MICRONEEDLES: AN EFFECTIVE TECHNIQUE FOR TRANSDERMAL DRUG DELIVERY

Optimization of drug delivery through human skin is important in modern therapy. With limitations of oral drug delivery, pain and needle phobias associated with traditional injections drug delivery research has focused on transdermal delivery route. A new approach to transdermal delivery that acts as a bridge between the user friendless of patches and the broad effectiveness of hypodermic needles has recently received attention.by using needles of micron dimensions, termed microneedles, skin can be pierced to effectively deliver the drugs, The mechanism of action is based on temporary mechanical disruption of skin. The drug, in the form of biomolecules, is encapsulated within the micro needles, which are then inserted into the skin in the same way a drug like nitroglycerine is released into the bloodstream from a patch. The needles dissolve within minutes, releasing the trapped cargo at the intended delivery site. The present review focus on various studies related to micro needles for transdermal drug delivery and technology applications in various fields.


INTRODUCTION
More recently, considerable interest has arisen regarding Microporation technologies that create micronizedmicro channelsor micro poresin the skin by using technologies such as laser ablation, thermal or radiofrequency ablation, or mechanical microneedles. [1] Researchers have described these minimally invasive technologies as third-generation technologies that will have a significant impact on medicine [2] Microneedles, a microstructure transdermalsystem, consists of an array of micro structuredprojections coated with a drug or vaccine that isapplied to the skin to provide intradermal deliveryof active agents, which otherwise would notcross the stratum corneum. [3] They are generally one micron in diameter andrange from 1-100 microns in length.These have been fabricated with variousmaterials such as: metals, silicon, silicon dioxide,polymers, glass and other materials. The major advantage of microneedles overtraditional needles is, when it is inserted into theskin it does not pass the stratum corneum, whichis the outer 10-15 µm of the skin. Conventionalneedles which do pass this layer of skin mayeffectively transmit the drug but may lead toinfection and pain.
IJBR2 [8][2011]451-465 www.ijbr.ssjournals.com As for microneedles they canbe fabricated to be long enough to penetrate thestratum corneum, but short enough not topuncture nerve endings. Thus reduces thechances of pain, infection, or injury. [4] Various types of needles have beenfabricated as well, for example: solid (straight,bent, filtered), and hollow. Solid microneedlescould eventually be used with drug patches toincrease diffusion rates; solid-increasepermeability by poking holes in skin, rub drugover area, or coat needles with drug. Hollowneedles could eventually be used with drugpatches and timed pumps to deliver drugs atspecific times. Arrays of hollow needles could beused to continuously carry drugs into the bodyusing simple diffusion or a pump system. Hollowmicroneedles could also be used to remove fluidfrom the body for analysis -such as bloodglucose measurements -and to then supplymicroliter volumes of insulin or other drug asrequired. The hollow needle designs includetapered and beveled tips, and could eventuallybe used to deliver microliter quantities of drugsto very specific locations. Verysmall microneedles could provide highlytargeted drug administration to individual cells.These are capable of very accurate dosing,complex release patterns, local delivery andbiological drug stability enhancement bystoring in a micro volume that can be preciselycontrolled. [5]

ADVANTAGES OF MICRONEEDLES
The major advantage of microneedles over traditional needles is, when it is inserted into the skin it does not pass the stratum corneum, which is the outer 10-15 µm of the skin [6] . Conventional needles which do pass this layer of skin may effectively transmit the drug but may lead to infection and pain. As for microneedles they can be fabricated to be long enough to penetrate the stratum corneum, but short enough not to puncture nerve endings. Thus reduces the chances of pain, infection, or injury. In terms of processing there are also many advantages. By fabricating these needles on a silicon substrate because of their small size, thousands of needles can be fabricated on a single wafer. This leads to high accuracy, good reproducibility, and a moderate fabrication cost.

