Determination of 6-mercaptopurine and azathioprine in plasma by high-performance liquid chromatography

Using 1-ml plasma samples, levels of 6-mercaptopurine (6MP) as low as 5 ng/ml and azathioprine (AZA) as low as 40 ng/ml can be detected using a high-performance liquid chromatography reversed-phase column procedure following extraction. Both compounds were stable in frozen plasma for seven weeks. AZA stability in blood was temperature dependent; the half-lives of AZA breakdown to 6MP at 37° were 28 and 46 min in blood drawn from two rhesus monkeys. Plasma levels of 6MP were measured in a rhesus monkey following 6MP (1.47 mg/kg) and AZA (3 mg/kg) intravenous administration. 6MP levels were also measured in three renal transplant patients on daily 50- and 100-mg AZA doses. Peak levels (45–75 ng/ml) were reached within an hour and 6MP levels were detected for up to 7 h.     

Assessment of Mercaptopurine (6MP) Metabolites and 6MP Metabolic Key-Enzymes in Childhood Acute Lymphoblastic Leukemia

Pediatric acute lymphoblastic leukemia (ALL) is treated with combination chemotherapy including mercaptopurine (6MP) as an important component. Upon its uptake, 6MP undergoes a complex metabolism involving many enzymes and active products. The prognostic value of all the factors engaged in this pathway still remains unclear. This study attempted to determine which components of 6MP metabolism in leukemic blasts and red blood cells are important for 6MP's sensitivity and toxicity. In addition, changes in the enzymatic activities and metabolite levels during the treatment were analyzed. In a cohort (N = 236) of pediatric ALL patients enrolled in the Dutch ALL-9 protocol, we studied the enzymes inosine-5′-monophosphate dehydrogenase (IMPDH), thiopurine S-methyltransferase (TPMT), hypoxanthine guanine phosphoribosyl transferase (HGPRT), and purine nucleoside phosphorylase (PNP) as well as thioguanine nucleotides (TGN) and methylthioinosine nucleotides (meTINs). Activities of selected enzymes and levels of 6MP derivatives were measured at various time points during the course of therapy. The data obtained and the toxicity related parameters available for these patients were correlated with each other. We found several interesting relations, including high concentrations of two active forms of 6MP—TGN and meTIN—showing a trend toward association with better in vitro antileukemic effect of 6MP. High concentrations of TGN and elevated activity of HGPRT were found to be significantly associated with grade III/IV leucopenia. However, a lot of data of enzymatic activities and metabolite concentrations as well as clinical toxicity were missing, thereby limiting the number of assessed relations. Therefore, although a complex study of 6MP metabolism in ALL patients is feasible, it warrants more robust and strict data collection in order to be able to draw more reliable conclusions.     

Deformability of stored erythrocytes as a criteria for their in vivo survival rate

The loss of deformability observed in erythrocytes stored as whole blood for 36 days (ACD-AG) or as buffy-coat free erythrocyte concentrate (EK) was characterized by measuring their filterability. During the first 3 weeks the index of filterability for ACD-AG erythrocytes increased only slightly and rose to about 140% of its initial value on the 36th day. In contrast, a heavy loss of deformability (increase of the filterability index to more than 600%) was detected for erythrocytes from EK, which, from a rheological point of view, is apt to raise doubts of using this stored blood. An incubation of 1 hour at 37 degrees C in fresh plasma did not result in improving the deformability. A cell volume loss of more than 20% connected with an increase of the inner viscosity to more than 400% was found to be the cause of this decrease of deformability. These rheological differences are also reflected in the 24 hours in vivo survival rate (SR), if the "early loss" of damaged erythrocytes immediately after transfusion is taken into account. Whereas the SR values of 80% for whole blood erythrocytes do not change significantly, the SR values for EK values can be found to reach 54% approximately.     

Quantification of naltrexone and 6,β-naltrexol in plasma and milk using gas chromatography–mass spectrometry: Application to studies in the lactating sheep

A selective gas chromatography–mass spectrometry method using solid-phase extraction has been developed for the detection and quantification of naltrexone and its metabolite, 6,β-naltrexol in plasma and milk from humans and sheep at pharmacologically relevant concentrations. Di- or tri-acetyl derivatives were formed and quantified by selected-ion monitoring. Recoveries of naltrexone (30 μg/l) and 6,β-naltrexol (250 μg/l) from both human plasma and milk were greater than 70%. Intra-assay and inter-day precision ranged from 3 to 21% for naltrexone and 2–18% for 6,β-naltrexol for all matrices investigated, with an overall mean accuracy of 104% for naltrexone, and 99% for 6,β-naltrexol. Human samples containing these analytes were stable for at least 3 weeks at −20°C or 6 weeks at −80°C. Analysis of the plasma and milk from the lactating sheep showed mean milk-to-plasma ratios of 55 for naltrexone and 3 for 6,β-naltrexol.     

