Increasing substrate concentration raises the reaction rate as more enzyme-substrate complexes form. 

However, if enzyme concentration is constant, a saturation point is reached where all active sites are occupied, therefore no more enzyme substrate complexes can be formed, and adding more substrate will no longer increase the reaction rate. Beyond this point, excess substrate molecules remain unbound.

Saturation point - when all available active sites are occupied.

As substrate concentration increases the increase in initial rate of reaction is linear, until the saturation point is reached. When the saturation point is reached the enzyme is operating at the maximum rate of reaction, know as Vmax.

If no substrate is added to a reaction mixture the amount of substrate will decrease as the reaction progresses, as it gets converted to product. The rate of reaction will decrease over time as there is less substrate available to form enzyme substrate complexes..

Limiting Factor - Enzyme Concentration

Higher enzyme concentration provides more active sites, increasing the chances of enzyme-substrate complex formation.

When sufficient substrate is available, the initial reaction rate rises linearly with increasing enzyme concentration.

If substrate is limited, increasing enzyme concentration beyond a certain point will not increase the reaction rate, as substrate availability becomes the limiting factor.

As temperature increases the rate of reaction increases. The substrate particles and enzymes gain more kinetic energy, so move faster, and collide more frequently. 

More frequent successful collisions results in more enzymes substrate complexes forming and a faster reaction rate, up to the optimum temperature.

The optimum temperature is the temperature at which the enzyme rate of reaction is at its maximum. Different organisms will be adapted to survive in different temperatures so their enzymes will have different optimum temperatures, for example certain Archea live around hot hydrothermal vents so have enzymes that can operate at higher temperatures than mammalian enzymes.

Above the optimum temperature the rate of reaction decreases. The bonds that hold the tertiary structure of the enzyme (such as hydrogen and ionic bonds) start to vibrate more and break.

As a result the tertiary structure changes. The enzyme has denatured. The active site will have changed shape and is no longer complementary to the substrate.

When the enzyme has denatured an enzyme substrate complex cannot form. 

Limiting Factor - pH

Enzymes are denatured at extremes of pH, anything higher or lower than the optimum pH.

The excess of H+ and OH- in solution will disrupt and break the hydrogen and ionic bonds in the tertiary structure of the enzymes and will denature the enzyme.

Once the enzyme has denatured the active site is no longer complementary to the substrate and no enzyme substrate complexes will form.

Different enzymes will have a different optimum pH. The optimum pH can indicate where in the body the enzyme may act. For example, a protease, pepsin, has an optimum pH of 2 because it is secreted in the stomach.

Some enzymes may be adapted to function over a wider range of pH values. For example, in the case of bacteria this could mean that they are more tolerant to changes in the environment.

When investigating the effect of pH on the action of enzymes a buffer solution can be used. This is very important if the products of the enzyme controlled reaction would lead to a pH change.