Balanced Amino Acid Solution Research Articles

Phosphate feeding improves high-cell-concentration NS0 myeloma culture performance for monoclonal antibody production.

Phosphorus depletion was identified in high-cell-concentration fed-batch NS0 myeloma cell cultures producing a humanized monoclonal antibody (MAb). In these cultures, the maximum viable and total cell concentration was generally ca. 5 x 10(9) and 7 x 10(9) cells/L, respectively, without phosphate feeding. Depletion of essential amino acids, such as lysine, was initially thought to cause the onset of cell death. However, further improvement of cell growth was not achieved by feeding a stoichiometrically balanced amino acid solution, which eliminated depletion of amino acids. Even though a higher cell viability was maintained for a longer period, no increase in total cell concentration was observed. Afterwards, phosphorus was found to be depleted in these cultures. By also feeding a phosphate solution to eliminate phosphorus depletion, the cell growth phase was prolonged significantly, resulting in a total cell concentration of ca. 17 x 10(9) cells/L, which is much greater than ca. 7 x 10(9) cells/L without phosphate feeding. The maximum viable cell concentration reached about 10 x 10(9) cells/L, twice as high as that without phosphate feeding. Apoptosis was also delayed and suppressed with phosphate feeding. A nonapoptotic viable cell population of 6.5 x 10(9) cells/L, as compared with 3 x 10(9) cells/L without phosphate feeding, was obtained and successfully maintained for about 70 h. These results are consistent with the knowledge that phosphorus is an essential part of many cell components, including phospholipids, DNA, and RNA. As a result of phosphate feeding, a much higher integral of viable cell concentration over time was achieved, resulting in a correspondingly higher MAb titer of ca. 1.3 g/L. It was also noted that phosphate feeding delayed the cell metabolism shift from lactate production to lactate consumption typically observed in recombinant NS0 cultures. The results highlight the importance of phosphate feeding in high-cell-concentration NS0 cultures.

Correlation between amino acid induced changes in energy expenditure and protein metabolism in humans

The relationship between energy expenditure and protein metabolism during amino acid (AA) administration wasevaluated in normal humans. A balanced AA solution was infused for 180 min at five different rates: 20 (study I), 40 (study II) , 80 (study III) , 160 (study IV), and 240 mg · m 2 −1 · min −1 (study V), on separate days, in seven normal, overnight-fasted subjects (age 25 ± 2 y; height 172 ± 5 cm; weight 68 ± 4 kg). Indirect calorimetry and [I- 14C] leucine infusion techniques were employed. Basal total plasma AA concentration averaged 1827 ± 121 μmol/L and increased to 2192 ± 142, 2576 ± 158, 3677 ± 195, 5638 ± 237, and 7185 ± 261 μmol/L in studies I–V, respectively. Basal energy expenditure averaged 0.60 ± 0.02 kcal· M 2 −1· min −1 and increased slightly in studies I and II (to 0.62 ± 0.03, 0.63 ± 0.02, respectively), and significantly in studies III–V (to 0.65 ± 0.03, 0.70 ± 0.04, and 0.77 ± 0.05 kcal· m 2 −1 · min −1, respectively; all P < 0.01 versus basal; P < 0.05−0.01 for each study versus preceding study). Basal nonoxidative leucine disposal (NOLD), an index of protein synthesis, averaged 73 ± 3 μmol·m 2 −1·min −1 and increased, albeit not significantly, in studies I and II (to 75 ± 5, 76 ± 4, respectively). In contrast, a significant increase in NOLD was observed in studies III–V (to 87 ± 7, 103 ± 7, and 127 ± 9 μmol · ml 2 −1· min −1, respectively; all P < 0.01 versus basal; P < 0.05−0.01 for each study versus preceding study). Basal respiratory quotient averaged 0.81 ± 0.02 and did not change significantly in studies I–V (0.80 ± 0.02, 0.79 ± 0.02, 0.80 ± 0.03, 0.82 ± 0.02 and 0.82 ± 0.03, respectively). The thermic effect of AA administration, calculated as percent of the AA energy infused, was constant and averaged 24 ± 4, 19 ± 3, 17 ± 4, 17 ± 3, and 18 ± 3% in studies I–V, respectively. When AA-induced increase in protein synthesis was plotted with the increment in energy expenditure, a positive correlation was obtained ( r = 0.792, P < 0.001). In summary, during AA administration ( 1) the absolute rise in energy expenditure is dose-dependent and does not show evidence of achieving a plateau; ( 2) it is positively correlated with AA-induced protein synthesis; and ( 3) the thermic effect is not dependent upon the AA dose administered. The data provide a quantitative assessment of AA-induced thermogenesis in normal humans and the energy needs associated with an acute stimulation of protein synthesis.

