INTERVIEW QUESTIONS

1. HR (HUMAN RESOURCE) INTERVIEW QUESTIONS 

2. QUALITY CONTROL INTERVIEW QUESTIONS ON WET CHEMICAL ANALYSIS

3. QUALITY CONTROL INTERVIEW QUESTIONS ON WET INSTRUMENTAL ANALYSIS

4. QUALITY CONTROL INTERVIEW QUESTIONS ON CHROMATOGRAPHY

5. INTERVIEW QUESTIONS ON SYSTEM SUITABILITY PARAMETERS - HPLC & GC

6. INTERVIEW QUESTIONS ON TROUBLESHOOTING - HPLC & GC

INTERVIEW QUESTIONS ON TROUBLE SHOOTING - HPLC & GC

1) PRESSURE PROBLEMS IN HPLC?

Low or no pressure:
› The solvent inlet lines are not being immersed in the solvent. (Immerse the inlets into the solvents)
› No solvent in the reservoir. (Fill the reservoir with mobile phase)
› Leaks in the system. (Identify leaks and close the leaks)
› Air lock in the pump line. (Remove all the air and purge the system with mobile phase)

High pressure:
› Blocked tubing around injector or column inlet. (Wash the system with proper solvent)
› High viscous solvents. (Better to use less viscous solvents)

Fluctuating pressure:
› Poorly primed (purged) lines. (Purge sufficient time)
› Badly degassed solvents. (Degas the mobile phase properly)
› Column contamination. (Check the system with known good column)

2) RETENTION PROBLEMS IN HPLC?

Retention time changes from injection to injection:
› Insufficiently equilibrated system. (Equilibrate the system up to steady baseline)
› Ambient Temperature fluctuation. (Maintain the system environment properly)
› Pressure fluctuating. (See above)
› Column contamination. (Check the system with known good column)


Continuously increasing or decreasing Retention time:
› Solvent preparation. (Prepare M.P freshly by mixing correct proportions)
› Flow rate changes. (Check the flow rate by graduated volumetric flask)
› Solvent delivery system blockage. (Wash the system with proper solvent)
› Ambient Temperature variation. (Maintain the system environment properly)
› Insufficient equilibration. (Equilibrate the system up to steady baseline)
› System leak. (Identify leaks and close the leaks)
› Column contamination. (Check the system with known good column)


Increasing or decreasing to a new constant Retention time:
› Mobile phase inconsistencies. (Prepare M.P freshly by mixing correct proportions)
› Gradient delays. (Check the gradient and curves)
› Incorrect column. (Use proper column)
› Changes in the solvent flow rate. (Check the flow rate by graduated volumetric flask)
› Temperature change. (Maintain the system environment properly)
› Column contamination. (Check the system with known good column)
› Leak in the system. (Identify leaks and close the leaks)


3) WHAT ARE THE REASON FOR NEGATIVE PEAKS IN HPLC?


› Absorption of analyte is lower than the absorption of mobile phase. (Use less absorptive mobile phase- use all HPLC grade solvents)
           Note: Generally in QC, all validated methods are used so Mobile phase may not be a problem. But we look for any contamination to mobile phase which are highly absorbing than analyte and also quality of solvents used.
› Air bubble observed in system. (Give purge injector & Needle wash)
› Detector signal polarity setting reversed-all peaks negative. (Correct it)
› Cabels reversed-all peaks negative. (Plug the cables correctly)


4) WHAT ARE THE REASONS FOR FLUCTUATION IN PEAK AREA WHEN THE SAME SAMPLE IS INJECTED MULTIPLE TIMES IN HPLC?