PROCESSING OF MICRONEEDLES
As stated previously, there are many types of materials, shapes and methods of processing of microneedles. Silicon microneedles are fabricated by using a wet etch method using a KOH solution, which requires a four step process [7] . The first step is todeposit a pad oxide (350 Å) and a nitride double layer (1000 Å) through LPCVD (low pressure chemical vapor deposition) on a (100) silicon wafer. By using photolithography, masking and plasma etching small micron sized circles are patterned on the silicon.  [6] . The sacrificial photoresist layer is then removed to release the electroplated/sputtered array, which is shown below in figure 1.

NEED FOR USING MICRONEEDLES
When oral administration ofdrugs is not feasible due to poor drug absorptionor enzymatic degradation in the gastrointestinaltract or liver, injection using a painful hypodermicneedle is the most common alternative. Anapproach that is more appealing to patients, andoffers the possibility of controlled release overtime, is drug delivery across the skin using apatch. However, transdermal delivery is severelylimited by the inability of the large majority ofdrugs to cross skin at therapeutic rates due tothe great barrier imposed by skin's outer stratumcorneum layer. [8] To increase skinpermeability, a number of different approacheshave been studied, ranging from chemical/lipidenhancers [9] to electric fields employingiontophoresis and electroporation to pressurewaves generated by ultrasound or photo acousticeffects. Although the mechanisms are alldifferent, these methods share the common goalto disrupt stratum corneum structure in order tocreate "holes" big enough for molecules topass through. The size of disruptionsgenerated by each of these methods isbelieved to be of nanometer dimensions, whichis large enough to permit transport of smalldrugs and, in some cases, macromolecules,but probably small enough to prevent causingdamage of clinical significance [10] An alternative approach involves creatinglarger transport pathways of micronsdimensions using arrays of microscopicneedles. These pathways are orders ofmagnitude bigger than molecular dimensionsand, therefore, should readily permit transportof macromolecules, as well as possiblysupramolecular complexes and micro particles.Despite their very large size relative to drugdimensions, on a clinical length scale theyremain small. Although safety studies need tobe performed, it is proposed that micron-scaleholes in the skin are likely to be safe, given thatthey are smaller than holes made byhypodermic needles or minor skin abrasionsencountered in daily life. [11] Transdermal drug delivery is a noninvasive,user-friendly delivery method fortherapeutics. However, its clinical use hasfound limited application due to the remarkablebarrier properties of the outermost layer ofskin, the stratum corneum (SC). Physical andchemical methods have been developed toovercome this barrier and enhance thetransdermal delivery of drugs. One of suchtechniques was the use of microneedles totemporarily compromise the skin barrier layer.This method combines the advantages ofconventional injection needles and transdermalpatches while minimizing their disadvantages.As compared to hypodermic needle injection,microneedles can provide a minimally invasivemeans of painless delivery of therapeuticmolecules through the skin barrier withprecision and convenience. The microneedlesseldom cause infection while they can allowdrugs or nanoparticles to permeate through theskin.
Increased micro needleassistedtransdermal delivery has been demonstratedfor a variety of compounds. For instance, the fluxof small compounds like calcein, diclofenacmethyl nicotinate was increased by micro needle arrays. In IJBR2 [8][2011]451-465 www.ijbr.ssjournals.com addition, microneedles also have beentested to increase the flux of permeation for largecompounds like fluorescein isothiocynate-labeledDextran, bovine serum albumin, insulin andplasmid DNA and nanospheres.Microneedles may create micro conduitssufficiently large to deliver drug-loadedliposomes into the skin. The combination ofelastic liposomes and microneedles may providehigher and more stable transdermal deliveryrates of drugs without the constraints oftraditional diffusion-based transdermal devices,such as molecular size and solubility. Though itcould offer benefits mentioned above, thecombined use of elastic liposomes andmicroneedles pretreatment has received littleattention.

