High performance liquid chromatography of plasma free amino acid patterns in liver disease and their diagnostic implications

The free amino acid levels in plasma of 24 chronic liver disease patient with chronic active hepatitis, cirrhosis and hepatoma and in 10 patients with acute hepatitis as well as 10 healthy controls were measured using high performance liquid chromatography. The aim of this work was to detect the free amino acid pattern in each group, to study the level of the molar ratio of branched chain amino acids, to aromatic amino acids, to correlate this ratio with conventional liver function tests and to detect its role in discrimination between chronic active hepatitis and cirrhosis. Plasma chromatograms of amino acids revealed significant increased fasting levels of aromatic amino acids and methionine in all chronic groups versus control group, while branched chain amino acids were significantly decreased. The mean molar ratio of branched chain amino acids/aromatic amino acids was 2.59, 1.43, 1.94 and 4.48 in chronic active hepatitis, cirrhosis, hepatoma and controls respectively. All groups showed significant difference compared with the controls [p<0.001]. The mean molar ratio of cirrhotic patients was significantly decreased compared to chronic active hepatitis [CAH] [p<0.001]. The molar ratio correlated significantly with albumin [r=0.73] and showed a significant negative correlation with aspartate aminotransferase in cirrhotic patients [r=0.62]. A decreased molar ratio was also detected in acute liver disease in comparison with the control group [p<0.001]. We concluded that the mean molar ratio decreases in chronic and acute liver disease. It correlates well with the synthetic capacity of the liver as indicated by albumin and correlates negatively with parenchymal damage as indicated by aspartate aminotransferase. It can be used as a useful indicator in discrimination between chronic active hepatitis and cirrhosis

Melatonin, the hormone of the pineal body: its role as an endogenous biological marker in major depression and breast cancer

The present study was designed to assess the relation of melatonin to two important pathological conditions namely; major depression [MD] and breast cancer [BC], with the intention of evaluating its role as an endogenous biological marker of both conditions and its potential clinical significance in follow-up of such cases. For this purposes 50 female patients with major depression [20 patients before treatment and 30 patients under treatment] in addition to 73 female patients with breast cancer [28 in stages I and II : early BC; 25 in stages III and IV late BC and 20 after radical mastectomy] were chosen for assessment of the serum melatonin levels. Nocturnal blood samples were collected from the MD group. whereas morning samples were collected from the BC group. Results were compared to those of an age-matched control group consisting of 20 healthy females. Nocturnal serum melatonin levels were significantly decreased in MD patients before the start of therapy as compared to the control group [P < 0.0001]. Meanwhile, the results of patients under antidepressant therapy [monoamine oxidase inhibitors and tricyclic antidepressants] showed no significant difference from the control group [P>0.05]. In cases of cancer breast, morning serum melatonin levels were significantly decreased in the early stages of the disease [P<0.0001], but became significantly elevated with progress of cancer and the occurrence of metastasis [P<0.0001]. Following radical mastectomy, the level of melatonin was insignificantly different from the control group [P >0.05]. Hence, we can conclude that decreased nocturnal melatonin could be considered an endogenous marker of major depressive illness, with such a decrease being masked by antidepressant therapy. Meanwhile, morning melatonin levels are of value in assessment and staging of breast cancer as well as post-operative follow-up of these patients. hopefully, aiming at early detection of recurrence

Improved resolution of liver and bone alkaline phosphatase isoenzymes using wheat germ lectin or neuraminidase pretreatment with the introduction of some technical modifications