› Air bubble in the injector. (Perform needle wash sufficient time)
› Carry over. (Needle wash with proper solvent)
› Temperature changes. (Inject the sample after attaining sample temperature or Injector temperature).
    Explanation: If you take the sample out of refrigerator and put it into auto injector it will slowly warm up to the injector temperature and the sample will expand. So that initially you will inject a larger mass of the sample than later. (when the sample has reached the injector temperature of the sample compartment). So peak areas decreases from run to run.
› Random variation of flow rate. (Monitor the back pressure & rectify if any problem).
› Integration problems. (Check and reprocess the data)
         Ghost peaks & negative peaks produced by the mobile phase contaminations affect the integration.
› Sample itself can be a source of problem. (Samples like proteins).
        a) Some proteins do not elute completely during the initial runs. An additional quantity is eluted in the subsequent runs. (If this occurs blank need to be run between the analysis)
       b) Some proteins irreversibly adsorbed (bind to active sites) on some columns. The peak area obtained with brand new columns may increase slowly with respective injections. (Once these active sites are saturated reproducible peak areas obtained. Saturate the active sites with your analyte or other proteins before the actual injection).
       c) Some proteins can also bind to the wall of the vial. So that peak area decreases from injection to injection. (Use specially made vials)

5) WHAT ARE THE REASONS FOR GHOST PEAKS?

› Column contamination (Flush the column)
› Carry over (Flush the column with strong solvent)
› Contaminated water (Use HPLC grade water)
› Unknown interferences in sample (Check for interferences)

6) WHAT ARE THE REASONS FOR PEAK DOUBLING?

› Blocked frit (Replace or clean the frit)
› Co-elution of interfering compound (Check for interferences)
› Column over loaded  (Flush the column)
› Sample volume too large (Decrease the sample amount)

7) HOW TO TROUBLE SHOOT NOISE,DRIFT & SPIKE?

Noise:
☻Detector lamp problem or Dirty flow cell (Replace UV lamp or clean dirty flow cell).
☻Gradient or Isocratic proportioning (Lack of proper solvent mixing) - (Check the mixing device)
☻Homogeneity of Mobile phase - (Degas and sonicate well the M.P before)

Drift:
☻Positive drift (Absorbance of M.P-B) - (Use non uv absorbing M.P)
☻Positive drift (Contamination) - (Flush the column with strong solvent)
☻Negative drift (Absorbance of M.P-A) - (Use non uv absorbing M.P)
☻Wavy or Undulating drift (Temperature changes) -(Control the environment)

Spike:
☻Column stored without caps (Store column always in solvent by tightly capping)
☻Bubble in the detector (Degas the M.P)
☻External electrical interferences (Use Voltage stabilizer or UPS power supply)
☻Column temperature higher than boiling point of solvent (Use low column temperature than solvent boiling point)  

HR INTERVIEW QUESTION

Even though you have an excellent knowledge on technical aspects and cleared the technical round, you may not selected until you pass HR interview. 

To clear HR interview you should have political approach while answering the question. Find below questions with possible best answers. 

1) TELL ME ABOUT YOUR SELF?
Follow the below format to describe yourself.
1. Name
2. Educational details (starting from higher).
3. Experience with work profile.
4. Professional strength.
5. Technical knowledge.
6. Positives.
7. Negatives.
8. Family details

(And conclusion is) If you consider my education & experience, I think i am suitable to your job.

2) WHY SHOULD I SELECT YOU? (or) WHY SHOULD I CHOOSE YOU?
First of all, I am sure i have the all eligibilities that match your job. Apart from the eligibilities, there are many reasons but to be very specific my USP is hard work, enthusiastic to learn, passionate to do work, which matches your job profile.

3) WHY ARE YOU LEAVING THE PRESENT JOB? (or) WHY DID YOU LEAVE THE LAST JOB?
That was a good job and had nice colleagues there. No doubt about it. But according to me we cannot grow in the field without taking more responsibilities & risks and also we can't enhance our skills (like team leading capabilities and Managerial skills) without expose to wide range of people. I think it’s time for me to grow and enhance my skills. And also I need some salary hike to make my life some more beautiful.

