Assigning Animals Randomly to different treatments

set.seed(3349)

conditions <- c("Control R1", 
                "Control R2", 
                "0.1 mg/L R1", 
                "0.1 mg/L R2", 
                "1 mg/L R1", 
                "1 mg/L R2", 
                "10 mg/L R1", 
                "10 mg/L R2", 
                "100 mg/L R1", 
                "100 mg/L R2", 
                "1000 mg/L R1", 
                "1000 mg/L R2")
animal_id <- c("DM022024_C01", 
               "DM022024_C02", 
               "DM022024_C03", 
               "DM022024_C04*", 
               "DM022024_C05", 
               "DM022024_C06", 
               "DM022024_C07", 
               "DM022024_C08",
               "DM022024_C09",
               "DM022024_C10", 
               "DM022024_C11", 
               "DM022024_C12")

treatment_assignments <- data.frame(
  Conditions = sample(conditions),
  animal_ID = sample(animal_id)
)
print(treatment_assignments)

     Conditions     animal_ID
1     1 mg/L R1  DM022024_C02
2   100 mg/L R1  DM022024_C01
3   0.1 mg/L R1  DM022024_C03
4    Control R1  DM022024_C05
5  1000 mg/L R2  DM022024_C11
6  1000 mg/L R1  DM022024_C12
7     1 mg/L R2  DM022024_C09
8    Control R2  DM022024_C06
9   100 mg/L R2 DM022024_C04*
10   10 mg/L R2  DM022024_C10
11  0.1 mg/L R2  DM022024_C07
12   10 mg/L R1  DM022024_C08

#save assignments in csv for downstream use as metadata

setwd('..')
write.csv(treatment_assignments, file = "treatment_assignments_LC50_E2.csv")

Nickel Spiking Calculations

Nickel Chloride Dilutions

Review resources prior to handling nickel chloride:

Nickel chloride SDS.
Chemical resistance of gloves.

All handling of stock solution and treatments to be done in a chemical fume hood. Once spiked water has been distributed, add animals, parafilm jars, place in secondary container, and move to environmental chamber.

Required Volume of Artificial Seawater

Calculations for spiking artificial seawater. 225 mL of seawater required to mostly submerge glass slide in wide mouth 16 oz Kerr mason jar.

Considerations:

2 replicates
225 mL per jar
6 conditions
Account for 10% loss of water during transferring
Additional 15 mL required for analytical chemistry

\[(2 * 225 * 6) + (15*6) + (0.1*(2*225*6)) + (0.1*(15*6)) = 3,069\]

Need 3,069 mL of artificial seawater total for exposure, particularly, 511.5 mL of artificial seawater per condition.

Calculations

Stock solution of NiCl₂ (10 g/L) made 04/08/24.

NiCl₂ (5g)
Suspended in MilliQ water (500 mL)

0.1 mg/L

\[ C_1*V_1 = C_2*V_2\] \[ 10 g/L * V_1 = 0.0001g/L*0.5115L\] \[V_1 = 0.000005115 L = 5.115 uL\] Spike 511.5 mL of aerated, artificial seawater (18 C) with 5.115 uL of stock NiCl₂ solution. Change in salinity negligible (+/- 1 PPT).

1 mg/L

\[ C_1*V_1 = C_2*V_2\] \[ 10 g/L * V_1 = 0.001g/L*0.5115L\] \[V_1 = 0.00005115 L = 51.15 uL\]

Spike 511.5 mL of aerated, artificial seawater (18 C) with 51.15 uL of stock NiCl₂ solution. Change in salinity negligible (+/- 1 PPT).

10 mg/L

\[ C_1*V_1 = C_2*V_2\] \[ 10 g/L * V_1 = 0.01g/L*0.5115L\] \[V_1 = 0.0005115 L = 511.5 uL\]

Spike 511.5 mL of aerated, artificial seawater (18 C) with 511.5 uL of stock NiCl₂ solution. Change in salinity negligible (+/- 1 PPT).

100 mg/L

\[ C_1*V_1 = C_2*V_2\] \[ 10 g/L * V_1 = 0.1g/L*0.5115L\] \[V_1 = 0.005115 L = 5.115 mL\]

Spike 511.5 mL of aerated, artificial seawater (18 C) with 5.115 mL of stock NiCl₂ solution. Change in salinity negligible (+/- 1 PPT).

1000 mg/L

\[ C_1*V_1 = C_2*V_2\] \[ 10 g/L * V_1 = 1g/L*0.5115L\] \[V_1 = 0.0759 L = 51.15 mL\]

Spike 511.5 mL of aerated, artificial seawater (18 C) with 51.15 mL of stock NiCl₂ solution. Change in salinity negligible (+/- 1 PPT).