Correlation between amino acid induced changes in energy expenditure and protein metabolism in humans.

The relationship between energy expenditure and protein metabolism during amino acid (AA) administration was evaluated in normal humans. A balanced AA solution was infused for 180 min at five different rates: 20 (study I), 40 (study II), 80 (study III), 160 (study IV), and 240 mg.m2(-1).min-1 (study V), on separate days, in seven normal, overnight-fasted subjects (age 25 +/- 2 y; height 172 +/- 5 cm; weight 68 +/- 4 kg). Indirect calorimetry and [1-14C] leucine infusion techniques were employed. Basal total plasma AA concentration averaged 1827 +/- 121 mumol/L and increased to 2192 +/- 142, 2576 +/- 158, 3677 +/- 195, 5638 +/- 237, and 7185 +/- 261 mumol/L in studies I-V, respectively. Basal energy expenditure averaged 0.60 +/- 0.02 kcal.m2(-1).min-1 and increased slightly in studies I and II (to 0.62 +/- 0.03, 0.63 +/- 0.02, respectively), and significantly in studies III-V (to 0.65 +/- 0.03, 0.70 +/- 0.04, and 0.77 +/- 0.05 kcal.m2(-1).min-1, respectively, all P < 0.01 versus basal; P < 0.05-0.01 for each study versus preceding study). Basal nonoxidative leucine disposal (NOLD), an index of protein synthesis, averaged 73 +/- 3 mumol.m2(-1).min-1 and increased, albeit not significantly, in studies I and II (to 75 +/- 5, 76 +/- 4, respectively). In contrast, a significant increase in NOLD was observed in studies III-V (to 87 +/- 7, 103 +/- 7, and 127 +/- 9 mumol.m2(-1).min-1, respectively; all P < 0.01 versus basal; P < 0.05-0.01 for each study versus preceding study). Basal respiratory quotient averaged 0.81 +/- 0.02 and did not change significantly in studies I-V (0.80 +/- 0.02, 0.79 +/- 0.02, 0.80 +/- 0.03, 0.82 +/- 0.02 and 0.82 +/- 0.03, respectively). The thermic effect of AA administration, calculated as percent of the AA energy infused, was constant and averaged 24 +/- 4, 19 +/- 3, 17 +/- 4, 17 +/- 3, and 18 +/- 3% in studies I-V, respectively. When AA-induced increase in protein synthesis was plotted with the increment in energy expenditure, a positive correlation was obtained (r = 0.792, P < 0.001). In summary, during AA administration (1) the absolute rise in energy expenditure is dose-dependent and does not show evidence of achieving a plateau; (2) it is positively correlated with AA-induced protein synthesis; and (3) the thermic effect is not dependent upon the AA dose administered. The data provide a quantitative assessment of AA-induced thermogenesis in normal humans and the energy needs associated with an acute stimulation of protein synthesis.

Beneficial effect of enteral glycine in intestinal ischemia/reperfusion injury.

It has been shown in vitro that glycine can protect renal tubules and hepatocytes from hypoxic injury. Glycine also attenuates ischemic injury in transplanted livers. The present study investigated the effect of enteral glycine in a murine model of ischemia/reperfusion injury of the small intestine. Mice (n = 12 in each group) were randomized to receive two gastric gavages of either a 20% glycine (Gly) or 23% balanced amino acid (AA) solution with a 6-hour interval between each gavage. One hour after the second gavage, mice underwent superior mesenteric artery clamping for 20 minutes. The clamp was then released for reperfusion. Another group of mice (n = 8) underwent a sham operation and served as additional control animals. Six hours after ischemia/reperfusion, the mice were killed in order to assess the intestinal injury (intestinal protein content, mucosal disaccharidase activity, and intestinal histologic findings) and the systemic consequences (bacterial translocation, serum interleukin-6, and lung myeloperoxidase activity). A second set of mice (n = 55) underwent identical gavages and ischemia/reperfusion and they were followed for survival. Compared to AA, enteral glycine administered prior to intestinal ischemia/reperfusion injury significantly preserved mucosal indices and intestinal histology and decreased lung myeloperoxidase activity. Survival was also significantly increased in animals receiving glycine compared to AA control mice. These data suggest that enteral glycine supplementation may be beneficial in attenuating intestinal ischemia/reperfusion injury and its related systemic effects in this murine model.

