Simultaneous measurement of whole blood vitamin B1 and vitamin B6 using LC-ESI-MS/MS.

Vitamin B1 and B6 have recently been included in the Dutch clinical guidelines for the general practitioner in the differential diagnosis of dementia. To keep up with the sharp rise in the number of requests, an LC-MS/MS method using stable isotopes as internal standards was developed. The active vitamers thiamine pyrophosphate (TPP) and pyridoxal-5'-phosphate (PLP) in whole blood are simultaneously measured with a short run time of 2min. Whole blood is mixed with internal standard solution containing both TPP-d3 and PLP-d3, followed by deproteinization with a trichloroacetic acid (TCA) solution. A UPLC-MS/MS system from Waters™ was used for chromatographic separation and subsequent detection by electrospray ionization in the positive mode with mass transitions of 425.1>121.85 for TPP and 247.9>149.9 for PLP. The method is linear across the range of 12-4870 nmol/L for TPP and 6-4850 nmol/L for PLP. The mean intra-assay and inter-assay precision are 3.5% and 7.6% respectively for TPP and 3.4% and 6.1% for PLP. The relative matrix effect (TPP 97%, PLP 93%), recovery (TPP 99%, PLP 94%) and lower limit of quantification (TPP 12 nmol/L, PLP 6 nmol/L) meet the applied acceptance criteria. The comparison of the new LC-ESI-MS/MS method for TPP with our current HPLC-Fluorescence method for total thiamine yields the following equation: TPP LC-MS/MS=0.97×total thiamine HPLC - 10.61 (r2=0.94). The comparison of the new LC-ESI-MS/MS method for PLP with our current LC-ESI-MS/MS method results in PLP LC-MS/MS new=1.01×PLP LC-MS/MS old - 1.58 (r2=0.99). In conclusion, this LC-MS/MS based assay is characterized by simple sample processing with a short run time and comparison with the current methods is excellent. The new LC-MS/MS method is a convenient method to determine TPP and PLP in whole blood for both clinical routine and research applications.

Estimation of severity of illness with APACHE II: age-related implications in cardiac arrest outcomes.

The ability to predict outcomes of cardiac arrest before starting cardiopulmonary resuscitation (CPR) would be useful for discussions of resuscitation with elders and their families. We thought CPR outcome might be dependent on the severity of pre-existing illnesses. The APACHE II is a severity-of-illness (SOI) scale based, in part, on physiologic parameters whereby points are given for degree of deviation from normal. Additionally, up to six points are given for increased age. We hypothesized that (1) patients with the highest APACHE II would be least likely to survive, and (2) because of the blunted physiologic responsiveness, the APACHE II would underestimate the SOI of elderly patients who were sufficiently ill to have a cardiac arrest. A retrospective study of 172 arrests was carried out to evaluate these hypotheses. For the young cohort (n = 126; age, < 70; mean age, 59 +/- 8), mean admission APACHE II was 16.5 +/- 7.9 and pre-arrest APACHE II regression analysis.2+ carried out with both APACHE II scores and factors previously correlated with CPR outcome. Witnessed arrests and those requiring a low number of medications were most likely to result in immediate success (restoration of blood pressure) and in a live discharge. APACHE II score (24 h pre-arrest) was associated with live discharge in the regression analysis.(ABSTRACT TRUNCATED AT 250 WORDS)

Measurement of stress-strain relationship and stress relaxation in various synthetic ligaments.

In an experimental study various synthetic augmentation devices for knee ligament surgery were tested in a servo-mechanical universal tensile testing machine under uniaxial loading. Two tests were done to elucidate the mechanical behaviour: stress relaxation and stress-strain relationship. Regarding the point of failure or rupture, the strongest ligament was the Trevira at 1800 N, followed by the 8-mm-wide Kennedy LAD at 1720 N. At a working load of 500 N the Gore-tex band, the Trevira, and the Kennedy-LAD stretched by between 2% and 3%. For synthetic augmentation in repair of proximally ruptured anterior cruciate ligaments we recommend a synthetic ligament that reaches failure point at a load of more than 1000 N with an alteration in length of less than 5%. Otherwise, stress protection of the biological reconstruction in full extension will be impossible. The requisite criteria were fulfilled by the Trevira, Kennedy-LAD and Gore-tex synthetic ligaments.

Pregnancy complicated by psittacosis acquired from sheep.

Two cases of chlamydial infection in pregnant women are described, the first serologically proved and the second suspected. In both cases the infection was probably contracted from sheep suffering with enzootic abortion. Both patients were farmers' wives who had helped their husbands and lambing and developed a non-specific febrile illness in late pregnancy. In the first case as there was no clinical improvement after 26 hours the patient was delivered by caesarean section of a live infant in good condition; the patient recovered fully. The second patient had presented a year earlier, the fetus had died in the uterus, and the patient himself died after spontaneous labour and forceps delivery 14 hours after admission. Both patients developed disseminated intravascular coagulation. As the casual agent in enzootic abortion in ewes has a predilection for the placenta, early delivery may be the management of choice in late pregnancy if infection with this organism if suspected.

The treatment of soil infested with the human whipworm, Trichuris trichiura.

The soil fumigants methyl bromide, chloropircrin and Basamid or mixtures of Basamid and chloropicrin proved to be of no use in filling T. trichiura ova in soil. However, on untreated plots the eggs died at a rate such than only 20% of the ova remained viable after 18 months.

The relationship between Trichuris trichiura (Linnaeus 1758) of man and Trichuris suis (Schrank 1788) of the pig.

Studies were made on the relationship between Trichuris suis of the pig and T trichiura of man. Comparative morphological and developmental studies revealed differences in sizes of the eggs and infective first stage larvae, in the rate of development to the infective larval stage and in development within young pigs following experimental infection. Successful cross infections of man with T suis ova and of pigs with T trichiura ova were achieved for the first time. T suis eggs recovered from one human experimental infection were used to produce a patent infection in a pig. It is argued that while the successful cross infections suggest a close evolutionary link, the differences in morphology, exogenous and endogenous development provide sufficient criteria for the two parasites to be maintained as separate species.

Synergism between Trichuris suis and the microbial flora of the large intestine causing dysentery in pigs.

The role of the microbial flora of the large intestine in experimental Trichuris suis infection was studied by comparing the clinical syndrome in conventionally reared (CR) pigs, specific pathogen-free pigs, and gnotobiotic pigs. Thedisease in CR pigs was characterized by a severe mucohemorrhagic enteritis; in contrast, a mild catarrhal enteritis was observed in specific pathogen-free and gnotobiotic pigs. Spirochaetes and vibrio-like organisms were observed only in CR pigs and increased during the clinical phase of the disease. The clinical syndrome was not transmitted by oral administration of intestinal or fecal material from infected CR pigs to CR pigs free of T. suis. Smaller numbers of T. suis produced diarrhea in CR pigs and significantly reduced the growth rates of infected animals; clinical signs and the reduction in growth rate was prevented by incorporating an antibacterial substance (dimetridazole) in the food. Although clinical trichuriasis closely resembles swin dysentery, the two syndromes seem to be distinct. The present results suggest that a microbial component acts synergistically with T. suis to produce the severe clinical syndrome in CR pigs, but identification of the microbial component and the mechanism by which clinical signs are produced await further studies of the bacterial flora of the large intestine of pigs.