Impact of excipient particle size on measurement of active pharmaceutical ingredient absorbance in mixtures using frequency domain photon migration.

A system of dual-component powder mixtures, varying in excipient particle size and concentration of active pharmaceutical ingredient (API), is analyzed using frequency domain photon migration (FDPM) techniques. The results show that the FDPM-measured absorption coefficient increases linearly with increasing API concentration whereas the isotropic scattering coefficient shows no sensitivity to changes in API concentration. It is further seen that the absorption coefficient of blends, owing to the API, is not only linearly dependent on its concentration, but that this relationship is furthermore related to the excipient particle size. Finally, a comparison between near-infrared absorbance and FDPM-measured isotropic scattering as a function of reciprocal particle size is made to highlight FDPM as a powerful particle sizing tool without need for calibration. Overall, this study presents FDPM as a comprehensive method for detection of API concentration independent of excipient particle size.

A long-term stable normothermic cardiopulmonary bypass model in neonatal swine.

BACKGROUND: An appropriate animal model of cardiopulmonary bypass (CPB) is critical in order to study the morbidity and mortality in newborn children undergoing long-term cardiac surgery. Since this has been reported to be technically difficult, this paper describes a neonatal porcine CPB model (3 days old, n = 18) for up to 8 h to study long-term bypass. METHODS: After anesthesia, neonates had arterial/venous monitoring lines inserted, they were heparinized (300 IU/kg), the aorta was cannulated for arterial retroperfusion, and a two-stage venous drainage catheter was placed in the right atrium. A Medtronic Minimax Plus oxygenator and the bypass circuit were primed with donor blood and CPB was instituted. RESULTS: Line and mean arterial pressures were kept at 147.7 +/- 73 and 62.7 +/- 9 mm Hg, respectively. Myocardial (38.1 +/- 1.0 degrees C) and rectal temperatures (37.7 +/- 1.0 degrees C) were maintained. Heart rate was 184.8 +/- 34.5 bpm. Hematocrits were 29.6 +/- 6.0%, activated clotting time was sustained above 400 s throughout bypass, blood gas parameters were maintained in the normal range (pH, 7.39 +/- 0.1; PO(2), 123.1 +/- 65.2 mm Hg; PCO(2), 37.2 +/- 8.5 mm Hg; and HCO(3)(-), 21.5 +/- 3.6 mmol/L). CPB was terminated after 8 h and no visceral edema or other imbalances normally associated with swine on bypass were observed. CONCLUSIONS: Results demonstrate a model of stable long-term bypass in neonatal swine which can be used to study issues critical to children requiring surgical correction and CPB at a young age. Overall effects of surgery and bypass on these younger patients have yet to be explored and therefore a stable long-term normothermic model of CPB would allow the study of numerous parameters associated with this complicated procedure.

Postischemic functional recovery in immature hearts is influenced by performance index and assessment technique.

In the in vivo immature heart, conflicting results are reported for postischemic functional recovery. This study determines whether interpretations of functional recovery are influenced by the contractile performance index (systolic pressure, developed pressure, and maximum rate of systolic pressure increase per unit time) reported or the assessment technique (isovolumetric and variable-volume) utilized. In neonatal pigs (n = 6) on cardiopulmonary bypass, each performance index was examined using both assessment techniques before myocardial ischemia and at 15, 30, and 60 min of reperfusion. With the use of the isovolumetric technique, all performance indexes had significantly different recovery. With the use of the variable-volume assessment technique, recovery of systolic pressure was significantly better than the other indexes. When recovery was compared between the two assessment techniques, systolic pressure recovered significantly better when assessed using the variable-volume technique. For each performance index, the correlation between isovolumetric and variable-volume techniques was positive before ischemia but negative during reperfusion, suggesting that the assessment techniques identified conflicting postischemic contractile performances. Thus both the contractile performance index reported and the assessment technique employed are ultimately important in interpreting postischemic functional recovery in the immature heart.

Metabolic and functional response of neonatal pig hearts to the development of ischemic contracture: is recovery possible?