Beneficial effect of enteral glycine in intestinal ischemia/reperfusion injury

It has been shown in vitro that glycine can protect renal tubules and hepatocytes from hypoxic injury. Glycine also attenuates ischemic injury in transplanted livers. The present study investigated the effect of enteral glycine in a murine model of ischemia/reperfusion injury of the small intestine. Mice (n = 12 in each group) were randomized to receive two gastric gavages of either a 20% glycine (Gly) or 23% balanced amino acid (AA) solution with a 6-hour interval between each gavage. One hour after the second gavage, mice underwent superior mesenteric artery clamping for 20 minutes. The clamp was then released for reperfusion. Another group of mice (n = 8) underwent a sham operation and served as additional control animals. Six hours after ischemia/reperfusion, the mice were killed in order to assess the intestinal injury (intestinal protein content, mucosal disaccharidase activity, and intestinal histologic findings) and the systemic consequences (bacterial translocation, serum interleukin-6, and lung myeloperoxidase activity). A second set of mice (n = 55) underwent identical gavages and ischemia/reperfusion and they were followed for survival. Compared to AA, enteral glycine administered prior to intestinal ischemia/reperfusion injury significantly preserved mucosal indices and intestinal histology and decreased lung myeloperoxidase activity. Survival was also significantly increased in animals receiving glycine compared to AA control mice. These data suggest that enteral glycine supplementation may be beneficial in attenuating intestinal ischemia/reperfusion injury and its related systemic effects in this murine model.

Hypermetabolism and increased peripheral release of amino acids after subarachnoidal hemorrhage and its operative treatment

The metabolic response to surgery for acute subarachnoidal hemorrhage and its modification by amino acid infusions was studied. Thirty patients with acute subarachnoidal hemorrhage were randomly assigned to receive for 12 h either an infusion of glucose and a balanced amino acid solution (1.68 MJ = 400 kcal/d and 0.15 gN · kg −1 · d −1; group AA) or a glucose and a solution containing 20% of total nitrogen as alanyl-glutamine (1.68 MJ = 400 kcal/d and 0.15 gN · kg −1 · d −1; group ALAGLN). A separate control group received glucose alone (1.68 MJ = 400 kcal/d). The infusions started 12 h after operation. All patients received corticosteroids. Despite a higher arterial glutamine concentration in the ALAGLN-group (791 ± 195 μmol/L vs. AA 581 ± 112 μmol/L, and control 571 ± 82 μmol/L; p < 0.05) the net release of glutamine from the leg was similar in all groups (ALAGLN: 39 ± 47 μmol/min, AA: 26 ± 18 μmol/min, and control: 24 ± μmol/min, NS). Also the release of alanine (ALAGLN: 35 ± 24 μmol/min, AA: 34 ± 24 μmol/min, and control: 30 ± 18 μmol/min) and total amino acids (ALAGLN: 133 ± 131 μmol/min, AA: 125 ± 98 μmo/min, and control: 112 ± 72 μmol/min) were similar in all groups. All groups were characterized by a pattern of preoperative hypermetabolism that persisted after the operation. The hypermetabolism was not related to increased peripheral oxygen consumption, since femoral oxygen consumption (VO 2) represented only 3% of the whole body VO 2.

Differential responsiveness of protein synthesis and degradation to amino acid availability in humans.