Effects of storage conditions on the stability and distribution of clinical trace elements in whole blood and plasma: Application of ICP-MS

BackgroundKnowledge of trace element stability during sample handling and preservation is a prerequisite to produce reliable test results in clinical trace element analysis.MethodAn alkaline dissolution method has been developed using inductively coupled plasma mass spectrometry to quantify eighteen trace element concentrations: vanadium, chromium, manganese, cobalt, nickel, copper, zinc, arsenic, selenium, bromine, molybdenum, cadmium, antimony, iodine, mercury, thallium, lead, and bismuth in human blood, using a small sample volume of 0.1 mL. The study evaluated the comparative effects of storage conditions on the stability of nutritionally essential and non-essential elements in human blood and plasma samples stored at three different temperatures (4 °C, −20 °C and −80 °C) over a one-year period, and analysed at multiple time points. The distribution of these elements between whole blood and plasma and their distribution relationships are illustrated using blood samples from 66 adult donors in Queensland.ResultsThe refrigeration and freezing of blood and plasma specimens proved to be suitable storage conditions for many of the trace elements for periods up to six months, with essentially unchanged concentrations. Substantially consistent recoveries were obtained by preserving specimens at −20 °C for up to one year. Ultra-freezing of the specimens at −80 °C did not improve stability; but appeared to result in adsorption and/or precipitation of some elements, accompanied by a longer sample thawing time. A population sample study revealed significant differences between the blood and plasma concentrations of six essential elements and their relationships also varied significantly for different elements.ConclusionBlood and plasma specimens can be reliably stored at 4 °C for six months or kept frozen at −20 °C up to one year to obtain high quality test results of trace elements.     

Developmental changes of 6-phosphofructo-1-kinase subunit levels in erythrocytes from normal dogs and dogs affected by glycogen storage disease type VII.

1. The subunit proportions (L:M:C) of the PFK isozymes from normal adult erythrocytes were 2:86:12. Affected adult erythrocyte 6-phosphofructo-1-kinase (PFK) isozymes contained normal L-type (31%) and C-type (61%) subunits as well as a small amount (8%) of truncated M-type subunit. 2. When measured within 24 hr of birth, both normal and affected dog erythrocytes contained high PFK activities due to elevated levels of the L-type subunit. As the dogs matured, PFK activity decreased due to a greater than 99% loss of the L-type subunit. 3. By 2 weeks of age, the M-type and C-type subunits in normal dog PFK isozymes increased several-fold and attained near adult levels. 4. During post-natal development, the L-type subunit from affected dog erythrocytes decreased more rapidly than from normal dog erythrocytes; but it was maintained at a higher level in the affected adult erythrocytes. Also, in the affected dog erythrocytes, truncated M-type subunits were detected; and the initially high levels of the C-type subunit decreased approximately 50% after 4 weeks.     

Vitamin C in mouse and human red blood cells: an HPLC assay

Although vitamin C (ascorbate) is present in whole blood, measurements in red blood cells (RBCs) are problematic because of interference, instability, limited sensitivity, and sample volume requirements. We describe a new technique using HPLC with coulometric electrochemical detection for ascorbate measurement in RBCs of humans, wild-type mice, and mice unable to synthesize ascorbate. Exogenously added ascorbate was fully recovered even when endogenous RBC ascorbate was below the detection threshold (25 nM). Twenty microliters of whole blood or 10 μl of packed RBCs was sufficient for assay. RBC ascorbate was stable for 24h from whole-blood samples at 4°C. Processed, stored samples were stable for >1 month at -80°C. Unlike other tissues, ascorbate concentrations in human and mouse RBCs were linear in relation to plasma concentrations (R=0.8 and 0.9, respectively). In healthy humans, RBC ascorbate concentrations were 9-57 μM, corresponding to ascorbate plasma concentrations of 15-90 μM. Mouse data were similar. In human blood stored as if for transfusion, initial RBC ascorbate concentrations varied approximately sevenfold and decreased 50% after 6 weeks of storage under clinical conditions. With this assay, it becomes possible for the first time to characterize ascorbate function in relation to endogenous concentrations in RBCs.     

