Enhanced exopolysaccharide production by Cordyceps militaris using repeated batch cultivation.

Cordyceps militaris exo-polysaccharides (EPS) have been reported to possess many benefits, such as anti-tumor, anti-inflammatory and antioxidant activities. In this study, the production of EPS via cultivation in a bioreactor was investigated. Glucose and yeast extract were determined to be the most suitable carbon and nitrogen sources for EPS production. The appropriate levels of glucose and yeast extract were 40 g/L and 10 g/L, respectively, resulting in EPS production of 1.686 g/L in a submerged culture. In the stirred-tank fermentor, an agitation rate of 150 rpm and aeration rate of 1.5 vvm were the most effective for EPS production. Due to the anchoring of mycelial cells on the wall of fermentor, a repeated batch approach was used. EPS production of C. militaris could be enhanced to a maximum of 5.713 g/L, with a productivity of 476 mg/L/day in the second run. The repeated batch approach was expected to generate higher EPS production, increase EPS yield and productivity and further simplify cultivation operations for bio-industrial application.

Kinetic analysis on precursors for iturin A production from Bacillus amyloliquefaciens BPD1.

In this study, the precursor effect for iturin A production was quantitatively analyzed. A strain identified as Bacillus amyloliquefaciens BPD1 (Ba-BPD1) was selected due to its ability to produce iturin A. The enhancement of iturin A production in a submerged culture was tested using various additives, including palmitic acid, oils, and complex amino acids. Among these, complex amino acids triggered the highest yield at 559 mg/L. The respective amino acids that contribute to the structure of iturin A were used as precursors. In fact, it was found that the addition of l-proline, l-glutamine, l-asparagine and l-serine could improve iturin A yield in the defined medium. However, during the kinetic analysis, all the amino acids exhibited a lower saturation level than l-serine, which exhibited a high saturation level at 1.2% resulting in an iturin A yield of 914 mg/L. In contrast, a negative effect was observed following the addition of l-tyrosine. To analyze the kinetic behavior of l-serine, three kinetic models were adopted: the kinetic order equation, the Langmuir kinetic equation, and a modified logistic equation. The regression results showed that the modified logistic model was the best fit for the kinetic behavior of l-serine as the major precursor, which could be further referred to the biosynthesis pathway of iturin A. Among the proposed processes for iturin A production, this study achieved the highest iturin A levels as a result of the addition of precursors.

Production of D-hydantoinase via surface display and self-cleavage system.

In this study, the cell surface expression system was the first time used to directly produce extracellular enzyme. In the plasmid construction, the truncated ice nucleation protein (INP) was fused with intein (INT) and target protein, D-hydantoinase (DHTase), to form the INP-INT-DHTase gene. The plasmid containing this gene was transformed into Escherichia coli ER2566 cells. The gene construct enables the expression of INP-INT-DHTase fusion protein, which might anchor on cell membrane surface. The induction conditions were studied and optimal conditions were as follows: E. coli ER2566 was incubated at 37°C and 200 rpm till OD600 reached 0.6. Then, 0.05 mM IPTG was added and the induction was conducted at 15°C for 24 h. The cell was harvested and resuspended in the cleavage buffer (50 mM Tris-HCl buffer, pH 6). The cleavage reaction was carried out at 25°C, and 100 rpm for 24 h. The DHTase with an activity of 0.225 U/ml and a purity of 63.2% was obtained via centrifugation. This study demonstrated the feasibility of direct extracellular enzyme production using E. coli via only two steps of centrifugation.

Ventricular fibrillation time constant for swine.

The strength-duration curve for cardiac excitation can be modeled by a parallel resistor-capacitor circuit that has a time constant. Experiments on six pigs were performed by delivering current from the X26 Taser dart at a distance from the heart to cause ventricular fibrillation (VF). The X26 Taser is an electromuscular incapacitation device (EMD), which generates about 50 kV and delivers a pulse train of about 15-19 pulses s(-1) with a pulse duration of about 150 micros and peak current about 2 A. Similarly a continuous 60 Hz alternating current of the amplitude required to cause VF was delivered from the same distance. The average current and duration of the current pulse were estimated in both sets of experiments. The strength-duration equation was solved to yield an average time constant of 2.87 ms +/- 1.90 (SD). Results obtained may help in the development of safety standards for future electromuscular incapacitation devices (EMDs) without requiring additional animal tests.

Taser blunt probe dart-to-heart distance causing ventricular fibrillation in pigs.

The maximum distance between the heart and a model Taser stimulation dart, called the dart-to-heart distance, at which the Taser can directly cause ventricular fibrillation (VF), was measured in pigs. A 9-mm-long blunt probe was advanced snugly through the surrounding tissues toward the heart. Five animals [pig mass=61.2+/-6.23 standard deviation (SD) kg] for ten dart-to-heart distances where the Taser caused VF were tested. The dart-to-heart distances where the Taser caused VF of the first stimulation site ranged from 4 to 8 mm with average 6.2 mm+/-1.79 (SD) and of the second stimulation site ranged from 2 to 8 mm with average 5.4 mm+/-2.41 (SD). The results help inform the evolving discussion of risks associated with Tasers.

Taser dart-to-heart distance that causes ventricular fibrillation in pigs.

Electromuscular incapacitating devices (EMDs), such as Tasers, deliver high current, short duration pulses that cause muscular contractions and temporarily incapacitate the human subject. Some reports suggest that EMDs can kill. To help answer the question, "Can the EMD directly cause ventricular fibrillation (VF)?", ten tests were conducted to measure the dart-to-heart distance that causes VF in anesthetized pigs [mass = 64 kg +/- 6.67 standard deviation (SD)] for the most common X26 Taser. The dart-to-heart distance that caused VF was 17 mm +/- 6.48 (SD) for the first VF event and 13.7 mm +/- 6.79 (SD) for the average of the successive VF events. The result shows that when the stimulation dart is close enough to the heart, X26 Taser current will directly trigger VF in pigs. Echocardiography of erect humans shows skin-to-heart distances from 10 to 57 mm (dart-to-heart distances of 1-48 mm). These results suggest that the probability of a dart on the body landing in 1 cm2 over the ventricle and causing VF is 0.000172.