4) WHAT ARE YOUR WEAKNESSES?
Well, my presentation skills are not the best one. But I read a lot of literature and regularly practice to improve it. And it was much better as it was one year back.

5) WHAT IS YOUR SALARY EXPECTATIONS?
Salary is not the first priority for me I applied for this job because I really like the role your offer. I know if I work well and achieve good results salary will automatically meet my expectations.

6) WHERE DO YOU SEE YOUR SELF IN FIVE YEARS?
I hope to be working in your company as manager, achieving great results in my department in 5 years time.

INTERVIEW QUESTIONS ON SYSTEM SUITABILITY PARAMETERS - HPLC & GC

1) WHAT IS SYSTEM SUITABILITY TEST? EXPLAIN SST PARAMETERS IN HPLC or GC?

The system suitability test is used to verify that the chromatographic system is suitable for the intended analysis or not. That is to ensure that the complete testing system including instruments, electronics, reagents, column & analyst is suitable for intended application.

The main system suitability parameters are
1. Precision
2. Capacity factor
3. Selectivity factor
4. Resolution
5. Theoretical plate count
6. Tailing factor
7. Signal to Noise ratio
8. Peak to valley ratio

2) WHAT IS PRECISION? EXPLAIN?
It is the closeness with which results of replicate analysis of a sample agree. Usually expressed in terms of %RSD.
%RSD = Standard Deviation*100/Mean.

For Assay by HPLC, the maximum permitted relative standard deviation does not exceed the appropriate value given in the below table as per USP & EP.

Note:

1.B (percent) means upper limit of your assay -100.
That is If your assay limit is 98%-102%, B (percent) is 102%-100% = 2.0
That implies your %RSD should not more than 0.85% for 6 injections and 0.73 for 5 injections.

2.This requirement does not apply to tests for related substances.

3) WHAT IS CAPACITY FACTOR or RETENTION FACTOR?
It is the ratio of the adjusted retention volume (or time) to the hold-up or Void volume (or time). (or) It is the migration rate of analyte on a column (or) It is a measure of time of sample component resides in the stationary phase relative to the time it resides in the mobile phase.

Simply it is measure of where the peak of interest located with respect to void volume (retention volume of unretained compound).

Capacity factor (K¹) = VR¹/Vm = tR¹/ tm = (tR-tm)/tm

VR¹= Retention volume of analyte.
VM=Retention volume of unretained compound
tR=Retention time of analyte.
tm or to=Retention time of unretained compound

Note:
1. Generally the ideal valve of K¹ is in between 2 to 5. But the acceptable valve is in between 1 to 20 .

4) WHAT IS SELECTIVITY FACTOR or RELATIVE RETENTION? EXPLAIN?
The relative retention of two peaks in a column is known as selectivity factor. It is the ratio of adjusted retention time of a compound to that of another used as reference obtained under identical conditions.

Selectivity factor (a ) = K2/K1 = (tR2-tm)/ (tR1-tm)

tR2=Retention time of our analyte
tR1=Retention time of reference compound.

Note:
1. As per USP the selectivity factor should be always greater than 1.

5) WHAT IS RESOLUTION? EXPLAIN?
Resolution is the ratio of distance of separation of band maxima to their average base width. (or) The distance between the peak centers of a two analyte peaks divided by the average base width of the peaks.

“It is the ability of a chromatographic column to separate peaks. It is usually expressed in terms of the separation between two adjacent peaks”

As per USP:


Resolution(R) = 2 (tR2-tR1)/ (w1+w2)

tR2&tR1 are retention times of two components.

w1&w2 are corresponding peak widths at base.

As per EP:

Resolution(R) = 1.18 (tR2-tR1)/ (wh1+wh2)

tR2&tR1 are retention times of two components

wh1&wh2 are corresponding peak widths at half height.

Note:
1. Base line resolution achieved at R=1.5. But more than two is desirable.
6) WHAT IS THEORITICAL PLATE? HOW NUMBER OF THEORITICAL PLATES EFFECT THE COLUMN EFFICIENCY? EXPLAIN?