The potential for functional and metabolic recovery in neonatal hearts after the development of ischemic contracture remains controversial and undefined. This study documents post-ischemic recovery of metabolism and function in the in vivo neonatal heart after the development of onset and peak ischemic contracture. In piglets on cardiopulmonary bypass, hearts were reperfused after the development of either onset (TICo) or peak (TICp) ischemic contracture. Systolic (developed and systolic function, contractility) and diastolic (diastolic function, relaxation) performance was assessed throughout reperfusion. Biopsies were obtained at end-ischemia or end-reperfusion to assess metabolism. By end-ischemia, the metabolic profiles of both TICo and TICp hearts confirmed energy-store depletion and purine degradation that was quantitatively greater in TICp hearts. Hearts reperfused at TICo had consistent moderate impairment of developed function, contractility, diastolic function, and relaxation, whereas hearts reperfused at TICp had much more profound functional impairment. Diastolic function showed the worst functional recovery. In contrast, systolic function was not significantly altered in either study group and, thus, did not reflect the actual extent of injury. In addition, TICo hearts either did not further deplete or partially regenerated energy stores during reperfusion, whereas TICp hearts had further energy-store depletion and lactate accumulation. In summary, neonatal hearts reperfused after TICo maintained or partially restored energy stores and had significant but incomplete functional recovery. In contrast, further metabolic deterioration and profound functional impairment occurred with reperfusion after TICp, potentially indicating ongoing mitochondrial injury and compromised oxidative phosphorylation.

Effects of hyperoxia on neonatal myocardial energy status and response to global ischemia.

BACKGROUND: This study examines the effect of neonatal exposure to clinically relevant hyperoxia levels on both in vivo myocardial metabolism and the subsequent metabolic response to global ischemia. METHODS: Three-day-old pigs were ventilated to normoxia (80 mm Hg, 2 or 5 hours, n = 11), mild hyperoxia (250 mm Hg, 2 hours, n = 9), or severe hyperoxia (500 mm Hg, 5 hours, n = 14). Ventricular biopsies obtained at the end of the ventilation period, and at early and late ischemia were analyzed for ATP, ADP, AMP, creatine phosphate, glycogen, and lactate. RESULTS: Hyperoxia did not significantly alter in vivo metabolism. During early ischemia, hearts exposed to severe hyperoxia had better ATP and glycogen preservation (p < 0.003). These hearts exhibited almost complete (92%) creatine phosphate depletion, in contrast to incomplete creatine phosphate use in all other neonatal hearts, even in the face of 30% ATP reductions. However, hearts exposed to severe hyperoxia also had a higher incidence of fibrillation during ischemia, which accelerated ATP and glycogen degradation. CONCLUSIONS: Although severe hyperoxia provided an energy-sparing effect during early ischemia, it also increased the incidence of ventricular fibrillation, which negated this beneficial effect.

Neonatal hemodynamic responses to extreme ranges of controlled graded hypoxia.

OBJECTIVES: To determine the hemodynamic responses to a wide range of specific, controlled, graded levels of hypoxic hypoxia over 120 mins in a neonatal porcine model and to identify the PaO2 threshold for altered hemodynamic homeostasis. DESIGN: Prospective, experimental, animal study. SETTING: University cardiovascular research laboratory. SUBJECTS: Three-day-old domestic swine. INTERVENTIONS: Anesthetized, intubated, and ventilated 3-day-old pigs (n = 88) were assigned to one of five predetermined graded PaO2 groups: Group I (normoxia, PaO2 = 80 torr [10.7 kPa]); group II (PaO2 = 60 torr [8.0 kPa]); group III (PaO2 = 40 torr [5.3 kPa]); group IV (PaO2 = 30 torr [4.0 kPa]); or group V (PaO2 = 20 torr [2.7 kPa]). MEASUREMENTS AND MAIN RESULTS: Hemodynamic parameters including heart rate, systolic blood pressure, diastolic blood pressure, mean arterial pressure (MAP), and pulse pressure were evaluated. Acid-base status (arterial pH and lactate) was monitored in each experimental group over the 120-min study period. Hemodynamic and acid-base parameters were unaltered in animals in groups I and II. In group III animals, blood pressure was maintained (partly by increased heart rate), and acid-base balance was unaltered. In contrast, group IV animals had a gradual and progressive decrease in systolic blood pressure, diastolic blood pressure, and MAP, and slightly decreased pulse pressure, despite sustained tachycardia. Group IV animals also developed mild lactic acidosis. Group V animals exhibited a biphasic hemodynamic response, while the heart rate response was characterized by tachycardia at the induction of hypoxia, which was reduced in magnitude by 120 mins. The biphasic hemodynamic response in this group of animals included an initial increase in systolic and pulse pressures, followed by a gradual and progressive decrease in systolic and diastolic blood pressures, MAP, and pulse pressure. In addition, group V animals also developed profound progressive lactic acidosis. CONCLUSIONS: In anesthetized neonatal pigs, tachycardia occurred in response to a PaO2 of 40 torr (5.3 kPa), and thus marked the threshold for altered hemodynamic homeostasis. Beyond this threshold, both the 30 torr (4.0 kPa) and 20 torr (2.7 kPa) groups had a PaO2- dependent "late" hypotension, while only the 20 torr (2.7 kPa) group had a significant biphasic hemodynamic response characterized by "early" hypertension. The "late" hypotension which occurred in these two profound hypoxia groups indicates an inability to adequately adjust hemodynamics during prolonged hypoxic hypoxia.

