The in vitro reduction of sodium [36Cl]chlorate in bovine ruminal fluid1,2

Abstract

Sodium chlorate effectively reduces or eliminates gram-negative pathogenic bacteria in the gastrointestinal tracts of live cattle. Limitations to the in vivo efficacy of chlorate are its rapid absorption from the gastrointestinal tract and its presumed reduction to chloride within the gastrointestinal tract. We hypothesized that chlorate would be reduced via ruminal bacteria in a ruminal in vitro system and that the reduction of chlorate would be influenced by the dietary for-age:concentrate ratio; thus, 4 ruminally cannulated steers were fed 20 or 80% concentrate diets in a crossover design. Ruminal fluid was collected in 2 periods and dispensed into in vitro tubes containing sodium [36Cl]chlorate, which was sufficient for 100 or 300 mg/L final chlorate concentrations. The tubes were incubated for 0, 1, 4, 8, 16, or 24 h; autoclaved, control ruminal fluid, fortified with sodium [36Cl]chlorate, was incubated for 24 h. Chlorate remaining in each sample was measured by liquid scintillation counting after [36Cl]chloride was precipitated with silver nitrate. A preliminary study indicated that chlorite, a possible intermediate in the reduction of chlorate, had a half-life of approximately 4.5 min in freshly collected (live) ruminal fluid; chlorite was, therefore, not specifically measured in ruminal incubations. The chlorate dose did not affect in vitro DM digestion (P > or = 0.11), whereas in vitro DM digestibility was decreased (P < or = 0.05) by 80% forage content. By 24 h, 57.5 +/- 2.6% of the chlorate remained in 100-mg/L incubations, whereas 78.2 +/- 2.6% of the chlorate remained in the 300-mg/L incubations. When the data were expressed on a concentration basis (mg/L), diet had no effect (P > or = 0.18) on chlorate reduction; however, when chlorate reduction was expressed on a percentage basis, chlorate reduction tended to be greater (P > or = 0.09) at 8 and 16 h in the incubations containing the low-concentrate diet. Chlorate remaining in autoclaved controls at 24 h was intermediate (P < 0.01) between chlorate remaining in live ruminal fluid samples incubated for 0 or 24 h. Attempts to isolate chlorate-respiring bacteria from 2 sources of ruminal fluid were not successful. These data indicate that microbial-dependent or chemical-dependent, or both, reduction of chlorate occurs in bovine ruminal fluid and that dietary concentrate had a negligible effect on chlorate reduction.

Chloroxyanion Residues in Cantaloupe and Tomatoes after Chlorine Dioxide Gas Sanitation

“…The methods 66 used by, and conclusions 59,60 of, Abdel Rahmen et al, who reported that chlorite is a stable residue in rats dosed with chlorine dioxide, chlorite, and chlorate have been refuted. 56 Instability of chlorite in biomatrices has been further demonstrated; for example 17.3 μg/mL of chlorite had a half-life of only 4.5 min in bovine ruminal fluid 67 and was a detectable, but transitory, metabolite of 36 Cl-chlorate in pure cultures of E. coli. 68 In the latter study, chlorite was only measurable by directly injecting culture fluid, without pretreatment, onto an ion chromatograph equipped with a radiochemical detector.…”

Chlorate Metabolism in Pure Cultures of Escherichia coli O157:H7 Pretreated with either Nitrate or Chlorate

“…Oliver et al (26) found that in vitro ruminal fluid cultures were capable of reducing moderate concentrations of chlorate (0.8 and 2.8 mM) during 24 h incubations. The degree of chlorate reduction (42% of total) for the 0.8 mM treatment of Oliver et al (26) was comparable to the degree of chlorate reduction (51%) for the 1 mM treatment of this study (Figure 2A,B).…”

“…In this study, where chlorite was directly detected, only low quantities (23 ( 8 μM) of chlorite were present in pure culture at 12 h (Figure 3B), the time when chlorate reduction rates were maximal. Previously, we determined that the half-life of modest quantities (28 μM) of [ 36 Cl]chlorite under anaerobic culture conditions (live ruminal fluid) was about 4.5 min (26). In preliminary work for this study, we measured the stability of low concentrations (0.24 μM) of chlorite in TSB to ensure that, if formed, chlorite could be detected in our matrix; after 1 h in TSB, 86% of the starting chlorite remained (data not shown), indicating that the TSB itself would not quench the presence of chlorite.…”

“…Parent [ 36 Cl]chlorate and [ 36 Cl]chloride were quantified after precipitation of [ 36 Cl]chloride with silver nitrate as described by Oliver et al ; reported chlorate concentrations are corrected for the starting radiochemical purity (i.e., T0 incubations contained 5.6% [ 36 Cl]chloride). Studies counting radioactivity in incubation supernatant before and after pelleting bacterial cells by centrifugation revealed no difference in supernatant activity.…”

“…For example, chlorite residues have not been measured in tissues or excreta of farm animals treated with [ 36 Cl]chlorate (19)(20)(21)(22) even though chlorite was reported by Abdel-Rahman et al (23,24) to be a metabolite excreted into the (25) were not able to replicate the findings of Abdel-Rhaman (23,24), but did demonstrate that chlorite is fairly unstable in rat urine. Further studies with [ 36 Cl]chlorite have demonstrated that its half-life was only 4.5 min in bovine ruminal fluid (26). In bacterial systems, studies supporting the bacterial reduction of chlorate to chlorite have relied on relatively insensitive (paper chromatography) and (or) nonspecific, indirect analyses (titration) for measurement of chlorite (2,4,27).…”