Martine & Synge used a chromatographic model involving a hypothetical division of column in to no. of plates known as theoretical plate. So, Theoretical plate is an imaginary part of the column.

The theoretical plate number (N) is a measure of the efficiency per unit length of the column.

As per USP:


Number of theoretical plates (N) = 16 (tR/W) 2

tR=Retention time of analyte

W=Peak width at base

As per EP:

Number of theoretical plates (N) = 5.54 (tR/Wh) 2

tR=Retention time of analyte

Wh =Peak width at half height.

7) WHAT IS SYMMETRY FACTOR or TAILING FACTOR? EXPLAIN?

1. Theory assumes an ideal symmetric peak which is known as Gaussian peak. The front side deviation from the Gaussian peak is known as peak fronting & rear side deviation is known as peak tailing.

2. It is a factor which describing shape of a chromatographic peak. And it is a measure of peak tailing.

Define Tailing factor:

“the distance from the front slope of the peak to the back slope divided by twice the distance from the center line of the peak to the front slope, with all measurements made at 5% of the maximum peak height”.
As per USP & EP:

Tailing factor (As) = w 0.05/2d

w 0.05=Width of peak height at one-twentieth of peak height

d=Distance between the perpendicular dropped from the peak maxima and the leading edge of the peak at one-twentieth of the peak height

Note:
1. As per EP, In related substance test or assay the symmetry factor should be in between 0.8 to 1.5 unless otherwise specified prescribed.


8) WHAT IS SIGNAL TO NOISE (S/N) RATIO? EXPLAIN?

The short term noise influences the precision of quantification. So S/N ratio is a useful system suitability parameter to identify noise effect on quantification of impurities..etc.

Signal to noise(S/N) ratio is calculated from following equation


S/N = 2H/h

Where

H= Height of concerned peak measured from the peak apex to the base extrapolated over a distance ≥5 times the peak width at its half height.

h= Difference between the largest and smallest noise valves observed over a distance ≥5 times width at half height of the peak.




9) WHAT IS PEAK TO VALLEY (p/v) RATIO EXPLAIN?

Peak to valley (p/v) ratio may be employed as system suitability criterion in a test for related substance when baseline separation between two peaks is not achieved.

p/v = Hp/Hv Where

Hp= Height above the extrapolated baseline of the minor peak.
Hv= Height above the extrapolated baseline at the lowest point of the curve separating the minor and major peaks.

QUALITY CONTROL INTERVIEW QUESTIONS ON CHROMATOGRAPHY

       1)WHAT IS THE PRINCIPLE OF CHROMATOGRAPHY (HPLC&GC)?

Separation of, mixture of components, Based on equilibrium distribution of analyte between two immiscible phases (like stationary phase and mobile phase).

HPLC: Solid stationary phase, Liquid mobile phase

GC     : Liquid stationary phase, Gas mobile phase.

       2) WHAT ARE REVERSE PHASE AND NORMAL PHASE CHROMATOGRAPHY?

These are the different type of techniques in chromatography.

Reverse phase chromatography:

Stationary phase: Non-polar

Mobile phase      : Polar

Normal phase chromatography:

Stationary phase: Polar

Mobile phase      : Non-polar

       3)IS CHIRAL CHROMATOGRAPHY & NORMAL PHASE CHROMATOGRAPHY ARE SAME or NOT?

No both are not 100% same.

Chiral chromatography:

             In this we determine the content of chiral isomer whether is it in normal phase or in reverse phase.

Normal phase chromatography:

             In this as above said we must use polar stationary phase and non-polar mobile phase whether is it to determine content of isomer or purity of analyte or whatever it may be.

       4) WHAT IS THE DIFFERENCE BETWEEN CHROMATOGRAPHIC PURITY & RELATED SUBSTANCE BY HPLC?

