Aluminium trilactate

Neurotoxicity

Description of key information

A study on neurodevelopmental toxicity of Aluminium citrate in rats and several publication studying neurotoxicity of Aluminium trilactate and other Aluminium salts are available. The most critical effect is described in a publication by Golub and Germann, 2001.
Based on the results, the NOAEL for neurotoxic effects of Aluminium is 10 mg/kg bw/d. Recalculation to the corresponding dose of Aluminium trilactate resulted in a NOAEL of 109.04 mg/kg bw/d.

Key value for chemical safety assessment

Effect on neurotoxicity: via oral route

Link to relevant study records

Reference

Endpoint:

neurotoxicity: oral

Remarks:

developmental

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Acceptable, well documented publication which meets basic scientific principles.

Qualifier:

no guideline followed

Principles of method if other than guideline:

The study investigated the long-term motor and cognitive effects of developmental exposure to aluminium (Al) in diet administered to mice at doses 100, 500 and 1000 μg Al/g diet or 10, 50, and 100 mg Al/kg bw/day through development (from conception to 35 days of age) and the influence of less than optimal diets on developmental toxicity of Al.

GLP compliance:

not specified

Limit test:

no

Species:

mouse

Strain:

Swiss Webster

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Harlan Sprague Dawley
- Age at study initiation: ca. 8 weeks
- Weight at study initiation: no data
- Fasting period before study: no data
- Housing: animals were kept in "a temperature- and photoperiod-controlled room"
no data for parental generation;
offspring: males were housed one per cage, females were housed two per cage in plastic tub-type cages with stainless steel lids and plastic water bottles
- Diet (e.g. ad libitum): basal purified diet ad libitum, composition is based on the recommendations of the National Research Council, but concentrations of some nutrients were reduced to correspond to the usual intake of these nutrients by young women
- Water (e.g. ad libitum): deionized water, ad libitum
- Acclimation period: 1 week

ENVIRONMENTAL CONDITIONS
housing in temperature- and photoperiod-controlled room

Route of administration:

oral: feed

Vehicle:

other: Al lactate mixed with purified diet

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
No data

DIET PREPARATION
diet was commercially prepared and fed ad libitum as preweighed portions in pelleted form to facilitate food intake recording

VEHICLE
No data

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

no data

Duration of treatment / exposure:

from conception to 35 days of age:

Dams were exposed to Al lactate during GD 0 - 21 and PND 1 - 21.
Offspring was exposed in utero on GD 0 - 21 and postnatally through maternal milk at PND 0 - 21 and Al contained diet at PND 21 - 35 (including peripubertal period of development).

Frequency of treatment:

daily

Remarks:

Doses / Concentrations:
7, 100, 500, or 1000 mg Al/g diet
Basis:
nominal in diet
Al content of the basal diet averaged 7 mg Al/g.

Remarks:

Doses / Concentrations:
< 1, 10, 50, or 100 mg Al/kg body weight/day
Basis:
actual ingested

No. of animals per sex per dose:

maternal: no data
offspring: 20 male, 16 female

Control animals:

yes, plain diet

Details on study design:

Only parental females F0 were exposed

Observations and clinical examinations performed and frequency:

Offspring: Body weight and food intake were measured at 2-week intervals from weaning to time of testing.

Neurobehavioural examinations performed and frequency:

Morris maze testing
The Morris maze was a 90-cm diameter pool filled with 15 cm of water (20–22°C) made opaque with powdered nontoxic paint (Tempura White 427217, Palmer Paint, Troy, MI). The maze was monitored from a video camera positioned above the pool. The video monitor was divided into four 90° quadrants. A 6x6-cm clear plastic platform was placed 1 cm below the surface 13 cm from the rim. The procedure was adapted from studies in aged mice so that the task would also be appropriate in older Al-exposed cohorts. Well defined cues were provided to compensate for potential loss of vision in older mice. The salient cue was a 15-cm 110° black plastic arc placed along the side of the tank, with the lower edge below the water surface, in the quadrant opposite the platform and extending into an adjacent quadrant. This cue could be accessed both visually and by touch. Potential visual cues in the room were occluded by a 90-cm-high gray plastic cylinder that surrounded the pool. By providing well defined cues, and by limiting room cues, the paradigm allowed cue manipulation sessions. It would be anticipated that place learning, rather than spatial navigation learning, would be involved in improved performance of the task. The cylinder had a door located in each quadrant to allow placement of the mouse at the beginning of each trial. An additional distal nonsalient visual cue was provided by leaving open the door (7.5 cm diameter semicircle) in the plastic cylinder in the quadrant adjacent to the salient cue.
Each session consisted of four trials, with the point of entry being randomly varied by quadrant. The maximum trial length was 120 s. For the intertrial interval the mouse was placed on the platform for 30 s. Escape latency, quadrant duration, and contacts with the platform were recorded by observers from the monitor either during the session or from videotape. Testing was conducted in the afternoon, during the light phase of the light/dark cycle, and the order of the mice was randomized each day.
For the learning phase, four daily sessions of four trials were conducted. A fifth session conducted 72 h after the fourth session measured longer-term retention. Both the fourth and fifth session were followed immediately by a probe trial, for which the platform was removed. Sessions 6– 8 were cue rotation sessions with the arc cue, door cue, and platform rotated 180° but maintained in the same relative positions. These sessions were also followed by a probe trial.
For the Session 6 probe trial, both salient and nonsalient cues were present; for the Session 7 probe trial, only the nonsalient cue was present; and for Session 8 probe trial, only the salient cue was present.

Motor testing
A test battery developed for rats was adapted for use. A single session consisted of five tests administered with 2 min between tests in the following order: rotarod, grip strength, wire suspension, mesh pole descent, beam traversal.
Two 120-s training trials using an automated rotarod apparatus were conducted at 20 rpm, 2 h apart on the day before the test session. For rotarod testing, one 120-s trial was conducted at 32 rpm. Grip strength was measured with an online strain gauge. Three trials each were conducted for fore- and hindlimbs.
Wire suspension was tested by allowing the mouse to grasp a horizontal wire (2 mm diameter) with its forepaws. During three 120-s trials at 30-s intervals, the mouse could either maintain suspension, fall to the surface (50 cm below), or move horizontally to a support pole and climb down.
For mesh pole descent, the mouse was placed at one end of a wooden pole (3 cm diameter, 82 cm long) covered with wire mesh, which was then moved to a vertical position. Time required to descend the pole (max 120 s) was recorded.
For beam traversal, round and square 10-mm diameter beams were elevated 80 cm above the surface. The mouse was placed in the middle of the beam and the time required to reach a platform at either end of the beam, or to fall from the beam, was recorded (maximum 120 s). Two testers participated in these assessments.
Reliabilities were established at the time testing began. Testing was conducted during the light phase of the cycle. The animal cages were blindcoded for group. For most tests, average (across trials), individual trials, as well as best performance were analyzed.

Sacrifice and (histo)pathology:

MATERNAL ANIMALS
No postmortem examinations reported

OFFSPRING
-Animals were sacrificed at PND 90.
Organ weights were obtained from a subsample of mice at the end of the study: brain, liver, heart, kidney, spleen, tibia

Statistics:

Dependent variables were analyzed via ANOVA with treatment groups compared to controls using Fisher LSD post hoc tests. Because only one offspring per litter was involved in any one assessment, statistical control for litter effects was not needed. If a significant treatment effect was identified, a linear regression analysis using administered dose as the independent variable was also conducted. Because growth retardation was found in this experiment, dependent variables showing treatment effects were entered into regression to determine if there was an association between body weight and performance variables in the
population as a whole. If this was the case, an ANCOVA was conducted with body weight as the covariate. If no group by covariate interaction was detected, the interaction term was removed for the final evaluation of group effects. Statview and SAS software (SAS Institute, Cary, NC) were used.

