Clarification and lessons learned for reporting studies with hydrates. Citation: Roberts et al., 2016. Toxicology Reports 3: 531–538

Abstract

To the Editor, We would like to submit clarifications for the recent article Roberts et al. (2016) and offer suggestions for addressing some of the challenges of reporting studies with test materials that are available in anhydrous and hydrated forms. We recently received a public inquiry pointing out that there is an inconsistency in the description of the vanadyl sulfate test material in this article compared to the description provided by the manufacturer (http://noahtech.com/products/15922). The manufacturer identifies the material as vanadyl sulfate hydrate, with an unspecified number of water molecules, while providing the CAS Registry Number and molecular weight for the anhydrous form. Based on our chemical analysis of the test material in October 2011 (prior to study conduct) and again in April 2017 when this issue was brought to our attention, the water content is approximately 33%. While we realized that the molecular weight for the vanadyl sulfate hydrate test material would need to be adjusted for water content, we failed to take this into account when formulating the drinking water solutions used in the 14 day toxicity studies. The impact of this oversight on our published article is that the concentrations of the vanadyl sulfate drinking water formulations used in the study (0, 125, 250, 500, 1000 and 2000 mg/L) actually contain 0, 83.8, 167.5, 335, 670, and 1340 mg/L vanadyl sulfate, respectively, when adjusted for water content. The only impact this revision has on the data presented and its interpretation is on the calculation of chemical and vanadium consumption presented in Table 3A, Table 3B. Updated tables are provided here: Table 3A Chemical Consumption (mg/kg/day): Rats. Table 3A Sodium Metavanadate Vanadyl Sulfate mg/L ♂ ♀ mg/L ♂ ♀ 0 0.00 0.00 0 0.00 0.00 125 12.9 (5.4) 14.5 (6.0) 83.8 8.7 (2.7) 10.2 (3.2) 250 24.1 (10.4) 25.3 (10.5) 167.5 17.4 (5.5) 19.5 (6.1) 500 39.1 (16.3) 42.5 (17.7) 335 30.9 (9.7) 34.6 (10.9) 1000 43.4 (18.1) 59.6 (24.8) 670 51.1 (16.0) 50.7 (15.9) 2000 48.2 (20.1) 43.8 (18.3) 1340 51.9 (16.3) 53.7 (16.8) Values represent group averages for chemical consumption and (total vanadium). Vanadium consumption was calculated from water consumption/chemical consumption; vanadium comprises 31% of vanadyl sulfate and 41.7% of sodium metavanadate. Averages for vanadyl sulfate and 0–500 mg/L sodium metavanadate are for study days 1–15. The averages presented for 1000 and 2000 mg/L sodium metavanadate are for study days 1–8, due to early removal. Table 3B Chemical Consumption (mg/kg/day): Mice. Based on the incidences of clinical observations and overt toxicity, our conclusions, with regard to sodium metavanadate being more toxic than vanadyl sulfate, have not changed. As communicated in the manuscript, the differences observed may be due to differences in total vanadium intake or differences in disposition or mechanism of toxicity. Additional studies are underway to help address these questions. The issue that we faced in this study is not unique to vanadyl sulfate and may be encountered with any hydrated or hygroscopic test material. Water may be present in the crystalline structure of a compound or associated via absorption. Water that is incorporated into the crystalline structure is likely to be consistent, while the amount of absorbed water can vary over time and with differing environmental conditions. It should be noted however that the presence of water in the test material does not change the chemical composition of the drinking water formulations i.e. animals in our study consumed a solution of vanadyl sulfate in water. While it is necessary to account for the water content when preparing solutions with these materials, it is not a trivial undertaking to determine the exact hydrated form or structure. Thus, assigning a suitable chemical name and molecular formula can be problematic. Even if the molecular formula is known, there may not be an existing CAS Registry Number that matches, and in this situation it may be desirable to create an alternative identifier. Given these considerations, in reporting our studies, we will continue to use the name vanadyl sulfate and the CASRN 27774-13-6, while also including a detailed description of