We investigated the effects of graded hyperaminoacidemia on protein metabolism in eight healthy, young (25 +/- 2 years), normal weight (BMI = 25 +/- 1 kg/m2), overnight-fasted human subjects. A balanced amino acid solution was infused for 180 min at five different rates: 0.5 (study I), 1.0 (study II), 2.0 (study III), 4.0 (study IV). and 6.0 (study V) mg x kg-1 x min-1 on separate days in random order. Studies were performed with [1-14C]leucine infusion and indirect calorimetry to calculate leucine oxidation (LOX), nonoxidative leucine disposal (NOLD) (an index of protein synthesis), and endogenous leucine flux (ELF) (an index of proteolysis). Basal total plasma amino acid concentrations averaged 1.85 +/- 0.1 mmol/l and increased to 2.27 +/- 0.1, 2.70 +/- 0.2, 3.84 +/- 0.2, 5.87 +/- 0.4, and 7.52 +/- 0.3 mmol/l in studies I-V, respectively. ELF decreased from a basal value of 2.27 +/- 0.2 to 2.12 +/- 0.2, 1.97 +/- 0.1, 1.73 +/- 0.2, 1.67 +/- 0.3, and 1.65 +/- 0.1 micromol x kg-1 x min-1 in studies I-V, respectively (P < 0.05 for study I vs. basal, P < 0.01 for studies II-V vs. basal, and NS for studies IV and V vs. study III). LOX increased from a basal value of 0.31 +/- 0.04 to 0.38 +/- 0.05, 0.41 +/- 0.02, 0.64 +/- 0.04, 1.11 +/- 0.07, and 1.56 +/- 0.05 micromol x kg-1 x min-1 in studies I-V (all P < 0.01 vs. basal; P < 0.05-0.01 for each study vs. preceding study). Basal NOLD averaged 1.96 +/- 0.2 and did not change significantly in studies I and II (2.03 +/- 0.2 and 2.10 +/- 0.1 micromol x kg-1 x min-1). In contrast, a significant increase in NOLD was observed in studies III, IV, and V (to 2.3 +/- 0.15, 2.74 +/- 0.2, and 3.25 +/- 0.7 micromol x kg-1 x min-1, respectively; all P < 0.01 vs. basal; P < 0.05-0.01 for each study vs. preceding study). The net leucine balance (difference between ELF and NOLD)(-0.31 +/- 0.06 micromol x kg-1 x min-1) became less negative in study I (P < 0.01 vs. basal) and positive during studies II-V when the rise in plasma total amino acid levels was >/= 50% above basal level (P < 0.01 vs. each preceding study). In conclusion, NOLD, ELF, and LOX exhibit a differential responsiveness to acute changes in substrate availability: 1) small increments (25-50%) in plasma amino acid levels inhibit ELF and stimulate LOX but have no effect on NOLD; 2) stimulation of NOLD is observed only with increments in plasma amino acid levels >/= 100% above basal values; and 3) increments in plasma amino acid concentrations > 100% above basal values cause a progressive dose-related increase in LOX and NOLD but do not induce any further inhibition of ELF.

Effect of prednisolone on amino acid-induced changes in renal haemodynamics and tubular function

The effect of oral prednisolone treatment on renal haemodynamics, tubular function and various hormones during amino acid infusion was studied in 14 normal men. A balanced amino acid solution was infused for 120 min, before and after 4 days of prednisolone treatment (40 mg day-1). During amino acid infusion before prednisolone glomerular filtration rate, renal plasma flow, urinary sodium excretion, fractional excretion of sodium, lithium clearance, fractional excretion of lithium, serum insulin (s-insulin), plasma glucagon (p-glucagon) and s-growth hormone increased, whereas p-atrial natriuretic peptide, p-aldosterone, p-vasopressin and s-insulin-like growth factor 1 were unchanged, and potassium excretion and fractional excretion of potassium fell. After prednisolone treatment the most important differences during amino acid infusion were a significantly lower fractional excretion of sodium after 120 min (before prednisolone 26%; after prednisolone-7%; p < 0.05), a more pronounced increase in s-insulin after 120 min (before 118%; after 200%; p < 0.05) and a lower s-potassium. In conclusion, amino acid infusion increased fractional sodium excretion in healthy men, and this increase was reduced by prednisolone due to increased reabsorption in the distal tubules. It is suggested that the more pronounced the increases in plasma insulin and the decrease in serum potassium are mediators of the increased distal tubular sodium reabsorption during amino acid infusion during prednisolone treatment.

Nutritional Aspects in Patients with Acute Renal Failure/ Multiorgan Failure

Nutrition support contributes to reducing morbidity and mortality in patients with the multiple-organ dysfunction syndrome (MODS). In cases when acute renal failure is a component of MODS, the goals of nutritional management should be to control general nutrient deficiencies and correct specific metabolic alterations caused by the loss of renal function or acute uraemia without worsening uraemic toxicity or disturbing electrolyte balance. In most cases energy requirements, which should ideally be estimated by indirect calorimetric measurements, amount to 30 kcal/kg/day and only seldom exceed 35 kcal/kg/day. Depending on the degree of protein catabolism, 1.0-1.5 g/kg/day of protein should be given. A balanced amino acid solution containing both essential and nonessential amino acids should be administered. Uraemia impairs protein synthesis and nitrogen administration should, therefore, not be started until the uraemic state is adequately controlled by dialysis or haemofiltration.