Chromatographic purity:

              Chromatographic purity is used when all the impurities are known in the given sample. Here we quantitate only known impurities.

Related substance:

               Related substance is used to quantitate both known as well as unknown impurities in the given sample.

       5)         WHAT ARE THE IMPURITY QUANTITATION METHODS IN HPLC?

There are mainly four types.

      1.      Area normalization method

      2.      Internal standard method

      3.      External standard method

      4.      Standard addition method

       6)  WHAT IS THE DIFFERENCE BETWEEN HPLC PURITY, ASSAY AND POTENCY?

Purity:

            It explains how pure our analyte is. It is not related to the amount of analyte present. Simply it is “100-% known or unknown impurity detected in HPLC”

Assay:

           It explains how much of our analyte is (i.e. content of our analyte). It is not related to purity of our analyte.

Potency:

           It explains how potent our analyte is (i.e. the original content of our analyte). It is calculated by mass balance technique. Simply it is 100-all possible impurities.

All possible impurities include Chromatographic impurities (HPLC, GC, and TLC), Heavy metals, Sulphated ash, MC, LOD etc.

Explanation:

            Let us assume an analyte of A with 99.5% purity, 100% assay and 99.0% potency. Then prepare 20%, 50% and 85% solution and analyze in HPLC.

For 20% solution you will get 99.5% purity, 20% assay and 99.0% potency.

For 50% solution you will get 99.5% purity, 50% assay and 99.0% potency.

For 85% solution you will get 99.5% purity, 85% assay and 99.0% potency.

Because we are diluting the concentration so purity will never change for dilution.  Potency is calculated from purity so it is also remains same.

       7)  WHAT IS A BUFFER SOLUTION? CAN YOU EXPLAIN ABOUT ACIDIC BUFFER & BASIC BUFFER?

Buffer solution:

          A solution which can resist change in its pH value on dilution or on the addition of solution of acid or base (Whole pH remains constant) is known as buffer solution.

Acidic buffer:

         Mixture of equimolar quantities of weak acid and salt of this weak acid with a strong base is known as acidic buffer.

Ex: Mixture of CH3COOH & CH3COONa

Basic buffer:

          Mixture of equimolar quantities of weak base and salt of this weak base with a strong acid is known as basic buffer.

Ex: Mixture of NH4OH & NH4Cl

       8)WHAT ARE PROTIC & APROTIC SOLVENTS?

Protic solvents:

              Any molecular solvent which contain dissociable H⁺ ion is called Protic solvent.

(The molecules of such solvents can donate an H⁺  proton.

Aprotic solvents:

              Any molecular solvent which does not contain dissociable H⁺ ion is called Aprotic solvent. (The molecules of such solvents cannot donate an H⁺  proton).

       9) WHAT IS DIFFERENCE BETWEEN HPLC & GC?

Principally both are same. Both are used for the separation of components present in the mixture of sample. Even though these two are principally same, these two also have some differences based applicability, Instrumentation.

Based on applicability:

·         HPLC is used for the separation of thermally stable (or non volatile) compounds.

·         GC is used for separation of volatile (thermally stable) compounds.

In nature 80% of the compounds are non volatile so we use HPLC more frequently than GC.

            Based on instrumentation:

·         In HPLC we use liquid mobile phase and solid stationary phase (Adsorption technique). And in HPLC we use small columns (more frequently up to 30 cm).

·         In GC we use gas mobile phase and liquid stationary phase (Partition technique). And in GC we use columns of length up to 105 cm.

Mobile phase, stationary phase, column, Detector...etc have to select based on sample properties in both the cases.

·         In HPLC we have the pumping system to pump the mobile phase with somewhat high pressure. But in GC we have no pumping system.

·         In HPLC we cannot change the column temperature with time. But we can change temperature with time in GC by giving oven program.

·         In HPLC we use detectors like UV, RI...etc where as in GC we use FID, ECD...etc.