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

not specified

Ophthalmological findings:

not examined

Clinical biochemistry findings:

not examined

Behaviour (functional findings):

effects observed, treatment-related

Gross pathological findings:

effects observed, treatment-related

Neuropathological findings:

not specified

Other effects:

no effects observed

Details on results:

There were no group differences in the number of dams completing pregnancy, gestation length, pregnancy weight gain (gestation day: GD 0–15), litter size at birth, or birth weight (data not shown). By weaning, both males and females in the Al500 and Al1000 groups weighed significantly less than controls, but by 35 days of age only the Al1000 group was smaller.

Food intake (grams per day) also differed by group in the same pattern as weights; however food intake per unit body weight was higher, rather than lower, in the Al500 and Al1000 groups than in controls.

Brain, liver, heart, kidney, spleen, and tibia weights were lower in the Al1000 group than in controls and the difference was significant for heart
and kidney. The absolute brain weight of the Al100 group was greater than controls. Al treatment did not affect relative organ weights, with the exception of brain. The relative brain weights of both the Al100 and Al1000 groups were greater than controls.

Dose descriptor:

NOAEL

Effect level:

10 mg/kg bw/day (nominal)

Based on:

element

Remarks:

Al3+

Sex:

male/female

Basis for effect level:

other: latency to fall off wire in wire suspension test, latency to locate the platform following cue relocation in the water maze test

Remarks on result:

other: Generation: offspring (migrated information)

No alterations in pregnancy weight gain or pup birth weights were observed. At PND 21, significant decreases in pup body weights were seen at 50 and 100 mg Al/kg bw/day. No information on maternal weight gain during lactation was given; however, the authors stated that the decrease in pup weight was not associated with reduced maternal food intake. At PND 35, a statistically significant decrease in body weight was observed at 100 mg Al/kg bw/day. On PND 90, female mice in the 100 mg Al/kg bw/day group weighed 15% less than controls. Decreases in heart and kidney weights were observed at 100 mg Al/kg bw/day in the females. Increases in absolute brain weight were seen in females at 10 mg Al/kg bw/day and relative brain weights were observed at 10 or 100 mg Al/kg/day, but not at 50 mg Al/kg bw/day. In the males, significant decreases in body weight were observed at 50 (10%) and 100 (18%) mg Al/kg bw/day at 5 months; an increase in food intake was also observed at these doses.

In the Morris maze (tested at 3 months in females), fewer animals in the 100 mg Al/kg/day group had escape latencies of <60 seconds during sessions 1-3 (learning phase) and a relocation of the visible cues resulted in increased latencies at 50 and 100 mg Al/kg bw/day. Body weight did not correlate with latency to find the platform or with the distribution of quadrant times. The authors concluded that controls used salient and/or nonsalient cues, 10 and 50 mg Al/kg bw/day animals used both cues, but had difficulty using only one cue, and 100 mg Al/kg bw/day animals only used the salient cues.

In the males tested at 5 months, a significant decrease in hindlimb grip strength was observed at 100 mg Al/kg bw/day, an increase in the number of rotations on the rotorod as observed at 100 mg Al/kg bw/day, and a shorter latency to fall in the wire suspension test was observed at 50 and 100 mg Al/kg bw/day. The authors stated that there were significant correlations between body weight and grip strength and number of rotations. When hindlimb grip strength was corrected for body weight, the aluminium-exposed mice were no longer significantly different from controls; the number of rotations was still significantly different from control after adjustment for body weight.

Conclusions:

Groups of 20 Swiss Webster mice were exposed to <1, 10, 50, or 100 mg Al/kg bw/d (as Aluminium lactate) via diet from conception to 35 days of age and tested behaviourally as adults.
Subtle deficits in several neuroparameters were observed, including impaired learning for the females in a maze in the 100 mg Al/kg bw/d group and poorer cue utilisation in the maze in both the 50 and 100 mg Al/kg bw/d groups. Performance of the males on the rotarod test was impaired in the 100 mg Al/kg bw/d group. A reduction in hind limb grip strength was reported in approximately 15% of animals in the 100 mg Al/kg bw/d group; this was no longer significant after adjustment for body weight. A dose-related and statistically significant difference between controls and rats given the 50 or 100 mg Al/kg bw/d were observed on wire suspension fall latency.
The data suggest that developmental Al exposure under normal, but less than optimal, dietary conditions can lead to subtle but long-term effects on growth and brain function in adulthood. The NOAEL identified in this study was 10 mg Al/kg bw/d.