Stress hormone and amino acid infusion in healthy volunteers: Short-term effects on protein synthesis and amino acid metabolism in skeletal muscle

To study the immediate effects of stress hormones and intravenous amino acid support, healthy male volunteers were administered a stress-hormone infusion including epinephrine, cortisol, and glucagon either alone (Triple, n = 8) or combined with a balanced glutamine-free amino acid solution (Triple AA, n = 8) over a period of 6 hours. The amino acid infusion was started 2 hours after the hormone infusion. A third group (AA, n = 8) received the balanced amino acid solution alone. After 6 hours of the stress-hormone infusion, a decrease was observed in skeletal muscle protein synthesis as measured by the size distribution and concentration of ribosomes. The decrease was prevented by an infusion of the balanced amino acid solution. Following the triple-hormone infusion, a decrease was noted in the content of the total free amino acids in both muscle and plasma. After including amino acids in the infusion solution, the significant decrease in muscle glutamine caused by the triple hormones was not seen. Plasma cortisol, insulin, and glucose increased in response to the triple-hormone infusion alone or in combination with amino acids. In summary, the results show that the signs of muscle protein catabolism elicited by administration of stress hormones can be attenuated by simultaneous administration of a conventional amino acid solution, although it does not contain glutamine.

Effects of valine, leucine, isoleucine, and a balanced amino acid solution on the seizure threshold to picrotoxin in rats

During infusion of branched-chain amino acids (BCAAs) in humans, changes in ventilatory drive, sleeping pattern, and appetite have been reported. The mechanism by which BCAA exerts their effects on CNS remains unclear. An infusion of a BCAA solution (300 mg/kg) has previously been found to increase the seizure threshold in rats to the proconvulsant drug picrotoxin, an antagonist on the GABA-benzodiazepine receptor complex. In this study, each of the BCAAs given separately (valine, leucine, isoleucine; 300 mg/kg) ( n = 10) increased the mean latency time to onset of seizures vs. placebo as an indication of an increased seizure threshold. A balanced amino acid solution (Vamin-Glucose ®) had no effect on the seizure threshold. Thus, these CNS effects are specific for BCAAs and occur with all three.

Different sensitivity of glucose and amino acid metabolism to insulin in NIDDM.

NIDDM subjects are characterized by impaired glucose tolerance and insulin resistance with respect to glucose metabolism. To examine whether the defect in glucose utilization extends to amino acid metabolism, 6 NIDDM subjects (64 +/- 4 yr of age; ideal body weight of 107 +/- 3%) and 7 control subjects (58 +/- 4 yr of age; ideal body weight of 105 +/- 2%) were studied with the euglycemic insulin clamp technique, in combination with [1-14C]leucine and indirect calorimetry. All subjects participated in two studies. In study 1, after 3 h of tracer equilibration, a 3-h insulin clamp (40 mU.m-2 x min-1) was performed to define the effect of insulin on leucine kinetics and glucose metabolism. In study 2, subjects received a repeat 3-h insulin clamp, and a balanced amino acid solution was infused to increase the plasma amino acid concentrations approximately 2-fold to examine the effect of combined physiological hyperinsulinemia-hyperaminoacidemia on the rate of leucine and glucose disposal. Insulin-mediated total body glucose uptake was significantly reduced in NIDDM during both study 1 (5.6 +/- 0.4 vs. 6.9 +/- 0.6 mg.kg-1 x min-1, P < 0.01) and study 2 (5.2 +/- 0.4 vs. 6.8 +/- 0.6, P < 0.01). Basal plasma leucine (120 +/- 10 vs. 123 +/- 11 microM) and alpha-ketoisocaproic acid concentrations (28 +/- 3 vs. 25 +/- 2 microM) were similar in NIDDM and control subjects, respectively. In contrast, the basal plasma glucose concentration (8.9 +/- 0.8 vs. 4.7 +/- 0.2 microM) and the HbA1c (8.5 +/- 0.2 vs. 5.7 +/- 0.2%) were significantly increased in NIDDM (P < 0.01). In the postabsorptive state, endogenous leucine flux, leucine oxidation, and nonoxidative leucine disposal were similar in NIDDM and control subjects. When insulin was infused without amino acids (study 1), the decrement in plasma leucine (53 +/- 5 vs. 48 +/- 4 microM), endogenous leucine flux (13 +/- 2 vs. 11 +/- 1 mumol.m-2 x min-1), leucine oxidation (1.6 +/- 0.2 vs. 1.3 +/- 0.1 mumol.m-2 x min-1), and nonoxidative leucine disposal (10 +/- 1 vs. 8 +/- 1 mumol.m-2 x min-1) was comparable in both groups. During combined insulin and amino acid infusion (study 2), plasma leucine concentration (185 +/- 20 vs. 190 +/- 15 microM) rose similarly in NIDDM and control subjects.(ABSTRACT TRUNCATED AT 400 WORDS)