This study aims to establish a routine reliable electrophoresis method with improved separation of liver and bone alkaline phosphatase isoenzymes, which allows for their quantitation. Modifications of the already available techniques will also be studied. Samples were collected from patients with liver diseases [n = 26], with bone diseases and from children [n = 24] and from pregnant females in the third trimester [n = 10]. Control sera containing liver and intestinal isoenzymes were also used. Samples were subjected to liver function tests, calcium and phosphorus determination, cellulose acetate electrophoresis : ordinary, with germ wheat lectin and with neuraminidase pretreatment. Agarose gel electrophoresis was done with and without lectin. Samples were also subjected to sequential heat inactivation. Results showed that cellulose acetate electro-phoresis gave better separation of liver and bone fractions when done with germ wheat lectin or when samples were pretreated with neuraminidase. Several modifications were suggested to improve the technique. Agarose gel affinity electrophoresis [i.e., with lectin] gave the best separation of liver and bone isoenzymes into sharply defined bands. Sequential heat inactivation was tedious and needed scrupulous control of time and temperature. It overestimated the liver isoenzyme due to inclusion of biliary and intestinal fractions in its estimation. Excellent correlation was found between the different methods used for both bone and liver isoenzymes, Biliary isoenzyme was best separated by ordinary electrophoresis whether on cellulose acetate or agarose gel. Placental isoenzyme separation required preheating the sample at 65°C for 10 minutes to destroy the bone fraction which had the same migration mobility as placental isoenzyme. It was concluded that agarose affinity gel electrophoresis gave the sharpest and clearest separation of liver and bone fractions. On the other hand, cellulose acetate electrophoresis was less expensive, more sensitive and precise. Both methods were more suitable than the heat separation analysis method

Comparison between two methods for measuring serum low density lipoprotein cholesterol

Two methods for low density lipoprotein cholesterol [LDL-C]. estimation were compared at different triglycerides [TG] levels. These were the quantitative determination by Helena cellulose acetate electrophoresis methods and the indirect calculation using Friedwald formula. Eighty fasting samples were selected and classified into 4 groups according to their TG levels. LDL-C was determined in each sample by both methods. Strongly significant correlation coefficients, between the 2 methods were obtained at normal, slightly, and moderately increased TG levels [r = 0.99, 0.97 and 0.94 respectively]. At TG levels >350 mg/dl the correlation became much weaker [r =0.77]. Thirty samples at different TG levels, were examined in duplicate for accuracy and precision. The electrophoretic quantitation of LDL-C was, much more precise [CV = 2.6%] than the indirectly calculated one. The latter depends on the imprecision of methods used in measuring total cholesterol, TG and HDL-C by precipitation method [CV = 2.7%, 3.8% and 8.5% respectively]. Paired t test showed no significant difference between LDL-C values obtained by both methods [P>0.05] However, in all cases of hypertriglyceridemia, the calculated LDL-C values showed constant insignificant lower values [6 - 12% lower] which might be sufficiently erroneous to cause misinterpretation of hypercholesterolemia at the critical LDL-C levels when medical interventions required. We recommended using the Fridewald formula as a screening test for LDL-C estimation and the electrophoresis method at TG levels> 350 mg/dl, LDL-C levels> 200 mg/dl or when abnormal lipoproteins are present

Evaluation of the O-cresolphthalein complexone procedure in comparison with the atomic absorption spectrophotometry for serum calcium determination

The automated 0.cresophthalein complexone dye method was recently introduced in the calcium chemistry module of the ASTRA-8 system present in Ain Shams University Hospital. This method was evaluated in comparison with the atomic absorption spectrophotometer reference method. Quality control material [normal and pathological levels] and 40 patient samples were used in the evaluation study. The coefficients of variation for the within batch precision were 1.5% and 1.1%, while those for the day to day precision were 3.2% and 2.3% for the normal and pathological levels respectively. Accuracy study showed minimal bias [0.12 mg/dl] and no statistically significant difference was observed between the two methods. Recovery was excellent. Linearity was 4.5 - 13 mg/ dl. No interference was detected by magnesium, while haemoglobin at a level of 1 g/dl showed a decrease in calcium concentration with an average of 0.6 mg/dl. The throughput was 70 samples/ hour, Very small sample size [12 micro l] or reagent [150 micro l] was required. There was good correlation between the ASTRA calcium method and atomic absorption spectrophotometry [Y = 1.97 + 0.8 X r=0.92]. The goals of analytical reliability, simplicity and convenience were nearly achieved by the -ASTRA-8 calcium method