Morphology of glutaraldehyde-fixed preserved red blood cells and 24-hr post-transfusion survival

Liquid-stored red blood cells and washed, previously frozen red blood cells were studied to determine whether a correlation existed between morphology and post-transfusion survival. Red cell concentrates were stored at 4 °C in citrate-phosphate-dextrose (CPD) for 21 days or in CPD-adenine (CPDA-1, CPDA-2, or CPDA-3) for as long as 35 days as liquid-preserved red cells. Both nonrejuvenated and rejuvenated red blood cells were frozen with 40%$\text{w}\text{v}$ glycerol at ?80 °C and were washed prior to testing.Samples of fresh, liquid-stored, and washed, previously frozen red blood cells were fixed with a 2% veronal glutaraldehyde solution. Phase, light, and electron microscopy were used to measure the numbers of discocytes, discoechinocytes, echinocytes, echinospherocytes, and spherocytes in each sample. A morphology score was assigned, with 100 representing all discocytes and 500 all spherocytes. In all samples phase and light microscopy gave nearly identical scores (r = 0.94), and phase and electron microscopy gave highly similar scores (r = 0.83).The morphology score proved to be a good indicator of 24-hr post-transfusion survival in liquid-stored red blood cells but not in washed, previously frozen red blood cells. Red blood cells stored in the liquid state at 4 °C in CPD, CPDA-1, CPDA-2, or CPDA-3 showed a significant inverse correlation between morphology and 24-hr post-transfusion survival (r = ?0.611) and a significant correlation between red cell ATP and 24-hr post-transfusion survival (r = 0.742). We saw no significant correlation between morphology scores and 24-hr post-transfusion values or between ATP levels and post-transfusion survival values in nonrejuvenated or rejuvenated washed, previously frozen red blood cells.     

Refrigeration of blood samples prior to separation is essential for the accurate determination of plasma or serum zinc concentrations

An evaluation of refrigeration (7°C) to prevent falsely high plasma or serum zinc concentrations owing to elapsed time between blood collection and centrifugation was performed. At room temperature (23°C), both plasma and serum zinc concentrations increased significantly, if blood samples were stored uncentrifuged. Plasma zinc concentrations increased 6.3% at 1 h and 40.7% at 24 h, whereas serum zinc concentrations increased only 0.9% at 1 h and 12.5% at 24 h at room temperature. When blood samples were stored uncentrifuged in the refrigerator for up to 24 h, there were no significant increases in zinc concentrations in either plasma or serum. These findings suggest that plasma or serum separation should be performed immediately after blood drawing to obtain accurate zinc concentrations, and if this is not feasible, the samples should be immediately refrigerated and separation performed within eight hours.     

Transport of ricin from endosomes to the Golgi apparatus is regulated by Rab6A and Rab6A'

Ricin is transported from early endosomes and/or the recycling compartment to the trans-Golgi network (TGN) and subsequently to the endoplasmic recticulum (ER) before it enters the cytosol and intoxicates cells. We have investigated the role of the Rab6 isoforms in retrograde transport of ricin using both oligo- and vector-based RNAi assays. Ricin transport to the TGN was inhibited by the depletion of Rab6A when the Rab6A messenger RNA (mRNA) levels were reduced by more than 40% and less than 75%. However, when Rab6A mRNA was reduced by more than 75% and Rab6A' mRNA was simultaneously up-regulated, the inhibition of ricin sulfation was abolished, indicating that the up-regulation of Rab6A' may compensate for the loss of Rab6A function. In addition, we found that a near complete depletion of Rab6A' gave approximately 40% reduction in ricin sulfation. The up-regulation of Rab6A mRNA levels did not seem to compensate for the loss of Rab6A' function. The depletion of both Rab6A and Rab6A' gave a stronger inhibition of ricin sulfation than what was observed knocking down the two isoforms separately. In conclusion, both Rab6A and Rab6A' seem to be involved in the transport of ricin from endosomes to the Golgi apparatus.     