MECHANISM OF ACTION
The mechanism for delivery is not based ondiffusion as it is in other transdermal drugdelivery products. Instead, it is based on thetemporary mechanical disruption of the skinand the placement of the drug or vaccine withinthe epidermis, where it can more readily reachits site of action.The drug, in the form of biomolecules, isencapsulated within the microneedles, whichare then inserted into the skin in the same waya drug like nitroglycerine is released into thebloodstream from a patch. The needlesdissolve within minutes, releasing the trappedcargo at the intended delivery site. They do notneed to be removed and no dangerous orbio hazardous substance is left behind on theskin, as the needles are made of abiodegradable substance.In microneedle devices, asmall area (the size of a traditional transdermalpatch) is covered by hundreds of microneedlesthat pierce only the stratum corneum (theuppermost 50 µm of the skin), thus allowing thedrug to bypass this important barrier ( Figure  1).The tiny needles are constructed in arrays todeliver sufficient amount of drug to the patientfor the desired therapeutic response . [12] 6. METHODOLOGY OF DRUG DELIVERY [13] A number of delivery strategies have beenemployed to use the microneedles fortransdermal drug delivery.These include • Poke with patch approach • Coat and poke approach • Biodegradable micro needles • Hollow microneedles • Dip and scrape Poke with patch approach: It involves piercing an array ofsolid micro needles into the skin followed byapplication of the drug patch at the treated site.Transport of drug across skin can occur bydiffusion or possibly by iontophoresis if an electricfield is applied. Coat and poke approach: In this approach needles arefirst coated with the drug and then inserted intothe skin for drug release by dissolution. The entiredrug to be delivered is coated on the needle itself. Biodegradable micro needles: It involves encapsulating thedrug within the biodegradable, polymericmicroneedles, followed by the insertion into theskin for a controlled drug release. Hollow microneedles: It involves injecting the drug through the needlewith a hollow bore. This approach is morereminiscent (suggestive of) of an injection than apatch. Dip and scrape: Dip and scrape approach, wheremicroneedles are first dipped into a drug solutionand then scraped across the skin surface to leavebehind the drug within the micro abrasions createdby the needles. The

PREPARATION
OF MICRONEEDLES [14] Molding: Micro molds were fabricated usingphotolithography and molding processes. Inbrief, a female micro needle master-mold wasstructured in SU-8 photoresist by UV exposureto create conical (circular cross section) orpyramidal (square cross section) microneedlestapering from a base measuring 300 µm to atip measuring 25 µm in width over amicro needle length of 600-800 µm. A malemicroneedle masterstructure made ofpolydimethylsiloxane was created using thismold. The PDMS master-structure was sputter coatedwith 100 nm of gold to preventadhesion with a second PDMS layer curedonto the male master-structure to create afemale PDMS replicate-mold. Excess PDMSon the female replicate-mold was trimmed sothat the mold fit within the 27-mm innerdiameter of a 50 ml conical tube. This metal coatedmale master-structure was repeatedlyused to make replicatemolds that wererepeatedly used to make microneedle devices. Other methods [12] Laser cutting:Microneedles were cut from stainless steelsheets using an infrared laser. The desiredmicroneedle shape and dimensions were firstdrafted in AutoCAD software. Using thisdesign, the infrared laser was operated at1000 Hz, 20 J/cm2 energy density and 40%attenuation of laser energy to cut themicroneedles. A total of three passes wererequired to completely cut through the stainlesssteel sheet. A cutting speed of 2 mm/s and airpurge at a constant pressure of 140 kPa wasused. Microneedles were either prepared asindividual rows of needles ('in-plane' needles)or as two-dimensional arrays of needles cutinto the plane of the stainless steel sheet andsubsequently bent at 90° out of the plane ('out of-plane' needles).