Executive summary:

Groups of 20 Swiss Webster mice were exposed to <1, 10, 50, or 100 mg Al/kg bw/d (as Aluminium lactate) via diet from conception to 35 days of age and tested behaviourally as adults.

Subtle deficits in several neuroparameters were observed, including impaired learning for the females in a maze in the 100 mg Al/kg bw/d group and poorer cue utilisation in the maze in both the 50 and 100 mg Al/kg bw/d groups. Performance of the males on the rotarod test was impaired in the 100 mg Al/kg bw/d group. A reduction in hind limb grip strength was reported in approximately 15% of animals in the 100 mg Al/kg bw/d group; this was no longer significant after adjustment for body weight. A dose-related and statistically significant difference between controls and rats given the 50 or 100 mg Al/kg bw/d were observed on wire suspension fall latency.

The data suggest that developmental Al exposure under normal, but less than optimal, dietary conditions can lead to subtle but long-term effects on growth and brain function in adulthood. The NOAEL identified in this study was 10 mg Al/kg bw/d.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

109.04 mg/kg bw/day

Study duration:

subacute

Species:

mouse

Quality of whole database:

All reported studies are of high quality (Klimisch score=2).

Effect on neurotoxicity: via inhalation route

Endpoint conclusion

Endpoint conclusion:

no study available

Effect on neurotoxicity: via dermal route

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

The nervous system is reported to be the most sensitive target of Aluminium toxicity. It has been suggested that Aluminium has the potential to produce neurotoxic effects and to be involved in the aetiology of Alzheimer’s disease and other neurodegenerative diseases in humans (EFSA (European Food Safety Authority), 2008;WHO IPCS EHC (World Health Organistion International Programme on Chemical Safety Environmental Health Criteria), 1997).

Human studies reviewed by EFSA (2008) were concluded to be not informative for a safety assessment of Aluminium from dietary intake. These epidemiological studies are not conclusive according to EFSA (2008) as some suggest an association with neurotoxicity and other studies do not.

Several publications studying neurotoxicity of Aluminium trilactate and other Aluminium salts in laboratory animals have been reviewed by DECOS(Dutch Expert Committee on Occupational Safety) (2010), EFSA (2008),US ATSDR (United States Agency for Toxic Substances and Disease Registry)(2008),WHO IPCS EHC(1997), Krewski et al. (2007) andFAO/WHO JECFA (Joint Food and Agriculture Organisation of the United Nations/World Health Organisation Expert Committee on Food Additives)(2007). NOAELs ranging from 10 to 62 mg Al/kg bw/d are reported for the developing nervous system. These reviews have been taken into account for the selection of key and supporting studies.

According to DECOS (2010), the oral administration of water-soluble aluminium compounds to dams during gestation had only effects on fetuses (such as decreased weights and retarded ossification) at dose levels inducing general toxicity effects. In post-natal studies pup mortality as well as neurodevelopmental and behavioural effects were also seen at doses not inducing general toxicity.

The most critical NOAEL is published by Golub and Germann, 2001:

Groups of 20 Swiss Webster mice were exposed to <1, 10, 50, or 100 mg Al/kg bw/d (as Aluminium lactate) via diet from conception to 35 days of age and tested behaviourally as adults. Subtle deficits in several neuroparameters were observed, including impaired learning for the females in a maze in the 100 mg Al/kg bw/d group and poorer cue utilisation in the maze in both the 50 and 100 mg Al/kg bw/d groups. Performance of the males on the rotarod test was impaired in the 100 mg Al/kg bw/d group. A reduction in hind limb grip strength was reported in approximately 15% of animals in the 100 mg Al/kg bw/d group; this was no longer significant after adjustment for body weight. A dose-related and statistically significant difference between controls and rats given the 50 or 100 mg Al/kg bw/d were observed on wire suspension fall latency. The data suggest that developmental Al exposure under normal, but less than optimal, dietary conditions can lead to subtle but long-term effects on growth and brain function in adulthood. The NOAEL identified in this study was 10 mg Al/kg bw/d.

