting coconut in half. Unique features and botanical terms of coconut seed development can be taught throughout the year. Teaching materials on seed germination of the two tropical plants are being developed by print-on-demand methods.

Influence of background air on microbial contamination during simulated i.v.-admixture preparation.

The effect of the cleanliness of environmental air on the microbial contamination of a simulated i.v.-admixture during its preparation by aseptic transfer was studied under three conditions: (i) in a laminar air flow (LAF) bench situated in a class 1000 clean room, (ii) in an LAF bench in a microbiology laboratory and (iii) on a bench in an ordinary laboratory. Three thousand bottles were manually filled with 10 ml of a nutrient broth under each of the three conditions. The liquid was transferred by means of a syringe and a needle that pierced through the rubber closures of the bottles. The numbers of contaminated bottles under those three conditions were 1, 1 and 2, respectively. Background contamination had no demonstrable effect on the incidence of contamination, provided that preparation of the simulated i.v.-admixture solution was performed by skilled personnel in an LAF cabinet and air contact is avoided.

Urinary platinum in hospital personnel occupationally exposed to platinum-containing antineoplastic drugs.

Urinary platinum levels of 21 nurses and hospital pharmacy personnel occupationally exposed to platinum containing antineoplastic drugs were determined in 24-h urine by voltammetric analysis after UV photolysis. All study participants applied standard safety measures, including a vertical laminar air-flow cabinet and latex gloves. The amount of platinum-containing drugs prepared for intravenous application ranged from 40-3260 mg/day. Urinary platinum was detected in 9 of 52 urine samples collected on days when platinum-containing drugs were mixed (limit of determination 4 ng/l). In comparison with a non-exposed control group, elevated urinary platinum levels were found in one pharmacist (35 ng/g creatinine) and one pharmacy technician (28 ng/g creatinine). The pharmacist's urinary platinum remained elevated after 2 days without occupational exposure to antineoplastic drugs. The urinary platinum level of the pharmacy technician dropped considerably after several weeks without handling cytostatic drugs. Voltammetric detection of urinary platinum is a highly sensitive method suitable for biological and environmental monitoring.

Sampling for airborne fluorouracil in a hospital drug preparation area

Fluorouracil content was studied in air samples from a hospital pharmacy work area where a vertical laminar-airflow biological safety cabinet (BSC) vented into the work area was used for the preparation of antineoplastic drugs. The BSC was run 24 hours per day and used approximately 6 hours per day. During one 56-hour period and one 95-hour period, a portable pump delivering room air at 2 L/min was positioned 12 inches outside the BSC. Fluorouracil content in extracts from 0.5-micron Teflon filters housed in the pump was assayed by high-performance liquid chromatography. The assay's lower limit of fluorouracil detection was 0.2 ng/cu m air. In the 56-hour sampling period and the 96-hour sampling periods, 25.4 g and 25 g, respectively, of fluorouracil were prepared. No fluorouracil was detectable in the filter extracts. Use of an appropriate biological safety cabinet in conjunction with good aseptic technique and the recommended procedures for safe handling of antineoplastic drugs may reduce the potential for dispersal of drug into the pharmacy workroom environment.

<i>In Vitro</i> Regeneration of Brinjal (<i>Solanum melongena</i> L.) Using Stem and Leaf Explants

DOI: 10.3329/jbs.v17i0.7126J. bio-sci. 17: 155-158, 2009

Development of a laminar air-flow biological cabinet.

Abstract Engineering and microbiological tests indicated that a typical, commercially available laminar flow cabinet was not effective in providing both product protection and agent containment. An analysis was conducted of the air flows within the cabinet and at the cabinet face, and design criteria were established. Several configurations were proposed for an actual air-flow evaluation in a mock-up unit. The configuration selected was evaluated for product protection and agent containment using controlled bacterial and viral aerosols. The aerosol challenges generated during these evaluations were considerably higher than normal in order to provide stringent test conditions. Test results indicated that the mock-up unit provided considerably better product protection than the commercial cabinet, and agent containment was improved. Based upon the results obtained in this study, detailed design drawings were prepared.

Microbiological studies on the performance of a laminar airflow biological cabinet.

Engineering and microbiological tests indicated that a typical, commercial laminar airflow cabinet was not effective in providing either product protection or agent containment. The cabinet was modified and tested through a series of alternate configurations to establish a set of design criteria. A mock-up cabinet was developed from these design criteria. The mock-up unit was evaluated for efficiency in providing both product protection and agent containment. In these evaluations, challenge methods were developed to simulate normal, in-use laboratory operations. Controlled bacterial or viral aerosol challenges were used at higher than normal levels to provide stringent test conditions. Test results indicated that the mock-up unit was considerably better in preventing agent penetration (0.1 to 0.2 particles per 100 ft(3) of air) than the commercial cabinet (5 to 6 particles per 100 ft(3) of air) during product protection tests. Similarly, agent containment was considerably better in the new cabinet (particle escape of 2 to 3 per 100 ft(3) of air at only one of the five test sites) than in the commercial cabinet (particle escape of 2 to 14 per 100 ft(3) of air at three of the five test sites).

Microbiological Studies on the Performance of a Laminar Airflow Biological Cabinet

Engineering and microbiological tests indicated that a typical, commercial laminar airflow cabinet was not effective in providing either product protection or agent containment. The cabinet was modified and tested through a series of alternate configurations to establish a set of design criteria. A mock-up cabinet was developed from these design criteria. The mock-up unit was evaluated for efficiency in providing both product protection and agent containment. In these evaluations, challenge methods were developed to simulate normal, in-use laboratory operations. Controlled bacterial or viral aerosol challenges were used at higher than normal levels to provide stringent test conditions. Test results indicated that the mock-up unit was considerably better in preventing agent penetration (0.1 to 0.2 particles per 100 ft(3) of air) than the commercial cabinet (5 to 6 particles per 100 ft(3) of air) during product protection tests. Similarly, agent containment was considerably better in the new cabinet (particle escape of 2 to 3 per 100 ft(3) of air at only one of the five test sites) than in the commercial cabinet (particle escape of 2 to 14 per 100 ft(3) of air at three of the five test sites).