the test material and its chemical characterization. This is a practical choice based on simplicity and has the added advantage that the studies will remain visible and properly tracked in electronic resources such as chemical toxicity databases. We hope that the lessons briefly communicated here will prove useful for other researchers who conduct studies with chemicals that exist as hydrates or are hygroscopic. We encourage others to pay close attention to chemical identity, purity and properties provided by the vendor, conduct their own chemical analyses when necessary, and ensure preparation and description of dose solutions properly account for water content. There is a wealth of information available to consult on these issues and researchers can take advantage of excellent public resources such as the National Library of Medicine’s PubChem and the USEPA’s Chemistry Dashboard databases to locate and reference chemical identity information. Table 3B Sodium Metavanadate Vanadyl Sulfate mg/L ♂ ♀ mg/L ♂ ♀ 0 0.00 0.00 0 0.00 0.00 125 18.4 (7.7) 15.3 (6.4) 83.8 11.7 (3.7) 10.0 (3.1) 250 41.1 (17.1) 26.5 (11.0) 167.5 23.2 (7.0) 16.9 (5.3) 500 58.2 (24.3) 38.5 (16.1) 335 35.6 (11.2) 28.4 (8.9) 1000 60.3 (25.1) 48.8 (20.3) 670 54.5 (15.8) 40.7 (12.7) 2000 112.5 (46.9) 46.7 (19.5) 1340 77.2 (24.2) 56.0 (17.6) Values represent group averages for chemical consumption and (total vanadium). Vanadium consumption was calculated from water consumption/chemical consumption; vanadium comprises 31% of vanadyl sulfate and 41.7% of sodium metavanadate. Averages for vanadyl sulfate and 0–500 mg/L sodium metavanadate are for study days 1–15. The averages presented for 1000 and 2000 mg/L sodium metavanadate are for study days 1–8, due to early removal. Based on the incidences of clinical observations and overt toxicity, our conclusions, with regard to sodium metavanadate being more toxic than vanadyl sulfate, have not changed. As communicated in the manuscript, the differences observed may be due to differences in total vanadium intake or differences in disposition or mechanism of toxicity. Additional studies are underway to help address these questions. The issue that we faced in this study is not unique to vanadyl sulfate and may be encountered with any hydrated or hygroscopic test material. Water may be present in the crystalline structure of a compound or associated via absorption. Water that is incorporated into the crystalline structure is likely to be consistent, while the amount of absorbed water can vary over time and with differing environmental conditions. It should be noted however that the presence of water in the test material does not change the chemical composition of the drinking water formulations i.e. animals in our study consumed a solution of vanadyl sulfate in water. While it is necessary to account for the water content when preparing solutions with these materials, it is not a trivial undertaking to determine the exact hydrated form or structure. Thus, assigning a suitable chemical name and molecular formula can be problematic. Even if the molecular formula is known, there may not be an existing CAS Registry Number that matches, and in this situation it may be desirable to create an alternative identifier. Given these considerations, in reporting our studies, we will continue to use the name vanadyl sulfate and the CASRN 27774-13-6, while also including a detailed description of the test material and its chemical characterization. This is a practical choice based on simplicity and has the added advantage that the studies will remain visible and properly tracked in electronic resources such as chemical toxicity databases. We hope that the lessons briefly communicated here will prove useful for other researchers who conduct studies with chemicals that exist as hydrates or are hygroscopic. We encourage others to pay close attention to chemical identity, purity and properties provided by the vendor, conduct their own chemical analyses when necessary, and ensure preparation and description of dose solutions properly account for water content. There is a wealth of information available to consult on these issues and researchers can take advantage of excellent public resources such as the National Library of Medicine’s PubChem and the USEPA’s Chemistry Dashboard databases to locate and reference chemical identity information.