Effects of Selenium Administration on Erythrocyte and Blood Plasma Glutathione Peroxidase Activity in Goats

The activity of glutathione peroxidase (GSH-Px, E.G. 1.11.1.9.) was determined in heparinized whole blood, blood plasma and washed erythrocytes from goats before and up to 4 weeks after the administration of selenium (0.4 mg/10 kg BW) and vitamin E (20 mg/10 kg BW) or only vit. E (20 mg/10 kg BW). It was found that Se administration caused a significant increase in enzyme activity in whole blood and washed erythrocytes first detected 2 weeks after the intramuscular injection of Se. No changes were observed in plasma from the treated animals. Minor and insignificant changes were seen in the vit. E treated control animals. It is concluded that GSH-Px activity in blood plasma or serum is of no value as a short-term indicator of the selenium status of goats but whole blood is a good indicator of the long-term status.     

High-performance liquid chromatographic assay for meropenem in serum

High-performance liquid chromatographic procedures have been developed for the measurement of meropenem in serum. The separation was performed on an Ultrasphere XL-ODS analytical column (75×4.6 mm I.D.). The mobile phase consisted of 10.53 mmol/l ammonium acetate-acetonitrile (95:5, v/v) (pH 4). The UV detection was at 298 nm. The quantitation limit both in serum and water was 0.25 μg/ml. The method was validated in serum and aqueous solution over the concentration range 0.25–50 μg/ml. The extraction recovery from serum spiked with meropenem was 99.7±3.4%. The intra- and inter-assay coefficients of variation were below 6%. Stored at −80°C for three months at various concentrations in serum and in aqueous solution, meropenem did not reveal any appreciable degradation. After 24 h, it was also stable at 4°C in serum, aqueous solution and supernatant of extraction but not at room temperature. The stability of the drug was also confirmed in serum after repeated freezing-thawing cycles at −80°C on four consecutive days.     

Density-gradient enrichment of newly-formed mouse erythrocytes. Application to the micronucleus test

To facilitate scoring micronuclei in peripheral blood erythrocytes, we have developed a centrifugation method to concentrate polychromatic and newly-formed normochromatic erythrocytes from microliter quantities of blood in a Percoll density gradient. Erythrocytes were separated into two discrete bands in a continuous gradient generated in situ in a microhematocrit capillary tube. The upper band contained white blood cells and a mixture of polychromatic and young normochromatic erythrocytes with a density of 1.080-1.082 g/ml. More than 75% of the polychromatic erythrocytes in samples of normal blood were recovered in the upper band. Older normochromatic erythrocytes migrated to the lower band. The frequency of polychromatic erythrocytes was increased from approximately 2% in whole blood to 60-80% in the upper band. After clastogen treatments, the elevated frequencies of micronuclei in the upper band polychromatic erythrocytes were similar to those in unfractionated blood. The frequencies of micronucleated normochromatic erythrocytes in the upper band were higher than those in whole blood at 48, 72 and 96 h after clastogen treatment, consistent with the expectation that the low-density normochromatic cells are newly derived from polychromatic erythrocytes. This density-gradient centrifugation technique enhances the efficiency of scoring micronuclei in the acute peripheral blood micronucleus test.     

Influence of gender, menstrual phase, and oral contraceptive use on immunological changes in response to prolonged cycling.

This study determined the influence of gender, menstrual phase (MP), and oral contraceptive (OC) use on immunological changes in response to endurance exercise. Twelve women and 11 men similar in age, aerobic power, and activity level cycled for 90 min at 65% maximal aerobic power. Women were OC users (n = 6) or nonusers (NOC) and cycled during the follicular (Fol) and the luteal (Lut) phases. Venous blood was collected before and after exercise to determine leukocyte counts, IL-6 concentrations, and cortisol. Higher resting levels of neutrophils (approximately 1.5-fold) and cortisol (approximately 2.5-fold) were found in OC vs. NOC and men. Exercise-induced immune cell count and IL-6 changes were similar between men and NOC, except for an approximately 38% greater lymphocyte response in NOC vs. men (P = 0.07). Neutrophil, monocyte, and lymphocyte responses to exercise during Lut in OC were greater than during Fol and also greater than the responses in men (P < or = 0.003). Changes in immune cell counts were consistently greater during Lut in OC vs. NOC, regardless of MP, but only neutrophil responses reached statistical significance (P = 0.01). The exercise-induced change in IL-6 was approximately 80% greater in NOC vs. OC during Fol (P = 0.06), but it was similar between these groups during Lut. Cortisol changes with exercise were not different between groups or MP. These results highlight the necessity to control for gender, and in particular OC use, when designing studies evaluating exercise and immunology.     