Effect of insulin and plasma amino acid concentration on leucine metabolism in cirrhosis

Clinically stable patients with cirrhosis demonstrate insulin resistance with regard to glucose metabolism. However, much less is known about the two major factors, insulin and plasma amino acid concentration, that regulate protein metabolism in cirrhotic patients. To examine this question, we performed paired euglycemic insulin clamp studies in combination with 14Cleucine and indirect calorimetry. In the first study insulin alone was infused, and the plasma amino acid concentration was allowed to decline, During the second study a balanced amino acid solution was infused with insulin to increase the total plasma amino acid concentration approximately twofold. Insulinmediated glucose disposal (4.68 vs. 6.45 mg/kg-min, p < 0.01) was significantly impaired by 30% in cirrhotic patients during both insulin clamp studies. In the postabsorptive state, cirrhotic patients manifested low plasma leucine (76 vs. 102 μmol/L) and α-ketoisocaproate (19 vs. 30 μmol/L) concentrations, but all parameters of leucine turnover were normal. When insulin alone was infused, the endogenous leucine flux (an index of protein degradation) declined similarly in cirrhotic patients (30.8 μmol/m2-min) and control (26.9) subjects, and this was accompanied by a similar decrease in plasma leucine concentration (31% vs. 33%). The decline in circulating leucine concentration was accompanied by a parallel decline in leucine oxidation (5.1 vs. 4.6 μmol/m2-min) and nonoxidative (28.9 vs. 26.0 μmol/m2-min) leucine disposal, which were of similar magnitude in cirrhotic patients and control subjects, respectively. In both cirrhotic patients and control subjects, combined hyperinsulinemia/hyperaminoacidemia elicited a similar stimulation of nonoxidative leucine disposal (an index of protein synthesis) and leucine oxidation while causing a greater suppression of endogenous leucine flux than observed with insulin alone. Thus the suppressive effect of insulin on protein degradation and the stimulatory effect of insulin/amino acid infusion on protein synthesis are not impaired in cirrhotic patients, demonstrating a clear-cut dissociation between the effects of insulin on protein and glucose metabolism.

Effect of insulin and plasma amino acid concentration on leucine metabolism in cirrhosis

Clinically stable patients with cirrhosis demonstrate insulin resistance with regard to glucose metabolism. However, much less is known about the two major factors, insulin and plasma amino acid concentration, that regulate protein metabolism in cirrhotic patients. To examine this question, we performed paired euglycemic insulin clamp studies in combination with 14C-leucine and indirect calorimetry. In the first study insulin alone was infused, and the plasma amino acid concentration was allowed to decline. During the second study a balanced amino acid solution was infused with insulin to increase the total plasma amino acid concentration approximately twofold. Insulin-mediated glucose disposal (4.68 vs. 6.45 mg/kg-min, p less than 0.01) was significantly impaired by 30% in cirrhotic patients during both insulin clamp studies. In the postabsorptive state, cirrhotic patients manifested low plasma leucine (76 vs. 102 mumol/L) and alpha-ketoisocaproate (19 vs. 30 mumol/L) concentrations, but all parameters of leucine turnover were normal. When insulin alone was infused, the endogenous leucine flux (an index of protein degradation) declined similarly in cirrhotic patients (30.8 mumol/m2-min) and control (26.9) subjects, and this was accompanied by a similar decrease in plasma leucine concentration (31% vs. 33%). The decline in circulating leucine concentration was accompanied by a parallel decline in leucine oxidation (5.1 vs. 4.6 mumol/m2-min) and nonoxidative (28.9 vs. 26.0 mumol/m2-min) leucine disposal, which were of similar magnitude in cirrhotic patients and control subjects, respectively. In both cirrhotic patients and control subjects, combined hyperinsulinemia/hyperaminoacidemia elicited a similar stimulation of nonoxidative leucine disposal (an index of protein synthesis) and leucine oxidation while causing a greater suppression of endogenous leucine flux than observed with insulin alone. Thus the suppressive effect of insulin on protein degradation and the stimulatory effect of insulin/amino acid infusion on protein synthesis are not impaired in cirrhotic patients, demonstrating a clear-cut dissociation between the effects of insulin on protein and glucose metabolism.