A study on neurodevelopmental toxicity of Aluminium citrate is available:

Pregnant Sprague-Dawley dams (n=20 per group) were administered aqueous solutions of aluminium citrate at 3 dosage levels (nominal 30, 100 and 300 mg Al/kg bw/d. Two control groups received either a sodium citrate solution (citrate control with 27.2 g/L) or plain water (control group). The Al citrate and Na citrate were administered to damsad libitumvia drinking water from gestation day 6 until weaning of offspring. Weaned offspring were dosed at the same levels as their dams. Pups were assigned to one of four cohorts (80 males, 80 females): a pre-weaning cohort that was sacrificed at PND 23, and cohorts that were sacrificed at PND 64, PND120 and PND 364. Endpoints were assessed in both female and male pups that targeted behavioral ontogeny (motor activity, T-maze, auditory startle, the Functional Observational Battery (FOB) with domains targeting autonomic function, activity, neuromuscular function, sensimotor function, and physiological function), cognitive function (Morris swim maze), brain weight, clinical chemistry, haematology, tissue/blood levels of aluminium and neuropathology at the different dose levels and time points PND 23, 64, 120 and 364. FOB observations showed no clear treatment-related effect among the neonatal Day 364 cohort pups that were assessed at PND 5 and 11 or in the juvenile pups assessed ca.PND 22. There was some evidence for dose-response relationships between neuromuscular measurements – hind-limb and fore-limb grip strength and Al-treatment in both males and females, although some of the effects may be secondary to body weight changes. Although the FOB endpoint most consistently associated with Al-citrate treatment, grip strength, measurements showed considerably variability and a consistent ordering of the Al-treatment group responses (dose-response) was not observed at all time points. No consistent treatment-related effects were observed in ambulatory counts (motor activity) in the different cohorts. No significant effects were observed for the auditory startle response, T-maze tests (pre-weaning Day 23 cohort) or the Morris Water Maze test (Day 120 cohort).

Whole body Al levels in neonatal pups from high dose females and males were greater than those in the control groups. There were no significant sex differences. These results suggest transfer of Al from dams to pupsin utero, although a contribution from breast milk PND 0 to 4 is also possible. Aluminium levels were assayed in several tissues in the pup cohorts. Levels of Al in whole blood were highest in the Day 23 cohort animals and declined with time, possibly due to the lower amounts of water (test solution) consumed once the pups matured. Although during the lactation period pups may have consumed some water/test solution, the results suggest that transfer of Al from dams to pups can occur through breast milk. Of the central nervous system tissues, Al levels were highest in the brainstem. Although levels of Al were relatively low in the cortex (< 1 µg/g), they were positively associated with Al levels in the liver and femur. In females, Al levels in the high dose group remained elevated relative to the other groups at all time points suggesting that accumulation might have occurred. Although representative of actual human exposures, extending the period of exposure beyond weaning until day 364 leads to ambiguity in interpretation of the results as effects observed later in the study may have resulted from either later exposures or exposures during periods critical for development.

The results from this study are informative for developmental and neurotoxic effects due to combined prenatal and chronic postnatal exposure of rats to high doses of aluminium (30, 100 and 300 mg Al/kg bw/d). As the offspring were dosed during the whole post-weaning period, it is difficult to differentiate between developmental or direct toxicity after weaning, however. The study showed no evidence of an effect of Al-citrate on memory or learning but a more consistent effect was observed in endpoints in the neuromuscular domain.

The critical effect was a deficit in fore- and hind-limb grip strength in the mid-dose group, supported by evidence of dose response and less consistently observed effects in the mid-dose animals: urinary tract lesions at necropsy; body weight; tail pinch; and foot-splay. Thus, a LOAEL of 100 mg Al/kg bw/day for aluminium toxicity is assigned. The NOAEL in this study was 30 mg Al/kg bw/d.

