A molecular dynamics run can be perfectly configured and still be frustrating to interpret: the temperature moves around, and it is not always clear whether the system is ready for the next step. NVT equilibration is where you bring the system toward its target temperature while keeping the number of particles and volume constant. The practical question is not whether the temperature becomes perfectly flat—it will fluctuate—but whether its behavior is stable enough to proceed.
This guide focuses on that decision: preparing an NVT run, reading its temperature plot, and knowing when to continue with NPT equilibration.
Start from a minimized structure
NVT comes after energy minimization. Before starting, make sure the minimization completed and the resulting structure looks reasonable. The NVT input can be the GRO file from minimization, or a batch project from a completed step. If you are continuing directly from the previous run, the auto-fill button can populate the input path for you.
In SAMSON, open the Equilibrate (NVT) tab and confirm that the chosen input corresponds to the system you intend to equilibrate. Checking the input before changing parameters helps avoid spending time running a job from the wrong structure or project.
Use the temperature plot as a readiness check
For a typical NVT run, the running average of the temperature should approach a plateau around the desired value. The instantaneous temperature does not need to sit exactly on the target at every point: small fluctuations are normal. Look at the overall trend and whether it appears stable enough for your workflow, rather than judging readiness from a single moment or endpoint.
The example plot shows temperature stabilizing around 300 K, the default reference temperature in the illustrated advanced parameters. It is an example, not a universal target. Your target should match your project’s requirements.
How long should NVT take?
The required time depends on the size and contents of the system. The tutorial gives 50–100 ps as a typical starting range for many NVT equilibration procedures, but the plot—not the number alone—should guide the decision. If the running average has not reached the desired value or still appears to be changing, continue equilibration. You can launch another NVT step using the results from the previous NVT run as input.
This makes the workflow iterative: run, inspect the temperature behavior, and extend equilibration if needed. A preset duration is useful for planning, but it is not proof that the system has stabilized.
Check the settings that control temperature
The NVT tab provides commonly adjusted parameters, including the integration time step and number of steps. More advanced options are available through All…. For many projects, the documentation suggests velocity rescaling with a stochastic term (v-rescale) and a time constant of about 1 ps. These are general starting points, not a substitute for choosing parameters appropriate to your system.
One detail is easy to overlook: the temperature used for velocity generation should correspond to the temperature coupling reference. If you couple separate groups—for example, Protein and non-Protein—the time constants and reference temperatures should be specified for the groups shown. The position-restraint options are filled automatically based on the input system. If you are following this tutorial, reset the advanced parameters to their defaults before running.
Run, then inspect the result
GROMACS Wizard can generate input files, run locally, or launch a cloud computation. For a local run, choose Equilibrate locally; progress is available in the Output window and the Local jobs list. Once the job finishes, import the trajectory or choose a subset of frames, then inspect the temperature plot in the Plots section. The plot is generated and saved when the job is finished and the results are loaded.
When the temperature has stabilized around the target and the behavior looks suitable to continue, proceed to NPT equilibration to stabilize density. If it has not, use the current NVT results as the input for additional equilibration. For the full walkthrough, see the GROMACS Wizard NVT Equilibration tutorial.
Note: SAMSON and all SAMSON Extensions are free for non-commercial use. Get SAMSON.