Responses of the bed bug,Cimex lectularius,to temperature extremes and dehydration: levels of tolerance,rapid cold hardening and expression of heat shock proteins

This study of the bed bug, Cimex lectularius, examines tolerance of adult females to extremes in temperature and loss of body water. Although the supercooling point (SCP) of the bed bugs was approximately −20°C, all were killed by a direct 1 h exposure to −16°C. Thus, this species cannot tolerate freezing and is killed at temperatures well above its SCP. Neither cold acclimation at 4°C for 2 weeks nor dehydration (15% loss of water content) enhanced cold tolerance. However, bed bugs have the capacity for rapid cold hardening, i.e. a 1‐h exposure to 0°C improved their subsequent tolerance of −14 and −16°C. In response to heat stress, fewer than 20% of the bugs survived a 1‐h exposure to 46°C, and nearly all were killed at 48°C. Dehydration, heat acclimation at 30°C for 2 weeks and rapid heat hardening at 37°C for 1 h all failed to improve heat tolerance. Expression of the mRNAs encoding two heat shock proteins (Hsps), Hsp70 and Hsp90, was elevated in response to heat stress, cold stress and during dehydration and rehydration. The response of Hsp90 was more pronounced than that of Hsp70 during dehydration and rehydration. Our results define the tolerance limits for bed bugs to these commonly encountered stresses of temperature and low humidity and indicate a role for Hsps in responding to these stresses.     

Fructose 2,6-bisphosphate in human erythrocytes

The fructose 2,6-bisphosphate concentrations in unwashed, washed, and leukocyte-free erythrocytes were compared. The concentration in washed red cells was 31 +/- 15 pmol per ml of cells (mean +/- S.D., n = 6). The concentration in unwashed erythrocytes was at least twofold higher, but the value in washed red cells was not due to leukocyte contamination because it did not decrease further when washed cells were passed through an Imgard column, which would have removed any remaining leukocytes. No platelets were detected among the washed erythrocytes. Thus, the concentration in erythrocytes after washing was ascribed solely to these cells. The fructose 2,6-bisphosphate concentration did not change when the glycolytic activity varied with pH, indicating that this compound is not involved in the regulation of carbohydrate metabolism in erythrocytes under these conditions.     

Effects of low-temperature acclimation on the survival and cold tolerance of an antarctic mite

The effects of long-term exposures to constant temperatures (+4, 0, −5, −10, −15 and −20°C) on the survival of a cryptostigmatid mite, Alaskozetes antarcticus, were studied during 1983–1984 on Signy Island, South Orkney Islands, in the maritime Antarctic. Field-fresh samples collected during the austral summer showed very large (e.g. about 60 percentage points) variations in survival when placed at constant temperatures, as a result of collection-date effects. Pretreatment acclimations (10-day) at +4 and 0°C (especially) reduced this variation. Short-term modulations in cold-hardiness levels were related to ambient temperature fluctuations. However, samples collected on the same occasion, from microhabitats 20 m apart, also showed significant cold-hardiness variation. For twelve summer samples, survival after 24 h at −15°C was highly correlated with supercooling capacity. Winter samples showed little variation in survival, interms of collection-date. Percentage survival remained greater than 85% at −5, −10 and −15°C, for exposures up to 100 days. Samples with median supercooling points of about −30°C, showed 52% survival after 250 days at −15°C, and 73% survival after 100 days at −20°C. At −15°C, supercooling capacity was used up at an estimated rate of 0.06 deg day⁻¹, as a result of a time-temperature interaction effect on the probability of heterogeneous nucleation. Adult mites showed 78% survival after 21 days encasement in distilled water ice, at −15°C. Survival differences between post-larval stages were not detected. In conclusion, survival ability under controlled laboratory conditions appeared to exceed the requirements of average winter-habitat temperatures, but the effects of fluctuating and extreme temperatures require investigation. Supercooling points are considered to be accurate indicators of low-temperature survival capability in this species.     

