Histamine assay

Muchowski recently reported -ll-deoxy-lla,12a-difuoro-methylene-PGE2 (HI) to have 5x the potency of I-PGE2 in a guinea pig assay (histamine agonist) when administered by aerosol, and to be equipotent when administered by the intravenous route (126,127). The 13,14-dihydro derivative IV was similarly active, as were the and Eq counterparts. Interestingly, the lla,12a-methylene analogs were essentially inactive. In a clinical trial with mildly asthmatic patients neither III or IV were sufficiently effective to be of interest (126). 

Histamine in the Blood. After its release, histamine diffuses rapidly into the blood stream and surrounding tissues (12). Histamine appears in blood within 2.5 min after its release, peaks at 5 min, and returns to baseline levels by 15 to 30 min. In humans, the diurnal mean of plasma histamine levels is 0.13 ng/g. In urine, elevations of histamine or metaboUtes are more prolonged than plasma elevations. Consequendy, abnormahties are more easily detected by urinary histamine assay. About one-half of the histamine in normal blood is in basophils, one-third in eosinophils, and one-seventh in neutrophils the remainder is distributed among all the other blood components. Increases in blood histamine levels occur in several pathological... 

Histamine concentrations are maximal almost immediately, decrease thereafter with a half-life of about 20 min, and should be assayed within the first hour of a reaction. The sensitivity of this test for the diagnosis of anaphylaxis was estimated at 75%, the specificity at 51%, the positive predictive value at 75% and the negative predictive value at 51%. Tryptase reaches a peak in the patient s serum 30 min after the first clinical manifestations. Its half-life is 90 min, and the levels usually decrease over time. In a recent series, the sensitivity was estimated at 64%, specificity at 89.3%, positive predictive value at 92-95%, and negative predictive value at 54.3% [9]. 

Black, J. W., Leff, R, and Shankley, N. R, Further analysis of anomalous pXj values for histamine H2-receptor antagonists on the mouse isolated stomach assay, Br. J. Pharmacol., 86, 581-587, 1985. 

Figure 4.7. Histamine release from mast cells in response to various dilutions of HRA generated from bovine serum albumin (BSA) by medium derived from stimulated rat neutrophils [156]. Neutrophils ((50-100) x K lml) were stimulated with FMLP (10 5 M), the medium removed and incubated with BSA (10 mg I ml) at pH 4.5 for 18 h at 37° C. It was then boiled and centrifuged (11,000 x g for 30 s), the supernatant fraction was removed, its pH was adjusted to 7.2 and it was added, at various dilutions, to suspensions of mast cells. Histamine release was then measured after 10 min. As the amount of generated HRA was increased histamine release increased to a maximum at 57 4%. Mean + S.E.M., n = 5. Inset Time-course of generation of HRA as assayed by histamine release from isolated mast cells. HRA was generated as before using 50 x 106 neutrophils/ml. Aliquots were removed at the indicated times and assayed (at 50% dilution) for HRA. Note that there is a significant generation of HRA by 2 h. Mean S.E.M., n = 3.

Castillo, J.C. and De Beer, E.J. (1947a). The guinea pig tracheal chain as an assay for histamine agonists. Fed. Proc. 6 315. 

Theory The gravimetric assay of histamine acid phosphate is based upon the formation of insoluble histamine-nitranilic acid complex as depicted in the following equation ... 

Biogenic amines are of great interest to researchers because of their potential roles in several psychiatric and neurological disorders. They include dopamine (DA), noradrenaline (NA), 5-hydroxytryptamine (5-HT, serotonin), histamine, and trace amines such as 2-phenylethylamine (PEA), tyramine, octopamine, phenylethanolamine, and tryptamine (Coutts and Baker, 1982). Although GC assays for DA, NA, and 5-HT are available, HPLC analysis with electrochemical detection has for many years now been the method of choice for analysis of these neurotransmitter amines. 