Blood lactate and acid-base balance in graded neonatal hypoxia: evidence for oxygen-restricted metabolism.

This study examines the neonatal response to graded hypoxia and determines the arterial PO2 (PaO2) threshold for oxygen-restricted metabolism as confirmed by the development of lactic acidosis and altered oxygen handling. Anesthetized, intubated, and ventilated 3-day-old pigs (n = 56) were randomly assigned to one of five predetermined acute (120 min) graded hypoxia groups: normoxia (PaO2 = 80 Torr) or mild (60 Torr), moderate (40 Torr), moderately severe (30 Torr), or severe (20 Torr) hypoxia. In moderate hypoxia, lactate and acid-base homeostasis were unaltered due to a significant increase in oxygen extraction (P < 0.05) that was sufficient to maintain the arteriovenous oxygen content difference (oxygen uptake). In moderately severe hypoxia, increased arterial lactate and decreased HCO3- and base excess were evidence of anaerobic metabolism, yet pH was unaltered, indicating adequate buffering. In this group, despite the increase in oxygen extraction, oxygen uptake was reduced, indicating the onset of oxygen-restricted metabolism. The severe hypoxia group had significantly increased lactate (21.7 +/- 3.9 mmol/l), decreased pH (7.01 +/- 0.07) and base excess (-21.5 +/- 3.0 mmol/l), and depletion of HCO3- (9.7 +/- 1.6 mmol/l) (P < 0.0001). Here, increases in oxygen extraction were severely limited by availability, resulting in significantly reduced oxygen uptake, anaerobic metabolism, and profound lactic acidosis.

Does the severity of acute hypoxia influence neonatal myocardial metabolism and sensitivity to ischemia?

The level of systemic hypoxia required to alter neonatal myocardial metabolism and its resultant effect on tolerance to global ischemia is unknown. This study examines myocardial purine nucleotides, glycogen (MG), lactate, creatine phosphate (CP) and the subsequent tolerance to ischemia in hearts exposed to varying levels of hypoxia (2 h). Three-day-old swine were randomly allocated into five study groups. Animals were anaesthetized and ventilated (2 h) with varying mixtures of medical air and nitrogen to achieve their target PaO2 (mmHg): normoxia (PaO2 = 80, n = 18), mild (PaO2 = 60, n = 10), moderate (PaO2 = 40, n = 12), moderately-severe (PaO2 = 30, n = 7) and severe (PaO2 = 20, n = 9). Arterial blood gases verified PaO2 and normal PaCO2 (39.5 +/- 0.5 mmHg). Subsequently, the heart was exposed and the metabolic profile determined from a freeze-clamp LV biopsy. The heart was excised and tolerance to ischemia determined by time (min) to ischemic contracture onset (TICo) and peak (TICp). The results demonstrated a tendency to decreased MG with progressive hypoxia which reached significance in severe hypoxia (6.6 +/- 2.7 mumol/g, P < 0.05). Despite a doubling of myocardial lactate with moderately-severe hypoxia, increases only reached significance with severe hypoxia (27.8 +/- 6.3 mumol/g, P < 0.0001). Despite the reduction in LV adenosine triphosphate (ATP) with severe hypoxia (2.16 +/- 0.68 mumol/g, P < 0.05), CP was unaltered.(ABSTRACT TRUNCATED AT 250 WORDS)

Multiple in vivo full-thickness myocardial biopsies by freeze-clamping.

A technique was developed to permit multiple large biopsies of the in vivo myocardium to measure certain metabolites (ATP, CP, myocardial glycogen, lactate) with minimal chance of ventricular fibrillation or severe alterations in the heart's ability to maintain BP. Adult pigs with normal hearts (n = 9) as well as with hypertrophy (n = 5) created by aortic banding 3-4 months prior were anesthetized, intubated, and ventilated. After a sternotomy, the pericardium was opened and the epicardial surface was cleared of any adhesions. Biopsies were taken from the distal to proximal end of the heart. A suture (3-O Prolene) was placed in a figure-eight pattern in the area to be biopsied. A piece of the myocardium (50-100 mg) was grasped using a pair of narrow dressing forceps and frozen in liquid nitrogen. When the sample was completely frozen, a No. 11 scalpel blade, held at a 90 degrees angle to the epicardium throughout the cut, was used to carve out the full thickness biopsy. The sample was placed immediately in liquid nitrogen. The myocardial defect was repaired by using the previously placed suture buttressed with pieces of subcutaneous fat to prevent tearing the epicardial surface. This in vivo biopsy method has negligible complications and can be repeated or taught with constant reliability.