Cleaning and bending microneedles:
Laser-cut stainless steelmicroneedle arrays were manually cleaned withdetergent to de-grease the surface and removeslag and oxides deposited during laser cutting,which was followed by thorough rinsing inrunning water. To prepare 'out-of-plane'microneedles, microneedles cut into stainlesssteel sheets were first manually pushed out ofthe sheet using either forceps or a hypodermicneedle (26 gage, 1/2 inch long) while viewingunder a stereo microscope, and then bent at 90°angle with the aid of a #9 single-edged razorblade.
Electro polishing: To clean microneedle edges and to makethe tips sharp, microneedles wereelectro polished in a solution containing glycerin,orthophosphoric acid (85%) and water in a ratioof 6:3:1 by volume. Electro polishing wasperformed in a 300 ml glass beaker at 70 °Cand a stirring rate of 150 rpm. A copper platewas used as the cathode, while microneedlesacted as the anode. The anode was vibrated ata frequency of 10 Hz throughout theelectropolishing process using a custom-builtvibrating device to help remove gas bubblesgenerated at the anodic surface duringelectropolishing. A current density of1.8 mA/mm2 was applied for 15 min toelectropolish the microneedles. Afterelectropolishing, microneedles were cleanedby dipping alternately three times in de-ionizedwater and 25% nitric acid for 30 s each. Thiswas followed by another washing step in hotrunning water and a final wash in running deionizedwater. Due to the electropolishingprocess, the thickness of the microneedles wasreduced to 50 µm. Microneedles were driedusing compressed air before storing in airtightcontainers until later use.  4A). Thebottom plate had a central feeding channel(1 mm deep × 0.5 mm wide) machined into oneof its faces, with a through-hole drilled acrossto the other face. This hole acted as the inletport to fill the channel with the coating solution.The cover plate had five holes (400 µmdiameter) drilled into it at the same interval asthe microneedles in the in-plane row to becoated. These 'dip-holes' acted as individualdipping reservoirs to coat each of themicroneedles in the row. The two plates(bottom and cover plates) were aligned andadhered to each other using solvent bondingwith methylene chloride (Fisher Scientific) asthe solvent.

Micro
(2) Micro positioning dip coater: To enable three-dimensional alignment anddipping of microneedle rows into the dipholes,three linear-micropositioners wereassembled on a 6.35-mm thick, flat, acrylicplate (Fig. 4B). The first micropositioner wasused to control the position of the in-planemicroneedle row. The other twomicropositioners were assembled one on topof the other on the acrylic plate to create acomposite Y-Z motion micropositioner thatwas used to control the position of thecoating-solution reservoir. The threemicropositioners together allowed thealignment of the inplane microneedle row tothe dip-holes. The X-micropositioner wasused to horizontally dip the microneedles intoand out of the dip-holes. The coating wasperformed manually while viewing under astereo microscope. Control over the length ofthe microneedle shaft to be coated wasexercised manually using the Xmicropositioner.Tolerance for misalignmentwas included by designing the dip-holediameter to be twice the width of themicroneedles. The fabricated microneedles for local delivery can generally be categorized into in-plane and out-of plane microneedles.

Out-of-Plane Microneedles:
In the literature, microneedle devices can be categorized as either out-of-plane (where the fluid flow channel is normal to the substrate) or in-plane (where the fluid flow channel is parallel to the substrate) [15].
For out-of-plane devices, bulk micromachining and/or micromolding fabrication techniques are utilized. Therefore, microneedle shape is typically dictated by some sort of removal process, which can be costly in terms of machine time, controllability, and maximum depth producible (100 to 500 µm). Furthermore, fluid access to the microneedle lumens is typically through the backside of the substrate which can also complicate the fabrication process. Due to their nature, out-of-plane microneedles are micro machined in two-dimensional arrays, making them ideal for patch-like transdermal drug delivery applications. However, this geometry may also make it difficult to integrate planar microfluidic components on-chip. Material selection is also limited by the fabrication techniques used to manufacture out-of-plane microneedles. For example, traditional bulk micromachining has produced several different microneedle designs in single crystal silicon [16]. Using micromolding, out-of-plane devices have been fabricated in both polymers [17] and electrodeposited metals [18]. However, silicon is brittle, polymers lack the required stiffness to penetrate into viable epidermis strata, and electroplated metals such as nickel are not considered to be biocompatible (nickel is a well-known skin irritant and carcinogen) [19].   [13] .