10)  WHAT IS PHASE COLLAPSE? (or)

 CAN I WASH MY HPLC REVERSE PHASE COLUMN WITH 100% WATER?

Before going to this question we should know the C18 column structure. In HPLC C18 column the inner wall of the column is packed with silica (Si-o-Si), this silica is bonded to C18 linkages.

So from this we can clearly understand that our washing solvent will flow on to C18 linkages.

If we wash our Reverse phase C18 column with 100% water, Due to polar (water) and non polar (C18 linkage) repulsion stationary phase matrix collapsed. This collapse is known as PHASE COLLAPSE. So we should not wash our reverse phase column with 100% water.

11)  WHAT IS NEEDLE WASH IN HPLC? (or)

 WHAT IS THE BEST COMPOSITION FOR NEEDLE WASH?

           The needle wash is used to clean the injector needle before and after the injection. The design of needle in the injector system varies for different manufacturers.

In some designs the inside of the needle is part of the flow path of the mobile phase and thus it is flushed continuously by the mobile phase between injections. In this case it is outside (upper layer) of the needle which is cleaned by the needle wash. In some other designs needle is separate from mobile phase flow path and thus the needle wash is used to clean inside and outside of the needle.

         In all the cases composition of needle wash needs to be matched to the sample since this is what you want to clean off the needle. Therefore typically the composition of needle wash is that which matches the proportion of aqueous and organic solvents in the mobile phase will be appropriate.

12)  WHAT IS SEAL WASH IN HPLC? (or)

 WHAT IS THE BEST COMPOSITION FOR SEAL WASH?

Before going to this question first of all we should know about seal in HPLC. In HPLC pump system there is a piston which moves back and forward in the pump head which drawing in and pushing out the mobile phase with each movement. Since it is a moving part, a seal around this piston is required to prevent mobile phase leaking out of the back of the pump, over a period of time small amounts of mobile phase solvents leach through the seal to the back of the piston. If these solvents contain buffers then the salts may precipitate out forming deposits, which can shorten the life of the seal.

So seal wash is to flush the back of the piston seals to remove any deposits and maximizing the lifetime of the seal.

Composition:

It should be aqueous to dissolve buffer and a small amount of organic is required to prevent the bacteria growth and also to reduce the surface tension of water. Typically seal wash composition is 90% water and 10% organic solvent. The organic solvent may be methanol, acetonitrile and IPA…etc.

13)  IN REVERSE PHASE CHRMATOGRAPHY PEAKS ELUTE EARLIER WHEN WE INCRESE ACETONITRILE.WHY?

Most of people confused that, Water is more polar than acetonitrile so when we increase % acetonitrile peaks should elute late because in reverse phase chromatography more polar one comes out first and non-polar comes out last.

In any chromatography the elution order of analyte depends on eluting strength of solvent. In reverse phase chromatography, the less polar, the greater the eluting strength. So the % of ACN increases the eluting strength also increases so the peaks elute earlier. Reverse is true for normal phase chromatography.

Note:

            1.      In above case except %ACN, remaining all same in both the cases.

14)  WHAT ARE DETECTORS USED FOR GAS CHROMATOGRAPHIC ANALYSIS? NAME AT LEAST 5?

     1.      Flame ionization detector.(FID)

     2.      Electron capture detector.(ECD)

     3.      Thermal conductivity detector.(TCD)

     4.      Photo ionization detector.(PID)

     5.      Flame photometric detector.(FPD)

     6.      Nitrogen-Phosphorous detector.(NPD)

15)  GIVE ICH LIMITS OF FOLLOWING MENTIONED SOLVENTS IN % W/W?

      1.      1,4-Dioxane (Class 2) – 380 ppm

      2.      Cyclo hexane (Class 2)  – 3880 ppm

      3.      Pyridine (Class 2) – 200 ppm

      4.      Xylene (Class 2) – 2170 ppm