The effect of varying arterial oxygen tension on neonatal acid-base balance.

Acute severe hypoxia, when the arterial partial pressure of O2 (PaO2) is sufficiently reduced, causes a shift from aerobic to anaerobic metabolism and can be fatal to neonates. The ability of the neonate to accommodate this shift, and prevent acidosis, could play a role in its ability to tolerate hypoxia. This study examines the effect of varying degrees of acute hypoxia on acid-base homeostasis in the neonate using a model of right to left shunting where PaO2 is decreased and, if severe, arterial partial pressure of CO2 is increased. Three-d-old swine were anesthetized, intubated, and mechanically ventilated. Ventilation with a gas mixture of medical air and 95% N2:5% CO2 resulted in four groups: 1) normoxia (n = 5); 2) mild hypoxia (n = 4); 3) moderate hypoxia (n = 3); and 4) severe hypoxia (n = 5), with PaO2 of 10.7, 8.0, 5.3, and 2.7 kPa (80, 60, 40, and 20 mm Hg), respectively. Acid-base status was evaluated via changes in arterial blood partial pressure of CO2, pH, HCO3-, and base excess at 0, 30, 60, and 120 min. Only the severe hypoxia group had significantly elevated (p less than 0.0001) arterial partial pressure of CO2 compared to the other groups at 60 and 120 min [8.7 +/- 0.5 and 8.0 +/- 0.5 kPa (65.5 +/- 3.7 and 60.0 +/- 3.7 mm Hg), respectively]. Base excess was unaltered in the normoxic and mild and moderate hypoxic groups, indicating no change in metabolic acid-base status. The severe hypoxic group had progressively decreased HCO3-, base excess, and pH at 60 and 120 min.(ABSTRACT TRUNCATED AT 250 WORDS)

Wikstromol, antitumor lignan from Wikstroemia foetida var. oahuensis Gray and Wikstroemia uva-ursi Gray (Thymelaeaceae).

The ethanol extracts of Wikstroemia foetida var. oahuensis and Wikstroemia uva-ursi showed antitumor activity against the P-388 lymphocytic leukemia (3PS) test system One PS-active constituent of both plants was the lignan wikstromol. Several inactive compounds were identified as daphnoretin, pinoresinol, and syringaresinol.

Separation of conformational isomers of bouvardin by high-pressure liquid chromatography.

Bouvardin, a bicyclic hexapeptide, was found to exhibit a solvent-dependent conformational distribution. HPLC was utilized to separate the two different conformers whose distribution ratio was dependent on the solvent of injection.

Odoratin and paucin: cytotoxic sesquiterpene lactones from Baileya pauciradiata (Compositae).

An ethanol extract of Baileya pauciradiata exhibited cytotoxic activity against the human epidermoid carcinoma of the nasopharynx and the lymphocytic leukemia test systems. Two constituents responsible for this activity were isolated and identified as odoratin and paucin. Their identities were proven by IR, PMR, and mass spectral data; elemental analysis; preparation of their acetates; and melting-point determinations. Odoratin was confirmed by comparison with an authentic sample.

Antitumor agents from Jatropha macrorhiza (Euphorbiaceae) III: acetylaleuritolic acid.

The triterpene acetylaleuritolic acid was isolated from Jatropha macrorhiza (Euphorbiaceae) and showed tumor-inhibitory properties toward the P-388 lymphocytic leukemia test system. The compound was identified by means of IR, PMR, and mass spectrometry and by its transformation into the known methyl acetyloleanolate.

Cytotoxic agents from Michelia champaca and Talauma ovata: parthenolide and costunolide.

The ethanol extract of Michelia champaca and the petroleum ether extract of Talauma ovata showed activity toward the human epidermoid carcinoma of the nasopharynx test system. The active constituents were sesquiterpene lactones, identified as parthenolide (C15H20O3) and costunolide (C15H20O2). Their identities were proven by elemental analyses, PMR, IR, and mass spectral data, melting-point and mixed melting-point determinations, and comparisons with authentic samples and spectra.