In-Plane
Microneedles: In-plane microneedle devices, where the lumen is parallel to the substrate, have several advantages over out-of-plane designs. First, microneedle length and shape can be defined lithographically. Also, in-plane microfluidic components can be easily integrated. But traditionally, in-plane microneedles have relied heavily on surface micromachining techniques which limit deposited film thicknesses and therefore overall device strength. Also, inplane microneedle arrays are typically limited to one-dimension, which can limit fluid throughput considerably. Existing technologies have utilized a number of different materials (see Fig.3), including polysilicon [20], electrodeposited metals [21], and single crystal silicon [22] However, similar to those commonly used for out-of-plane microneedles, these micromechanical materials impose limitations on device performance which ultimately constrains their utility and efficacy. For example, the low fracture toughness of silicon and polysilicon can detrimentally affect device reliability and damage tolerance. And the materials associated with electrode position (e.g. nickel) may not be well-suited for microneedle applications. Consequently, there is a distinct need for the development of micromechanical

APPLICATIONS OF MICRONEEDLETECHNOLO GY
Microneedle technology has been developedas a platform technology for delivery of highmolecular weight and hydrophilic compoundsthrough the skin. The first ever study oftransdermal drug delivery by microarraytechnology was conducted by Henry et al whodemonstrated an increase in the permeability ofskin to a model compound calcein usingmicroarray technology. In a follow up study, Mc-Allister et al found a change in the permeabilityof cadaver skin to insulin, latex nanoparticlesand bovine serum albumin after treatment withmicroneedles, and unleashed the mechanism oftransport as simple diffusion. Oligonucleotide delivery: Lin and coworkers extended the in vitrofindings of microarray drug delivery to in vivoenvironment. An oligonucleotide, 20-merphosphorothioated oligodeoxynucleotide wasdelivered across the skin of hairless guinea pigeither alone or in combination with iontophoresis.Lin and coworkers used solid microneedlesetched from stainless steel or titanium sheetprepared with the poke with patch approach.This delivery system increased the absorption ofthe molecules relative to the intact skin.Iontophoresis combined with microneedles wasable to increase the transdermal flux by 100 foldcompared to the iontophoresis alone. DNA vaccine delivery [13] : The cells of Langerhans present in the skinserve as the first level of immune defense of thebody to the pathogens invading from theenvironment. These cells locate the antigensfrom the pathogens and present them toTlymphocytes, which in turn stimulate theproduction of antibodies. Mikszta et al reportedthe delivery of a DNA vaccine using microneedletechnology prepared with the dip and scrapeapproach. The arrays were dipped into asolution of DNA and scrapped multiple timesacross the skin of mice in vivo. Expression ofluciferase reporter gene was increased by2800 fold using micro enhancer arrays. Inaddition, microneedle delivery induced immuneresponses were stronger and less variablecompared to that induced by the hypodermicinjections. Similar results were obtained byresearchers at Beckett-Dickinson™ in ananimal study for antibody response to HepBnaked plasmid DNA vaccine. This approachhas a potential to lower the doses and thenumber of boosters needed for immunization. Solidmicroneedles of stainless steel having 1mmlength and tip width of 75 µm were inserted intothe rat skin and delivered insulin using pokewith patch approach. Over a period of 4 hours,blood glucose level steadily decreased by asmuch as 80% with the decrease in glucoselevel being dependent on the insulinconcentration.