Effect of Balanced Amino Acid Solution on Protein Metabolism after Surgery

The aim of this study was to investigate the nitrogen-sparing effect of hypocaloric parenteral nutrition (HPN) with a balanced amino acid supply compared with that of a standard Rose's formula. Eighteen well-nourished patients with colorectal cancer were randomly distributed into 3 groups: I control, II Study group, III Rose's formula. The amount of non-protein calories was similar in both groups: 8 kcal/kg administered as glucose, and 1.33 g of amino acid/kg was also infused during 5 days. There were no differences in the plasma protein levels of total proteins, albumin, and transferrin, but retinol-binding protein was higher in group II compared with the control. Nitrogen balance was significantly better in group II than in group III during the 3rd and 4th days. There were differences in the concentration of aspartic acid and glycine in the plasmatic aminograms between groups II and III. It was concluded that the balanced amino acid solution does not significantly improve a patient's nutritional state. Nor have we observed that the balanced amino acid solution causes fewer modifications in the plasma aminograms. However, it does achieve a slight improvement in the nitrogen balances and the synthesis of retinol-binding protein.

Psychomotor, subjective and neuroendocrine effects of acute tryptophan depletion in the healthy volunteer

SummaryThe effects of acute tryptophan depletion were investigated in 20 healthy volunteers. Ten of them received a balanced amino acid solution and 10 a tryptophan-free solution. The fall in tryptophan levels induced by the oral administration of a mixture tryptophan-free of L-amino acids was - 77% for free tryptophan and - 81% for total tryptophan. Before treatment, there were intergroup differences affecting alertness parameters (critical flicker threshold, recognition reaction time), plasma levels of prolactin and baseline performance in the proofreading correction test. In the tryptophan-depleted group, the number of errors reported during the unpleasant sound signal was increased (+ 48%) after treatment, whereas the number of errors fell (— 15%) in the group receiving the tryptophan supplement. Conversely the levels of prolactin were correlated with those of serum tryptophan. Few subjective effects were reported.

The effects of branched chain amino-acids upon postoperative muscle protein synthesis and nitrogen balance

Patients undergoing elective cholecystectomy were given total parenteral nutrition containing adequate amounts of calories and nitrogen postoperatively. One group (n=8) received a solution enriched with branched chain amino-acids (BCAA) and a control group (n=9) was given a balanced amino-acid solution. Muscle biopsy specimens were taken before surgery and on the third postoperative day. Muscle protein synthesis was assessed by comparing the concentration and size distribution of ribosomes between these two occasions. The total ribosome concentration per mg of DNA was decreased by 24% postoperatively at 3 days in both groups as compared with the preoperative value (p<0.05), indicating a diminished capacity for protein synthesis. The percentage proportion of polyribosomes was lower after than before surgery (p<0.05), reflecting a decrease in the activity of muscle protein synthesis. No significant difference between the two groups was detected. The nitrogen balance was negative throughout the study period and the cumulative nitrogen balance did not differ between the two groups. In this study the effect of BCAA upon muscle protein synthesis after elective abdominal surgery was evaluated. It is concluded that BCAA-enrichment of the amino-acid support failed to prevent the decrease in protein synthesis in skeletal muscle and the negative nitrogen balance observed when a conventional TPN programme is given.

Anti-cancer therapy with valine-depleted amino acid imbalance solution.