Due to technical difficulties in detecting vivo histamine release in humans, in vivo animal experiments or in vitro assays are often used. 

Histamine release from pig lung occurs in response to extracts from cotton pericarp, seed, leaf, root and bract (112). Extracts of sisal release histamine from both pig and human lung, but the reactivity is less than cotton dust extracts. The use of chopped lung for histamine assay is not always reliable, however, nor is it sufficiently sensitive. 

Pig platelets have recently been shown to release histamine when exposed to cotton dust or plant extracts (115) and this assay compares closely with chopped lung assays. Mill dust and gin trash extracts give some release and extracts of leaves from other plant sources, including pecan and grape, give similar reactions. Byssinosan (116). an aminopolysaccharide isolated from cotton dust, and THF antigen (83) are relatively inactive. 

The platelet hist UIline release assay demonstrated that cotton mill dust extract, cotton bract extract, cotton leaf extract, dialyzed CMD extract, polyphenols, compound 48/80, rutin, trimethylamine HCl, quercetin, catechin, tannic acid, ellagic acid and sodium metasilicate all release histamine directly (48). Thus not only do tannin compounds induce histamine release, but they may also form higher molecular weight polymers and contain components that survive acid hydrolytic conditions (48). Tannins are widely distributed in the plant kingdom. 

The histamine- and leukotriene B4-releasing potential of the RCBPA P743 has been investigated in vitro on the RBL-2H3 mast cell line [25]. RBL-2H3 cells (with permission of Dr RP Siraganian, National Institutes of Health, Bethesda, MD, USA) were replated on Eagle s minimum essential medium and incubated for 30 minutes at 37 °C in 5% CO2 with the test-solutions. Calcium ionophore A23187 was used as a positive control. The viability of mast cells was determined by means of a quantitative colorimetric assay that is based on the abiUty of viable cells to cleave the reagent MTT [33]. 

Crude and three diethyl ether extracted, acetone treated, fractions were isolated from large-scale cultures of Gambierdiscus toxicus. Crude extracts at. 04 mg/ml inhibited the histamine contraction response in smooth muscle of the guinea pig ileum. Three semi-purified fractions at 5 ng/ml, effectively inhibited the guinea pig ileum preparation. Two of these fractions followed Michaelis-Menten kinetics for a competitive inhibition. The third fraction inhibited in a non-reversible manner. This study has established the presence of three lipid extracted toxins in toxicus, outlined a method for their assay in small quantities, and identified at least two of the effects of these toxic extracts in animals. 

In summary, we have isolated three toxins from lipid extracts of tqxicus. We have demonstrated that the guinea pig ileum preparation is an effective assay for these toxins in nanogram quantities. All three fractions effectively inhibited the guinea pig ileum preparation in its response to histamine at nanogram concentrations. The GT-3 toxic fraction is quite different from the other two (GT-1, GT-2) in being very slow acting and nonreversible in nature. 

Bacterial Growth and Histamine Formation. The quantitative relationship between histamine formation and the microbial flora in skipjack tuna at 38 C is shown in Figure 3. At intervals during incubation samples were removed from the second section and assayed for histamine content and bacterial numbers. After 24 h the anaerobic bacterial count was 3.5 x 10 per g, and the histamine content was 297 mg per 100 g. Anaerobic counts were used to measure the microbial population because over 92% of the bacteria found in decomposed skipjack tuna were obligate or facultative anaerobes (Table I). 

Fig. 1.1 Interaction between allergen and adenosine receptor agonists with respect to histamine release from guinea-pig chopped lung (left) or bronchoconstriction in the guinea pig in vivo (right). Tissues and animals were passively sensitized to ovalbumin. In the histamine release assay, a threshold response to allergen is augmented concentration-dependently by NECA. In the whole animal, a single intravenous injection of APNEA markedly enhances the bronchoconstrictor response to allergen (J.R. Fozard and H.J. Pfannkuche, unpublished observations 1994)...

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