Porphyrin
Precursor 5-Aminolevulinic Acid(ALA) Delivery: Photodynamic therapy of deep or nodularskin tumours is currently limited by the poortissue penetration of the porphyrin precursor 5-aminolevulinic acid (ALA). Ryan F. Donnelly andco workers have shown that, in vivoexperiments using nude mice showed thatmicroneedle puncture could reduce applicationtime and ALA dose required to induce high levelsof the photosensitizer protoporphyrin IX in skin.This clearly has implications for clinical practice,as shorter application times would meanimproved patient and clinician convenience andalso that more patients could be treated in thesame session. In vitro transdermal delivery of monoclonalantibody: In all the previously mentioned studies,purified human IgG was used as a model drugfor large proteins in transdermal delivery, andlater the feasibility of microneedlemediatedtransdermal delivery was further investigatedusing a human monoclonal antibody IgG todemonstrate the applicability of this techniquefor delivery of macromolecules.

COMMERCIAL MICRONEEDLETECHNOLO GIES
A decade after the first microneedles werereported, many commercial technologies havecome into the market including the Macrofluxtechnology, h-patch, Micro-Trans and manymore are given in table1 .

FUTURE TRENDS
Integration of solid microneedles withtransdermal patch provides a minimal invasivemethod to increase the skin permeability ofdrugs, including the macromolecules such asproteins. Till date, microneedles made up ofsilicon, metal, glass and plastics have beenutilized for transdermal delivery. [23]

FUTURE PROSPECTS
Early microneedles were made of single crystal silicon [24] The device wafer was sacrificed or dissolved away in silicon etchant leaving the microneedle behind. The fluid channels of these microneedles only occupy a small fraction of the interior volume of the needle resulting in a small fluid carrying capacity. These microneedles are useful for delivering fluid at low (50.1 ml/sec) flow rates but cannot deliver sufficient fluid for many therapeutic injections such as insulin. Micromolded needles leave the majority of the interior volume free, and allow larger fluid flow rates for the same size needle outer diameter. Other approaches to microneedles have been investigated. One approach uses SF6/O2 plasma to create high aspect ratio barbs for piercing the skin to allow therapeutics to diffuse across the skin [3] However, these needles are not sufficient for injecting large amounts of fluids or clinically relevant dosages of much therapeutics. Another approach uses electroplated palladium as the needle structural material and thick photoresist to define the needle channel [25] . However, because these needles are electroplated, they have very blunt tips. Future work needs to be performed to determine the biological response to needles. The first response to tissue distress from needle insertion is an inflammatory , polyethylene glycol (PEG) [28] , or plasma enhanced chemical vapor deposition (PECVD) of a Teflon-like fluropolymer. Any of these coatings could be incorporated into needle fabrication to improve biocompatibility. Since microneedles are designed for short term intradermal drug delivery, fibrous encapsulation is not expected because the needle is not inserted long enough for encapsulation to occur. Thrombosis is also not expected since the needle will not be in contact with the blood stream. Due to the small size of microneedles, strength and robustness are the major factors in determining the range of their applications. Needles must be able to tolerate forces associated with insertion, intact removal and normal human movements if they are to be integrated into portable biomedical devices.

CONCLUSION
Micro channel based Transdermal DeliverySystem by using Microneedles is a NovelApproach for Drug delivery system. It is aconvenient, painless, and less invasivealternative to injection & it can be used acommon method for administering largeproteins and peptides, antibiotics, vaccines inlow manufacturing cost. In contrast to oraldelivery, microneedles avoid first pass effectand offer the benefit of immediate cessation ofdrug administration in case of an adverseeffect or overdose. In contrast to passivedelivery, this allow for the delivery of water-solubledrugs. In contrast to Iontophoresis, thisis use for long time. There is also no molecularsize limitation, no molecular electrical chargerequirement, and no specific formulation pHconstraint. In contrast to conventional TDDS,this is using for potent & less potent the drug, themore extended release the delivery system.  Micro Needle Therapy System Clinical resolution lab Microneedle Dermaroller   [13] .