For the purpose of developing amino acid imbalance solution applicable to cancer treatment, we prepared seven kinds of amino acid imbalance solutions based on a 10% balanced amino acid solution and investigated the anti-tumor effects of each solution. The administration of valine-depleted amino acid solution for 8 days at a daily dose of 53 ml/rat (79.5 kcal/rat) resulted in the most significant inhibitory effects on the growth of hepatoma (AH109A) and mammary tumors (MRMT-1), the rate being 82.8% and 90.8%, respectively. Side effects observed were inhibition of increases in host body weight, weight loss of the spleen and thymus, loss of hair, and a decrease in the amounts of total plasma protein and albumin. When the daily dosage of valine-depleted amino acid solution was reduced to 40 ml/rat (58.3 kcal/rat), anti-tumor effects were still noticed, while side effects were abated. These findings indicate that side effects accompanying the use of this solution can be alleviated by controlling the ingredients of the solution as well as the amount administered. It is thus suggested that valine-depleted amino acid imbalance solution is a valuable tool in the treatment of cancer.

Transport kinetics of amino acids across the resting human leg.

Flux rates of amino acids were measured across the leg after an overnight fast in resting human volunteers. A balanced amino acid solution was, after a primed infusion, continuously infused for 2 h at each of three step-wise and increasing rates corresponding to 8.3, 16.7, 33.2 mg N/kg per h that were equivalent to 0.2, 0.4, 0.8 g N/kg per d. Flux of amino acids across the leg was compared with the flux of glucose, glycerol, lactate, free fatty acids, and oxygen. The size of the muscular tissue pool of amino acids was measured. Whole body amino acid oxidation was estimated by means of the continuous infusion of a 14C-labeled mixture of amino acids. Arterial steady state levels were obtained for most amino acids within 30 to 45 min after the primed constant infusion. Leg flux of amino acids switched from a net efflux after an overnight fast to a balanced flux between infusion rates corresponding to 0.2-0.4 g N/kg per d. At 0.8 g N/kg per d essentially all amino acids showed uptake. The infusion of amino acids stimulated leg uptake of glucose and lactate production and decreased FFA release. Oxygen uptake and leg blood flow increased significantly with increased infusion of amino acids. There was significant variability in transport rate among individual amino acids. Branched chain amino acids showed rapid transport and methionine slow transport rate. Only small changes in the muscle tissue concentration of certain amino acids were registered after 6 h of amino acid infusion despite uptake for several hours. When amino acids were infused at a rate corresponding to 0.8 g N/kg per d, the leg uptake of amino acids was 6% and the simultaneous whole body oxidation of infused amino acids was approximately 10%. Net uptake of leucine across the leg per hour was 62% of the muscle pool of free leucine when amino acids were infused at a rate corresponding to 0.4 g N/kg per d. Multiple regression analysis showed that the arterial concentration of an amino acid was the most important factor for uptake, more so than insulin concentration and blood flow. It is concluded that leg exchange of amino acids is large enough to rapidly change the pool size of the amino acids in skeletal muscle, if not counter-regulated by changes in rates of protein synthesis and degradation. Estimates of the capacity for protein synthesis and transfer RNA acceptor sites in muscles agree in order of magnitude with the net uptake of amino acids at high infusion rates of amino acids. Therefore, measurements of the balance of tyrosine, phenylalanine, and particularly methionine at steady state may reflect net balance of proteins across skeletal muscles even in short-time experiments.

Effect of branched-chain amino acids and insulin on postinjury protein catabolism in growing animals.

Muscle proteolysis continues to occur in hypercatabolic states despite the administration of carbohydrates and proteins. Recent clinical and experimental studies have demonstrated that, under catabolic conditions, treatment with either branched-chain amino acids (BCAA) or insulin may decrease negative nitrogen balance. However, the use of BCAA-enriched solutions to inhibit muscle proteolysis has never been tested in growing animals. A study was therefore undertaken to assess the effectiveness of such solutions, with or without insulin, as compared to a more balanced amino acid solution, in preventing or diminishing postinjury protein catabolism in growing animals. Fifteen-day-old rabbits, exposed to standard moderate trauma in the form of crushing the muscle mass of one rear thigh, received one of two amino acid formulations--a balanced amino acid solution (18.8% BCAA) or a 35% BCAA-enriched solution--for 96 hr. Insulin was given to subgroups of both series. The results indicate that: (1) nitrogen balance in nontraumatized animals is clearly superior when balanced amino acids are administered; (2) BCAA-enriched solutions may decrease postinjury muscle protein catabolism; (3) after trauma, insulin also has a nitrogen-conserving effect, which is demonstrated when it is combined both with BCAA-enriched (35%) and balanced amino acid (18.8%) solutions. However, a better nitrogen balance is achieved when insulin is associated with the balanced amino acid solution.