Urinary cortisol analysis for monitoring adrenal activity in elephants

Cortisol was measured in dichloromethane-extracted elephant urine using an ¹²⁵I solid-phase radioimmunoassay (RIA). The cortisol RIA was validated by demonstrating 1) parallelism between dilutions of pooled urinary extracts and the standard curve, 2) significant recovery of exogenous cortisol added to elephant urine, and 3) a relationship between changes in peripheral and urinary cortisol after an adrenocorticotropin hormone (ACTH) challenge. One African (Loxodonta africana) and one Asian (Elephas maximus) elephant were given three injections of ACTH (1.25 mg) at 2 h intervals. Serum cortisol increased four- to eightfold within 30 min after the first injection and peaked (nine- to twelvefold increase) after the second injection. Serum concentrations began to decline 2–3 h after the last injection but were still approximately fourfold higher than baseline at the end of the collection period (hour 8). In the urine, cortisol concentrations were increased in the first sample postinjection (1.5–4 h) and peaked twenty- to fortyfold by ～6 h. Urinary cortisol remained elevated at 8 h, but returned to baseline the following morning. Analysis of high performance liquid chromatography fractions of extracted urine revealed that immunoactivity was associated with free cortisol (～90% of total immunoactivity) and a more polar, unidentified metabolite. A method for preserving urine was developed to allow storing unfrozen samples. One pool of urine from each of one African and two Asian elephants was divided into aliquots, placed in tubes containing absolute ethanol (10%), sodium azide (0.1%) or distilled water (control), and frozen after 0, 1, 2, 3, 4, 6, 8, 10, 12, and 24 weeks of storage at ～25°C. In unpreserved samples, cortisol concentrations were reduced 46% by 2 weeks and 95% by 24 weeks. In contrast, ethanol- and sodium azide-preserved samples retained 100 and 95% of cortisol immunoactivity through 8 weeks and 93 and 85% of activity through 12 weeks, respectively. We infer from these data that changes in urinary cortisol excretion in the elephant reflect fluctuations in adrenal activity and may be a useful indicator of stress. Additionally, urine samples can be collected and stored unfrozen for at least 2 months before any appreciable loss in cortisol immunoactivity occurs, a finding potentially useful to field application of this technique. © 1995 Wiley-Liss, Inc.

1 This article is a US Government work and, as such, is in the public domain in the United States of America

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Studies with dextran 40 in cryopreservation of blood

Whole blood from healthy donors was washed twice with phosphate-buffered saline (PBS) and then resuspended in sufficient PBS to give a final concentration of 2 × 10⁹/cells/ml. Aliquots were combined with equal volumes of the required diluents to give final dextran 40 concentrations of 0, 5, 10, 15, and 20% in PBS. Fifty-lambda samples in 50-lambda Micropets (Clay Adams) were frozen in alcohol baths at temperatures ranging from ?10 to ?80 °C. The specimens were frozen either for 1 min or 16 min, rapidly thawed, and resuspended in PBS or PBS plus dextran. Percentage of hemolysis was determined colorimetrically. Results indicate that concentraitons as low as 5% dextran exert a cryoprotective effect. Increased dextran concentration increases cryoprotection at high subzero bath temperatures (?10 ° and ?20 °C). Dextran concentrations beyond 12% have a damaging effect at low subzero bath temperatures (below ?30 °C). Based on this a two-factor hypothesis for cryopreservation is proposed. Apparent partial recovery of red blood cells without dextran or with 5% dextran during subzero storage was demonstrated.     

Effective low-cost preservation of human stools in field-based studies for helminth and microbiota analysis

Molecular studies of gastrointestinal infections or microbiotas require either rapid sample processing or effective interim preservation. This is difficult in remote settings in low-income countries, where the majority of the global infectious disease burden exists. Processing or freezing of samples immediately upon collection is often not feasible and the cost of commercial preservatives is prohibitive. We compared fresh freezing (the ‘gold standard’ method), with low-cost chemical preservation in (i) a salt-based buffer consisting of DMSO, EDTA and NaCl (DESS) or (ii) 2.5% potassium dichromate (PD), for soil-transmitted helminth detection and microbiota characterisation in pre-school and school-aged children from north-western Thailand. Fresh frozen samples were frozen at −20°C on collection and maintained at −80°C within ~3 days of collection until molecular analysis, with international shipping on dry ice. In contrast, chemically preserved samples were collected and stored at ~4°C, transported on wet ice and only stored at −20°C on arrival in Australia ~8 weeks after collection, with international shipping on wet ice. DESS and PD provided better sensitivity for STH diagnosis, estimating higher infection rates (>80% for Ascaris lumbricoides and >60% for Trichuris trichiura; versus 56% and 15% for these parasites in fresh frozen samples) and egg abundance (inferred as gene copy number estimates). All methods performed similarly for microbiota preservation, showing no significant differences in alpha-diversity based on overall richness or inverted Simpson’s Index. All three methods performed similarly for RNA and protein preservation in a small subset of samples. Overall, DESS provided the best performance, with the added benefit of being non-toxic, compared with PD, hence making it particularly applicable for studies in remote and resource-poor settings.