 

The latter study has not yet been included into the existing reports by EFSA and other authorities, but it does not rise more concern than those data taken into account.

No evidence for neurotoxicity of Lactic acid exists. Lactic acid is a common biological molecule, to which humans are continuously exposed, from diet, from bacterial generation in the gut, and from intramitochondrial processes. It is a metabolic intermediate derived e.g. from glycogen and amino acids metabolism that is produced by most mammalian cells and other organisms (Andersen, 1998).

Based on these results, the NOAEL for neurotoxic effects of Aluminium is 10 mg/kg bw/d. This is consistent with the NOAEL used by EFSA (2008) for the derivation of the Provisional Tolerable Weekly Intake of Aluminium from all dietary sources.

Recalculation to thecorresponding dose of Aluminium trilactate resulted in a NOAEL of109.04mg/kg bw/d.

  

References:

Andersen (1998) FINAL REPORT ON THE SAFETY ASSESSMENT OF GLYCOLIC ACID, [...], AND SODIUM GLYCOLATES, [...] , AND LACTIC ACID, AMMONIUM, CALCIUM, POTASSIUM, SODIUM, AND TEA-LACTATES, METHYL, ETHYL, ISOPROPYL, [...],International Journal of Toxicology, Vol.17, Suppl.1.

DECOS (Dutch Expert Committee on Occupational Safety) (2010): Aluminium and aluminium Compounds, Health-based recommended occupational exposure limit. (No. 2010/05OSH, The Hague, July 15, 2010); available via internet: http://www.gezondheidsraad.nl/sites/default/files/201005OSH.pdf

EFSA (European Food Safety Authority) (2008) Safety of aluminium from dietary intake, The EFSA Journal 754, 1-34, available via internet: http://www.efsa.europa.eu/de/efsajournal/pub/754.htm

FAO/WHO JECFA (Joint Food and Agriculture Organisation of the United Nations/World Health Organisation Expert Committee on Food Additives)(2007) Safety evaluation of certain food additives and contaminants. WHO FOOD ADDITIVES SERIES: 58, World Health Organization, Geneva, 2007, available via internet: http://whqlibdoc.who.int/publications/2007/9789241660587_eng.pdf

Krewski, et al. (2007). Human Health Risk Assessment for Aluminium, Aluminium Oxide, and Aluminium Hydroxide, A Report Submitted to the US Environmental Protection Agency. J Toxicol Environ Health B Crit Rev. 10 Suppl 1:1-269. Available via internet: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2782734/

US ATSDR (United States Agency for Toxic Substances and Disease Registry)(2008) Toxicological profile for Aluminium, U.S. DEPARTMENT OF HEALTH AND HUMAN SERVICES, Public Health Service, Agency for Toxic Substances and Disease Registry, available via internet: http://www.atsdr.cdc.gov/toxprofiles/tp.asp?id=191&tid=34

WHO IPCS EHC (World Health Organisation International Programme on Chemical Safety Environmental Health Criteria)(1997) Aluminium (Environmental health criteria; 194), IPCS, World Health Organization, Geneva, available via internet: http://www.inchem.org/documents/ehc/ehc/ehc194.htm  

Justification for selection of effect on neurotoxicity via oral route endpoint:
The most critical effects (subtle deficits in several neuroparameters including impaired learning and poorer cue utilisation in the maze, reduction in hind limb grip strength, wire suspension fall latency) were reported in this publication.

Justification for selection of effect on neurotoxicity via inhalation route endpoint:
no relevant route of exposure

Justification for selection of effect on neurotoxicity via dermal route endpoint:
very low dermal absorption

Justification for classification or non-classification

Comparison of the available data on neurotoxicity with the classification criteria and guidance values indicates that Aluminium trilactate, does not need to be classified for neurotoxicity according to Directive 67/548/EEC as well as CLP, EU GHS (Regulation 1272/2008/EC) and therefore labelling is not necessary.